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rap1gap rabbit polyclonal antibody 19174 1 ap proteintech group  (Proteintech)


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    Proteintech rap1gap rabbit polyclonal antibody 19174 1 ap proteintech group
    Fig. 5. Rap1 overexpression reverses the expression of key proteins in CD147 knockdown mediated cell proliferation, apoptosis and EMT. (A, B) Western blot analysis of Rap1 and <t>Rap1GAP</t> protein levels in HCT116 and SW620 cells after CD147 knockdown (shCD147). β-actin served as a loading control. Data represent mean ± SD (n = 3; **P < 0.01, ***P < 0.001 vs. shNC, Student’s t-test). Original blots are presented in Supplementary Fig. 3. (C) qRT-PCR validation of Rap1 mRNA overexpression in CD147-knockdown cells. Expression normalized to β-actin (n = 3; ***P < 0.001 vs. HCT116 or SW620, Student’s t-test). (D, E) Western blot analysis of c-Myc, Bcl-2, Bax, N-cadherin, and E-cadherin protein levels in CD147-Knockdown cells with Rap1 overexpression (n = 3; *P < 0.05, **P < 0.01, ***P < 0.001 vs. shNC, #P < 0.05, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). Original blots are presented in Supplementary Fig. 4.
    Rap1gap Rabbit Polyclonal Antibody 19174 1 Ap Proteintech Group, supplied by Proteintech, used in various techniques. Bioz Stars score: 93/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/rabbit+polyclonal+proteintech+group/RAP1GAP+Antibody/pm40254691-88-47-52
    Average 93 stars, based on 4 article reviews
    rap1gap rabbit polyclonal antibody 19174 1 ap proteintech group - by Bioz Stars, 2026-10
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    1) Product Images from "CD147 regulates the Rap1 signaling pathway to promote proliferation, migration, and invasion, and inhibit apoptosis in colorectal cancer cells."

    Article Title: CD147 regulates the Rap1 signaling pathway to promote proliferation, migration, and invasion, and inhibit apoptosis in colorectal cancer cells.

    Journal: Scientific reports

    doi: 10.1038/s41598-025-98266-8

    Fig. 5. Rap1 overexpression reverses the expression of key proteins in CD147 knockdown mediated cell proliferation, apoptosis and EMT. (A, B) Western blot analysis of Rap1 and Rap1GAP protein levels in HCT116 and SW620 cells after CD147 knockdown (shCD147). β-actin served as a loading control. Data represent mean ± SD (n = 3; **P < 0.01, ***P < 0.001 vs. shNC, Student’s t-test). Original blots are presented in Supplementary Fig. 3. (C) qRT-PCR validation of Rap1 mRNA overexpression in CD147-knockdown cells. Expression normalized to β-actin (n = 3; ***P < 0.001 vs. HCT116 or SW620, Student’s t-test). (D, E) Western blot analysis of c-Myc, Bcl-2, Bax, N-cadherin, and E-cadherin protein levels in CD147-Knockdown cells with Rap1 overexpression (n = 3; *P < 0.05, **P < 0.01, ***P < 0.001 vs. shNC, #P < 0.05, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). Original blots are presented in Supplementary Fig. 4.
    Figure Legend Snippet: Fig. 5. Rap1 overexpression reverses the expression of key proteins in CD147 knockdown mediated cell proliferation, apoptosis and EMT. (A, B) Western blot analysis of Rap1 and Rap1GAP protein levels in HCT116 and SW620 cells after CD147 knockdown (shCD147). β-actin served as a loading control. Data represent mean ± SD (n = 3; **P < 0.01, ***P < 0.001 vs. shNC, Student’s t-test). Original blots are presented in Supplementary Fig. 3. (C) qRT-PCR validation of Rap1 mRNA overexpression in CD147-knockdown cells. Expression normalized to β-actin (n = 3; ***P < 0.001 vs. HCT116 or SW620, Student’s t-test). (D, E) Western blot analysis of c-Myc, Bcl-2, Bax, N-cadherin, and E-cadherin protein levels in CD147-Knockdown cells with Rap1 overexpression (n = 3; *P < 0.05, **P < 0.01, ***P < 0.001 vs. shNC, #P < 0.05, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). Original blots are presented in Supplementary Fig. 4.

    Techniques Used: Over Expression, Expressing, Knockdown, Western Blot, Control, Quantitative RT-PCR, Biomarker Discovery

    Fig. 7. Rap1 restores CD147-mediated migration and invasion in colorectal cancer cells. (A, C) Scratch wound healing assay in HCT116 cells. Healing rates were quantified at 24 h and 48 h (n = 3; ***P < 0.001 vs. shNC, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). (B, D) Transwell migration and Matrigel invasion assays. Migrated/invaded cells were counted and normalized to shCtrl (n = 3; ***P < 0.001 vs. shNC, ###P < 0.001 vs. shCD147, one-way ANOVA). SW620 cells showed consistent trends. (E) Mechanistic diagram of CD147 promoting tumor progression through Rap1/Rap1GAP signaling in colorectal cancer cells.
    Figure Legend Snippet: Fig. 7. Rap1 restores CD147-mediated migration and invasion in colorectal cancer cells. (A, C) Scratch wound healing assay in HCT116 cells. Healing rates were quantified at 24 h and 48 h (n = 3; ***P < 0.001 vs. shNC, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). (B, D) Transwell migration and Matrigel invasion assays. Migrated/invaded cells were counted and normalized to shCtrl (n = 3; ***P < 0.001 vs. shNC, ###P < 0.001 vs. shCD147, one-way ANOVA). SW620 cells showed consistent trends. (E) Mechanistic diagram of CD147 promoting tumor progression through Rap1/Rap1GAP signaling in colorectal cancer cells.

    Techniques Used: Migration, Wound Healing Assay

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    Article Snippet: .. Table S4 Primer sequence used in this study Gene sequence β-actin-F 5′ TCAAGATCATTGCTCCTCCTGA 3′ β-actin-R UPK1A-F UPK1A-R HIF1A-F HIF1A-R GLUT1-F GLUT1-R 5′ CTCGTCATACTCCTGCTTGCTG 3′ 5′ GAGGCTGCAGACAGAGAAGG 3′ 5′ TGAAGAAGGAGAAGCCGCAG 3′ 5′ TCTGGATGCTGGTGATTTGG 3′ 5′ GCACCAAGCAGGTCATAGGT 3′ 5′ CGGGCCAAGAGTGTGCTAAA 3′ 5′ TGACGATACCGGAGCCAATG 3′ HK2-F 5′ AACAGCCTGGACGAGAGCAT 3′ HK2-R 5′ GTCCACGCTTGGTGAAATCG 3′ LDHA-F 5′ ATCTTGACCTACGTGGCTTGGA 3′ LDHA-R 5′ CCATACAGGCACACTGGAATCTC 3′ PDK1-F 5′ ACCAGGACAGCCAATACAAG 3′ PDK1-R 5′ CCTCGGTCACTCATCTTCAC 3′ PKM2-F 5′ TCCGACACAGTCTCCTGGAC 3′ PKM2-R 5′ AAGCTTCCCCAGCTGTTCTC 3′ G6P1-F HRE1-F HRE1-R HRE2-F HRE2-R HRE3-F HRE3-R LDHA-HRE-F LDHA-HRE-R 5′ GCATTCCTGTGA CTGAGCAG 3′ 5’GTAGAGATAGGGTTTCACCATGTTAG 3’ 5’ CACTGGTTTGGTAATAAGATACATG 3’ 5’CTGACCTAAGGTGATCCCCCC 3’ 5’CACTGAAAATCCCATCGTC 3’ 5’GATGATACCCATTTCGCAGATGGGC 3’ 5’GGAGAAGAACGTGCTAGAAAATGAG 3’ 5’TTGGAGGGCAGCACCTTACTTAGA 3’ 5’GCCTTAAGTGGAACAGCTATGCTGAC 3’ Table S5 siRNA sequence used in this study siRNA sense sequence anti-sense sequence si-UPK1A-1 5’GCGUCAUGAUUGAG CAAGATT 3’ 5’UCUUGCUCAAUCAUGA CGCTT 3’ si-UPK1A-2 5’CACAUGGACUACCU GUUCATT 3’ 5’UGAACAGGUAGUCC AUGUGTT 3’ si-HIF-1α 5’GGACACAGAUUUAG ACUUGTT 3’ 5’CAAGUCUAAAUCUG UGUCCTT 3’ Table S6 Information on antibodies used in this study Antibody WB IHC IF Specificity Company UPK1A (ab185970) 1: 1000 / / Rabbit polyclonal Abcam UPK1A (sc-366947) / 1:100 1:100 Rabbit polyclonal Santa Cruz Biotechnology β-actin (sc-8432) 1:2000 / / Mouse monoclonal Santa Cruz Biotechnology HIF-1α (20960-1-AP) 1:1000 / / Rabbit polyclonal Proteintech Group ..

    Western Blot:

    Article Title: Reciprocal regulation of HIF-1α and Uroplakin 1A promotes glycolysis and proliferation in Hepatocellular Carcinoma
    Article Snippet: .. Table S4 Primer sequence used in this study Gene sequence β-actin-F 5′ TCAAGATCATTGCTCCTCCTGA 3′ β-actin-R UPK1A-F UPK1A-R HIF1A-F HIF1A-R GLUT1-F GLUT1-R 5′ CTCGTCATACTCCTGCTTGCTG 3′ 5′ GAGGCTGCAGACAGAGAAGG 3′ 5′ TGAAGAAGGAGAAGCCGCAG 3′ 5′ TCTGGATGCTGGTGATTTGG 3′ 5′ GCACCAAGCAGGTCATAGGT 3′ 5′ CGGGCCAAGAGTGTGCTAAA 3′ 5′ TGACGATACCGGAGCCAATG 3′ HK2-F 5′ AACAGCCTGGACGAGAGCAT 3′ HK2-R 5′ GTCCACGCTTGGTGAAATCG 3′ LDHA-F 5′ ATCTTGACCTACGTGGCTTGGA 3′ LDHA-R 5′ CCATACAGGCACACTGGAATCTC 3′ PDK1-F 5′ ACCAGGACAGCCAATACAAG 3′ PDK1-R 5′ CCTCGGTCACTCATCTTCAC 3′ PKM2-F 5′ TCCGACACAGTCTCCTGGAC 3′ PKM2-R 5′ AAGCTTCCCCAGCTGTTCTC 3′ G6P1-F HRE1-F HRE1-R HRE2-F HRE2-R HRE3-F HRE3-R LDHA-HRE-F LDHA-HRE-R 5′ GCATTCCTGTGA CTGAGCAG 3′ 5’GTAGAGATAGGGTTTCACCATGTTAG 3’ 5’ CACTGGTTTGGTAATAAGATACATG 3’ 5’CTGACCTAAGGTGATCCCCCC 3’ 5’CACTGAAAATCCCATCGTC 3’ 5’GATGATACCCATTTCGCAGATGGGC 3’ 5’GGAGAAGAACGTGCTAGAAAATGAG 3’ 5’TTGGAGGGCAGCACCTTACTTAGA 3’ 5’GCCTTAAGTGGAACAGCTATGCTGAC 3’ Table S5 siRNA sequence used in this study siRNA sense sequence anti-sense sequence si-UPK1A-1 5’GCGUCAUGAUUGAG CAAGATT 3’ 5’UCUUGCUCAAUCAUGA CGCTT 3’ si-UPK1A-2 5’CACAUGGACUACCU GUUCATT 3’ 5’UGAACAGGUAGUCC AUGUGTT 3’ si-HIF-1α 5’GGACACAGAUUUAG ACUUGTT 3’ 5’CAAGUCUAAAUCUG UGUCCTT 3’ Table S6 Information on antibodies used in this study Antibody WB IHC IF Specificity Company UPK1A (ab185970) 1: 1000 / / Rabbit polyclonal Abcam UPK1A (sc-366947) / 1:100 1:100 Rabbit polyclonal Santa Cruz Biotechnology β-actin (sc-8432) 1:2000 / / Mouse monoclonal Santa Cruz Biotechnology HIF-1α (20960-1-AP) 1:1000 / / Rabbit polyclonal Proteintech Group ..

    Article Title: Expression Status And Prognostic Value Of M6A-associated Genes in Gastric Cancer
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    Immunohistochemistry:

    Article Title: Reciprocal regulation of HIF-1α and Uroplakin 1A promotes glycolysis and proliferation in Hepatocellular Carcinoma
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    Figure 5. <t>HORMAD1</t> and HORMAD2 accumulate on the lateral elements of synapsed autosomes in Trip13-
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    Figure 1. <t>TRIP13</t> localizes to the synaptonemal complex and telomeres in spermatocytes. (A) Western blot analysis of TRIP13 in adult mouse tissues. Heart and skeletal muscle lack ACTB. (B) Immunofluorescence of TRIP13 in sections of 3-month-old wild type and Trip13-/- testes. Lep, leptotene; Zyg, zygotene; Pac, pachytene; eS, elongating spermatids; ES, elongated spermatids. (C) Immunofluorescence of TRIP13 in spread nuclei of spermatocytes from wild type P20 testes. (D) Super-resolution localization of TRIP13 to the central element (CE) but not lateral element (LE) of the synaptonemal complex at early pachytene stage. The enlarged view of the boxed chromosome is shown at the bottom.
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    Figure 1. <t>TRIP13</t> localizes to the synaptonemal complex and telomeres in spermatocytes. (A) Western blot analysis of TRIP13 in adult mouse tissues. Heart and skeletal muscle lack ACTB. (B) Immunofluorescence of TRIP13 in sections of 3-month-old wild type and Trip13-/- testes. Lep, leptotene; Zyg, zygotene; Pac, pachytene; eS, elongating spermatids; ES, elongated spermatids. (C) Immunofluorescence of TRIP13 in spread nuclei of spermatocytes from wild type P20 testes. (D) Super-resolution localization of TRIP13 to the central element (CE) but not lateral element (LE) of the synaptonemal complex at early pachytene stage. The enlarged view of the boxed chromosome is shown at the bottom.
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    Fig. 5. Rap1 overexpression reverses the expression of key proteins in CD147 knockdown mediated cell proliferation, apoptosis and EMT. (A, B) Western blot analysis of Rap1 and Rap1GAP protein levels in HCT116 and SW620 cells after CD147 knockdown (shCD147). β-actin served as a loading control. Data represent mean ± SD (n = 3; **P < 0.01, ***P < 0.001 vs. shNC, Student’s t-test). Original blots are presented in Supplementary Fig. 3. (C) qRT-PCR validation of Rap1 mRNA overexpression in CD147-knockdown cells. Expression normalized to β-actin (n = 3; ***P < 0.001 vs. HCT116 or SW620, Student’s t-test). (D, E) Western blot analysis of c-Myc, Bcl-2, Bax, N-cadherin, and E-cadherin protein levels in CD147-Knockdown cells with Rap1 overexpression (n = 3; *P < 0.05, **P < 0.01, ***P < 0.001 vs. shNC, #P < 0.05, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). Original blots are presented in Supplementary Fig. 4.

    Journal: Scientific reports

    Article Title: CD147 regulates the Rap1 signaling pathway to promote proliferation, migration, and invasion, and inhibit apoptosis in colorectal cancer cells.

    doi: 10.1038/s41598-025-98266-8

    Figure Lengend Snippet: Fig. 5. Rap1 overexpression reverses the expression of key proteins in CD147 knockdown mediated cell proliferation, apoptosis and EMT. (A, B) Western blot analysis of Rap1 and Rap1GAP protein levels in HCT116 and SW620 cells after CD147 knockdown (shCD147). β-actin served as a loading control. Data represent mean ± SD (n = 3; **P < 0.01, ***P < 0.001 vs. shNC, Student’s t-test). Original blots are presented in Supplementary Fig. 3. (C) qRT-PCR validation of Rap1 mRNA overexpression in CD147-knockdown cells. Expression normalized to β-actin (n = 3; ***P < 0.001 vs. HCT116 or SW620, Student’s t-test). (D, E) Western blot analysis of c-Myc, Bcl-2, Bax, N-cadherin, and E-cadherin protein levels in CD147-Knockdown cells with Rap1 overexpression (n = 3; *P < 0.05, **P < 0.01, ***P < 0.001 vs. shNC, #P < 0.05, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). Original blots are presented in Supplementary Fig. 4.

    Article Snippet: Supplier Antibody dilution used WB/IF Anti CD147 mouse monoclonal antibody 66443-1-AP ProteinTech Group, Inc. 1:5000 /1:200 Anti N-cadherin rabbit polyclonal antibody 22018-1-AP ProteinTech Group, Inc. 1:5000 Anti E-cadherin rabbit polyclonal antibody 20874-1-AP ProteinTech Group, Inc. 1:10000 Anti c-myc mouse monoclonal antibody 67,447-AP ProteinTech Group, Inc. 1:5000 Anti RAP1GAP rabbit polyclonal antibody 19174-1-AP ProteinTech Group, Inc. 1:2000 Anti RAP1 rabbit polyclonal antibody 14595-1-AP ProteinTech Group, Inc. 1:1000 Anti β-actin rabbit polyclonal antibody 20536-1-AP ProteinTech Group, Inc. 1:8000 HRP-conjugated Goat Anti-Mouse IgG(H + L) SA00001-1 ProteinTech Group, Inc. 1:10000 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) ab150116 Abcam 1:200 Table 1.

    Techniques: Over Expression, Expressing, Knockdown, Western Blot, Control, Quantitative RT-PCR, Biomarker Discovery

    Fig. 7. Rap1 restores CD147-mediated migration and invasion in colorectal cancer cells. (A, C) Scratch wound healing assay in HCT116 cells. Healing rates were quantified at 24 h and 48 h (n = 3; ***P < 0.001 vs. shNC, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). (B, D) Transwell migration and Matrigel invasion assays. Migrated/invaded cells were counted and normalized to shCtrl (n = 3; ***P < 0.001 vs. shNC, ###P < 0.001 vs. shCD147, one-way ANOVA). SW620 cells showed consistent trends. (E) Mechanistic diagram of CD147 promoting tumor progression through Rap1/Rap1GAP signaling in colorectal cancer cells.

    Journal: Scientific reports

    Article Title: CD147 regulates the Rap1 signaling pathway to promote proliferation, migration, and invasion, and inhibit apoptosis in colorectal cancer cells.

    doi: 10.1038/s41598-025-98266-8

    Figure Lengend Snippet: Fig. 7. Rap1 restores CD147-mediated migration and invasion in colorectal cancer cells. (A, C) Scratch wound healing assay in HCT116 cells. Healing rates were quantified at 24 h and 48 h (n = 3; ***P < 0.001 vs. shNC, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). (B, D) Transwell migration and Matrigel invasion assays. Migrated/invaded cells were counted and normalized to shCtrl (n = 3; ***P < 0.001 vs. shNC, ###P < 0.001 vs. shCD147, one-way ANOVA). SW620 cells showed consistent trends. (E) Mechanistic diagram of CD147 promoting tumor progression through Rap1/Rap1GAP signaling in colorectal cancer cells.

    Article Snippet: Supplier Antibody dilution used WB/IF Anti CD147 mouse monoclonal antibody 66443-1-AP ProteinTech Group, Inc. 1:5000 /1:200 Anti N-cadherin rabbit polyclonal antibody 22018-1-AP ProteinTech Group, Inc. 1:5000 Anti E-cadherin rabbit polyclonal antibody 20874-1-AP ProteinTech Group, Inc. 1:10000 Anti c-myc mouse monoclonal antibody 67,447-AP ProteinTech Group, Inc. 1:5000 Anti RAP1GAP rabbit polyclonal antibody 19174-1-AP ProteinTech Group, Inc. 1:2000 Anti RAP1 rabbit polyclonal antibody 14595-1-AP ProteinTech Group, Inc. 1:1000 Anti β-actin rabbit polyclonal antibody 20536-1-AP ProteinTech Group, Inc. 1:8000 HRP-conjugated Goat Anti-Mouse IgG(H + L) SA00001-1 ProteinTech Group, Inc. 1:10000 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) ab150116 Abcam 1:200 Table 1.

    Techniques: Migration, Wound Healing Assay

    Fig. 1. CD147 overexpression correlates with poor prognosis and RAP1 co-expression in colorectal cancer. (A, B) CD147 mRNA expression levels in tumor vs. normal tissues across multiple cancers (UCSC Xena database) and specifically in colorectal cancer (CRC) (GEPIA database). **P < 0.01, unpaired t-test. (C) Kaplan-Meier survival analysis of CRC patients stratified by CD147 expression (log-rank test, P < 0.05). (D) Representative immunohistochemical images from The Human Protein Atlas showing CD147 expression in normal colorectal tissue (negative) and CRC tissue (cytoplasmic/membrane staining). (E, F) Western blot analysis of CD147 protein levels in paired CRC and adjacent normal tissues (n = 12). β-actin served as loading control. ***P < 0.001, paired t-test. Origin blots are presented in Supplementary Fig. 1. (G, H) Correlation analyses between CD147 and RAP1A/B mRNA expression using GEPIA (Pearson’s correlation, P < 0.05) and TIMER2.0 (no significant correlation). (I) qRT-PCR validation of CD147 and Rap1 expression correlation in 12 CRC tissues (Pearson’s correlation, P < 0.01).

    Journal: Scientific reports

    Article Title: CD147 regulates the Rap1 signaling pathway to promote proliferation, migration, and invasion, and inhibit apoptosis in colorectal cancer cells.

    doi: 10.1038/s41598-025-98266-8

    Figure Lengend Snippet: Fig. 1. CD147 overexpression correlates with poor prognosis and RAP1 co-expression in colorectal cancer. (A, B) CD147 mRNA expression levels in tumor vs. normal tissues across multiple cancers (UCSC Xena database) and specifically in colorectal cancer (CRC) (GEPIA database). **P < 0.01, unpaired t-test. (C) Kaplan-Meier survival analysis of CRC patients stratified by CD147 expression (log-rank test, P < 0.05). (D) Representative immunohistochemical images from The Human Protein Atlas showing CD147 expression in normal colorectal tissue (negative) and CRC tissue (cytoplasmic/membrane staining). (E, F) Western blot analysis of CD147 protein levels in paired CRC and adjacent normal tissues (n = 12). β-actin served as loading control. ***P < 0.001, paired t-test. Origin blots are presented in Supplementary Fig. 1. (G, H) Correlation analyses between CD147 and RAP1A/B mRNA expression using GEPIA (Pearson’s correlation, P < 0.05) and TIMER2.0 (no significant correlation). (I) qRT-PCR validation of CD147 and Rap1 expression correlation in 12 CRC tissues (Pearson’s correlation, P < 0.01).

    Article Snippet: Supplier Antibody dilution used WB/IF Anti CD147 mouse monoclonal antibody 66443-1-AP ProteinTech Group, Inc. 1:5000 /1:200 Anti N-cadherin rabbit polyclonal antibody 22018-1-AP ProteinTech Group, Inc. 1:5000 Anti E-cadherin rabbit polyclonal antibody 20874-1-AP ProteinTech Group, Inc. 1:10000 Anti c-myc mouse monoclonal antibody 67,447-AP ProteinTech Group, Inc. 1:5000 Anti RAP1GAP rabbit polyclonal antibody 19174-1-AP ProteinTech Group, Inc. 1:2000 Anti RAP1 rabbit polyclonal antibody 14595-1-AP ProteinTech Group, Inc. 1:1000 Anti β-actin rabbit polyclonal antibody 20536-1-AP ProteinTech Group, Inc. 1:8000 HRP-conjugated Goat Anti-Mouse IgG(H + L) SA00001-1 ProteinTech Group, Inc. 1:10000 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) ab150116 Abcam 1:200 Table 1.

    Techniques: Over Expression, Expressing, Immunohistochemical staining, Membrane, Staining, Western Blot, Control, Quantitative RT-PCR, Biomarker Discovery

    Fig. 5. Rap1 overexpression reverses the expression of key proteins in CD147 knockdown mediated cell proliferation, apoptosis and EMT. (A, B) Western blot analysis of Rap1 and Rap1GAP protein levels in HCT116 and SW620 cells after CD147 knockdown (shCD147). β-actin served as a loading control. Data represent mean ± SD (n = 3; **P < 0.01, ***P < 0.001 vs. shNC, Student’s t-test). Original blots are presented in Supplementary Fig. 3. (C) qRT-PCR validation of Rap1 mRNA overexpression in CD147-knockdown cells. Expression normalized to β-actin (n = 3; ***P < 0.001 vs. HCT116 or SW620, Student’s t-test). (D, E) Western blot analysis of c-Myc, Bcl-2, Bax, N-cadherin, and E-cadherin protein levels in CD147-Knockdown cells with Rap1 overexpression (n = 3; *P < 0.05, **P < 0.01, ***P < 0.001 vs. shNC, #P < 0.05, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). Original blots are presented in Supplementary Fig. 4.

    Journal: Scientific reports

    Article Title: CD147 regulates the Rap1 signaling pathway to promote proliferation, migration, and invasion, and inhibit apoptosis in colorectal cancer cells.

    doi: 10.1038/s41598-025-98266-8

    Figure Lengend Snippet: Fig. 5. Rap1 overexpression reverses the expression of key proteins in CD147 knockdown mediated cell proliferation, apoptosis and EMT. (A, B) Western blot analysis of Rap1 and Rap1GAP protein levels in HCT116 and SW620 cells after CD147 knockdown (shCD147). β-actin served as a loading control. Data represent mean ± SD (n = 3; **P < 0.01, ***P < 0.001 vs. shNC, Student’s t-test). Original blots are presented in Supplementary Fig. 3. (C) qRT-PCR validation of Rap1 mRNA overexpression in CD147-knockdown cells. Expression normalized to β-actin (n = 3; ***P < 0.001 vs. HCT116 or SW620, Student’s t-test). (D, E) Western blot analysis of c-Myc, Bcl-2, Bax, N-cadherin, and E-cadherin protein levels in CD147-Knockdown cells with Rap1 overexpression (n = 3; *P < 0.05, **P < 0.01, ***P < 0.001 vs. shNC, #P < 0.05, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). Original blots are presented in Supplementary Fig. 4.

    Article Snippet: Supplier Antibody dilution used WB/IF Anti CD147 mouse monoclonal antibody 66443-1-AP ProteinTech Group, Inc. 1:5000 /1:200 Anti N-cadherin rabbit polyclonal antibody 22018-1-AP ProteinTech Group, Inc. 1:5000 Anti E-cadherin rabbit polyclonal antibody 20874-1-AP ProteinTech Group, Inc. 1:10000 Anti c-myc mouse monoclonal antibody 67,447-AP ProteinTech Group, Inc. 1:5000 Anti RAP1GAP rabbit polyclonal antibody 19174-1-AP ProteinTech Group, Inc. 1:2000 Anti RAP1 rabbit polyclonal antibody 14595-1-AP ProteinTech Group, Inc. 1:1000 Anti β-actin rabbit polyclonal antibody 20536-1-AP ProteinTech Group, Inc. 1:8000 HRP-conjugated Goat Anti-Mouse IgG(H + L) SA00001-1 ProteinTech Group, Inc. 1:10000 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) ab150116 Abcam 1:200 Table 1.

    Techniques: Over Expression, Expressing, Knockdown, Western Blot, Control, Quantitative RT-PCR, Biomarker Discovery

    Fig. 6. Rap1 overexpression rescues the regulatory effects of CD147 knockdown on proliferation and apoptosis. (A) CCK-8 assay showing proliferation of HCT116 (n = 3; ***P < 0.001 vs. shNC, ###P < 0.001 vs. shCD147, one-way ANOVA). (B, C) Colony formation assays in HCT116 cells. Colonies were counted and normalized to shCtrl (n = 3; **P < 0.01, ***P < 0.001 vs. shNC, ###P < 0.001 vs. shCD147, one-way ANOVA). (D, E) Flow cytometry analysis of apoptosis in HCT116 cells. Percentages of apoptotic cells are shown (n = 3; *P < 0.05 vs. shNC, #P < 0.05 vs. shCD147, one-way ANOVA). Similar results were observed in SW620 cells.

    Journal: Scientific reports

    Article Title: CD147 regulates the Rap1 signaling pathway to promote proliferation, migration, and invasion, and inhibit apoptosis in colorectal cancer cells.

    doi: 10.1038/s41598-025-98266-8

    Figure Lengend Snippet: Fig. 6. Rap1 overexpression rescues the regulatory effects of CD147 knockdown on proliferation and apoptosis. (A) CCK-8 assay showing proliferation of HCT116 (n = 3; ***P < 0.001 vs. shNC, ###P < 0.001 vs. shCD147, one-way ANOVA). (B, C) Colony formation assays in HCT116 cells. Colonies were counted and normalized to shCtrl (n = 3; **P < 0.01, ***P < 0.001 vs. shNC, ###P < 0.001 vs. shCD147, one-way ANOVA). (D, E) Flow cytometry analysis of apoptosis in HCT116 cells. Percentages of apoptotic cells are shown (n = 3; *P < 0.05 vs. shNC, #P < 0.05 vs. shCD147, one-way ANOVA). Similar results were observed in SW620 cells.

    Article Snippet: Supplier Antibody dilution used WB/IF Anti CD147 mouse monoclonal antibody 66443-1-AP ProteinTech Group, Inc. 1:5000 /1:200 Anti N-cadherin rabbit polyclonal antibody 22018-1-AP ProteinTech Group, Inc. 1:5000 Anti E-cadherin rabbit polyclonal antibody 20874-1-AP ProteinTech Group, Inc. 1:10000 Anti c-myc mouse monoclonal antibody 67,447-AP ProteinTech Group, Inc. 1:5000 Anti RAP1GAP rabbit polyclonal antibody 19174-1-AP ProteinTech Group, Inc. 1:2000 Anti RAP1 rabbit polyclonal antibody 14595-1-AP ProteinTech Group, Inc. 1:1000 Anti β-actin rabbit polyclonal antibody 20536-1-AP ProteinTech Group, Inc. 1:8000 HRP-conjugated Goat Anti-Mouse IgG(H + L) SA00001-1 ProteinTech Group, Inc. 1:10000 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) ab150116 Abcam 1:200 Table 1.

    Techniques: Over Expression, Knockdown, CCK-8 Assay, Flow Cytometry

    Fig. 7. Rap1 restores CD147-mediated migration and invasion in colorectal cancer cells. (A, C) Scratch wound healing assay in HCT116 cells. Healing rates were quantified at 24 h and 48 h (n = 3; ***P < 0.001 vs. shNC, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). (B, D) Transwell migration and Matrigel invasion assays. Migrated/invaded cells were counted and normalized to shCtrl (n = 3; ***P < 0.001 vs. shNC, ###P < 0.001 vs. shCD147, one-way ANOVA). SW620 cells showed consistent trends. (E) Mechanistic diagram of CD147 promoting tumor progression through Rap1/Rap1GAP signaling in colorectal cancer cells.

    Journal: Scientific reports

    Article Title: CD147 regulates the Rap1 signaling pathway to promote proliferation, migration, and invasion, and inhibit apoptosis in colorectal cancer cells.

    doi: 10.1038/s41598-025-98266-8

    Figure Lengend Snippet: Fig. 7. Rap1 restores CD147-mediated migration and invasion in colorectal cancer cells. (A, C) Scratch wound healing assay in HCT116 cells. Healing rates were quantified at 24 h and 48 h (n = 3; ***P < 0.001 vs. shNC, ##P < 0.01, ###P < 0.001 vs. shCD147, one-way ANOVA). (B, D) Transwell migration and Matrigel invasion assays. Migrated/invaded cells were counted and normalized to shCtrl (n = 3; ***P < 0.001 vs. shNC, ###P < 0.001 vs. shCD147, one-way ANOVA). SW620 cells showed consistent trends. (E) Mechanistic diagram of CD147 promoting tumor progression through Rap1/Rap1GAP signaling in colorectal cancer cells.

    Article Snippet: Supplier Antibody dilution used WB/IF Anti CD147 mouse monoclonal antibody 66443-1-AP ProteinTech Group, Inc. 1:5000 /1:200 Anti N-cadherin rabbit polyclonal antibody 22018-1-AP ProteinTech Group, Inc. 1:5000 Anti E-cadherin rabbit polyclonal antibody 20874-1-AP ProteinTech Group, Inc. 1:10000 Anti c-myc mouse monoclonal antibody 67,447-AP ProteinTech Group, Inc. 1:5000 Anti RAP1GAP rabbit polyclonal antibody 19174-1-AP ProteinTech Group, Inc. 1:2000 Anti RAP1 rabbit polyclonal antibody 14595-1-AP ProteinTech Group, Inc. 1:1000 Anti β-actin rabbit polyclonal antibody 20536-1-AP ProteinTech Group, Inc. 1:8000 HRP-conjugated Goat Anti-Mouse IgG(H + L) SA00001-1 ProteinTech Group, Inc. 1:10000 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) ab150116 Abcam 1:200 Table 1.

    Techniques: Migration, Wound Healing Assay

    Figure 2. Loss of Trip13 leads to meiotic arrest in males. (A) Image of testes from 2- to 3-month-old mice. (B) Western blot analysis of TRIP13 in P21 testes. SYCP3 serves as a meiosis-specific marker. ACTB serves as a loading control. (C) Testis to body weight ratio of 2–3-month-old mice. n = 3 males. Statistics, one-way ANOVA. (D) Sperm count of 2–3-month-old Trip13+/+ and Trip13+/- males. n = 3 males. Statistics, one-way ANOVA. (E) Histological analysis of 2-month-old testes. Sertoli, Sertoli cell; Zyg, zygotene; Pa-like, pachytene-like; Dip, diplotene; eS, elongating spermatids.

    Journal: eLife

    Article Title: TRIP13 localizes to synapsed chromosomes and functions as a dosage-sensitive regulator of meiosis

    doi: 10.7554/elife.92195

    Figure Lengend Snippet: Figure 2. Loss of Trip13 leads to meiotic arrest in males. (A) Image of testes from 2- to 3-month-old mice. (B) Western blot analysis of TRIP13 in P21 testes. SYCP3 serves as a meiosis-specific marker. ACTB serves as a loading control. (C) Testis to body weight ratio of 2–3-month-old mice. n = 3 males. Statistics, one-way ANOVA. (D) Sperm count of 2–3-month-old Trip13+/+ and Trip13+/- males. n = 3 males. Statistics, one-way ANOVA. (E) Histological analysis of 2-month-old testes. Sertoli, Sertoli cell; Zyg, zygotene; Pa-like, pachytene-like; Dip, diplotene; eS, elongating spermatids.

    Article Snippet: or resource Designation Source or reference Identifiers Additional information Gene (Mus musculus) Trip13 GenBank Gene ID: 69716 Genetic reagent (M. musculus) Trip13tm1.1(KOMP)Vlcg/JMmucd MMRCC MMRRC_050223- UCD Genetic reagent (M. musculus) Hormad1 knockout PMID:21079677; Shin et al., 2010 Rajkovic lab Genetic reagent (M. musculus) Sycp2 knockout PMID:16717126; Yang et al., 2006 Wang lab Genetic reagent (M. musculus) Rec8 knockout PMID:32232159; Guan et al., 2020 Wang lab Genetic reagent (M. musculus) 3×FLAG-Trip13 This paper Wang Lab Genetic reagent (M. musculus) Trip13-3×FLAG This paper Wang Lab Antibody Anti- ACTB (mouse monoclonal) Sigma Cat# A5441, RRID:AB_476744 WB (1:2000) Antibody Anti- centromere (CREST) (human polyclonal) Antibodies Incorporated Cat# 15- 234, RRID:AB_2687472 IF (1:500) Antibody Anti- SYCP1 (rabbit polyclonal) Abcam Cat# ab15090, RRID:AB_301636 IF (1:300) Antibody Anti- SYCP2 (guinea pig polyclonal) PMID:16717126 Custom made IF (1:150) Antibody Anti- SYCP3 (mouse monoclonal) Abcam Cat# ab97672, RRID:AB_10678841 IF (1:500) Antibody Anti- SYCP3 (rabbit polyclonal) ProteinTech Group Cat# 23024- 1- AP, RRID:AB_11232426 IF (1:500), WB (1:2000) Antibody Anti- HORMAD1 (rabbit polyclonal) ProteinTech Group Cat# 13917- 1- AP, RRID:AB_2120844 IF (1:300) Antibody Anti- HORMAD2 (rabbit polyclonal) PMID:19851446; Wojtasz et al., 2009 A gift from Toth lab IF (1:500) Antibody Anti- REC114 (rabbit polyclonal) PMID:31003867; Boekhout et al., 2019 A gift from Keeney Lab IF (1:100) Antibody Anti- REC8 (rabbit polyclonal) Custom made A gift from Mengcheng Luo lab IF (1:200) Antibody Anti- FLAG (mouse monoclonal) Sigma Cat# F3165, RRID:AB_259529 IF (1:300), WB (1:4000) Antibody Anti- SKP1 (rabbit polyclonal) Cell Signaling Cat# 12248S, RRID:AB_2754993 IF (1:200), WB (1:1000) Antibody Anti- TRIP13 (rabbit polyclonal) ProteinTech Group Cat# 19602- 1- AP IF (1:150), WB (1:1000) Chotiner et al. eLife 2023;12:RP92195.

    Techniques: Western Blot, Marker, Control

    Figure 4. Normal localization of centromere and telomere markers in Trip13-deficient spermatocytes from juvenile mice. (A–D) Immunofluorescent analysis of centromere and telomere markers in Trip13+/+ and Trip13-/- pachytene spermatocytes: CREST (A), CENPC (B), TRF1 (C), and MAJIN (D). SYCP3 labels the lateral elements of the synaptonemal complex. Scale bars, 10 µm.

    Journal: eLife

    Article Title: TRIP13 localizes to synapsed chromosomes and functions as a dosage-sensitive regulator of meiosis

    doi: 10.7554/elife.92195

    Figure Lengend Snippet: Figure 4. Normal localization of centromere and telomere markers in Trip13-deficient spermatocytes from juvenile mice. (A–D) Immunofluorescent analysis of centromere and telomere markers in Trip13+/+ and Trip13-/- pachytene spermatocytes: CREST (A), CENPC (B), TRF1 (C), and MAJIN (D). SYCP3 labels the lateral elements of the synaptonemal complex. Scale bars, 10 µm.

    Article Snippet: or resource Designation Source or reference Identifiers Additional information Gene (Mus musculus) Trip13 GenBank Gene ID: 69716 Genetic reagent (M. musculus) Trip13tm1.1(KOMP)Vlcg/JMmucd MMRCC MMRRC_050223- UCD Genetic reagent (M. musculus) Hormad1 knockout PMID:21079677; Shin et al., 2010 Rajkovic lab Genetic reagent (M. musculus) Sycp2 knockout PMID:16717126; Yang et al., 2006 Wang lab Genetic reagent (M. musculus) Rec8 knockout PMID:32232159; Guan et al., 2020 Wang lab Genetic reagent (M. musculus) 3×FLAG-Trip13 This paper Wang Lab Genetic reagent (M. musculus) Trip13-3×FLAG This paper Wang Lab Antibody Anti- ACTB (mouse monoclonal) Sigma Cat# A5441, RRID:AB_476744 WB (1:2000) Antibody Anti- centromere (CREST) (human polyclonal) Antibodies Incorporated Cat# 15- 234, RRID:AB_2687472 IF (1:500) Antibody Anti- SYCP1 (rabbit polyclonal) Abcam Cat# ab15090, RRID:AB_301636 IF (1:300) Antibody Anti- SYCP2 (guinea pig polyclonal) PMID:16717126 Custom made IF (1:150) Antibody Anti- SYCP3 (mouse monoclonal) Abcam Cat# ab97672, RRID:AB_10678841 IF (1:500) Antibody Anti- SYCP3 (rabbit polyclonal) ProteinTech Group Cat# 23024- 1- AP, RRID:AB_11232426 IF (1:500), WB (1:2000) Antibody Anti- HORMAD1 (rabbit polyclonal) ProteinTech Group Cat# 13917- 1- AP, RRID:AB_2120844 IF (1:300) Antibody Anti- HORMAD2 (rabbit polyclonal) PMID:19851446; Wojtasz et al., 2009 A gift from Toth lab IF (1:500) Antibody Anti- REC114 (rabbit polyclonal) PMID:31003867; Boekhout et al., 2019 A gift from Keeney Lab IF (1:100) Antibody Anti- REC8 (rabbit polyclonal) Custom made A gift from Mengcheng Luo lab IF (1:200) Antibody Anti- FLAG (mouse monoclonal) Sigma Cat# F3165, RRID:AB_259529 IF (1:300), WB (1:4000) Antibody Anti- SKP1 (rabbit polyclonal) Cell Signaling Cat# 12248S, RRID:AB_2754993 IF (1:200), WB (1:1000) Antibody Anti- TRIP13 (rabbit polyclonal) ProteinTech Group Cat# 19602- 1- AP IF (1:150), WB (1:1000) Chotiner et al. eLife 2023;12:RP92195.

    Techniques:

    Figure 5. HORMAD1 and HORMAD2 accumulate on the lateral elements of synapsed autosomes in Trip13-

    Journal: eLife

    Article Title: TRIP13 localizes to synapsed chromosomes and functions as a dosage-sensitive regulator of meiosis

    doi: 10.7554/elife.92195

    Figure Lengend Snippet: Figure 5. HORMAD1 and HORMAD2 accumulate on the lateral elements of synapsed autosomes in Trip13-

    Article Snippet: or resource Designation Source or reference Identifiers Additional information Gene (Mus musculus) Trip13 GenBank Gene ID: 69716 Genetic reagent (M. musculus) Trip13tm1.1(KOMP)Vlcg/JMmucd MMRCC MMRRC_050223- UCD Genetic reagent (M. musculus) Hormad1 knockout PMID:21079677; Shin et al., 2010 Rajkovic lab Genetic reagent (M. musculus) Sycp2 knockout PMID:16717126; Yang et al., 2006 Wang lab Genetic reagent (M. musculus) Rec8 knockout PMID:32232159; Guan et al., 2020 Wang lab Genetic reagent (M. musculus) 3×FLAG-Trip13 This paper Wang Lab Genetic reagent (M. musculus) Trip13-3×FLAG This paper Wang Lab Antibody Anti- ACTB (mouse monoclonal) Sigma Cat# A5441, RRID:AB_476744 WB (1:2000) Antibody Anti- centromere (CREST) (human polyclonal) Antibodies Incorporated Cat# 15- 234, RRID:AB_2687472 IF (1:500) Antibody Anti- SYCP1 (rabbit polyclonal) Abcam Cat# ab15090, RRID:AB_301636 IF (1:300) Antibody Anti- SYCP2 (guinea pig polyclonal) PMID:16717126 Custom made IF (1:150) Antibody Anti- SYCP3 (mouse monoclonal) Abcam Cat# ab97672, RRID:AB_10678841 IF (1:500) Antibody Anti- SYCP3 (rabbit polyclonal) ProteinTech Group Cat# 23024- 1- AP, RRID:AB_11232426 IF (1:500), WB (1:2000) Antibody Anti- HORMAD1 (rabbit polyclonal) ProteinTech Group Cat# 13917- 1- AP, RRID:AB_2120844 IF (1:300) Antibody Anti- HORMAD2 (rabbit polyclonal) PMID:19851446; Wojtasz et al., 2009 A gift from Toth lab IF (1:500) Antibody Anti- REC114 (rabbit polyclonal) PMID:31003867; Boekhout et al., 2019 A gift from Keeney Lab IF (1:100) Antibody Anti- REC8 (rabbit polyclonal) Custom made A gift from Mengcheng Luo lab IF (1:200) Antibody Anti- FLAG (mouse monoclonal) Sigma Cat# F3165, RRID:AB_259529 IF (1:300), WB (1:4000) Antibody Anti- SKP1 (rabbit polyclonal) Cell Signaling Cat# 12248S, RRID:AB_2754993 IF (1:200), WB (1:1000) Antibody Anti- TRIP13 (rabbit polyclonal) ProteinTech Group Cat# 19602- 1- AP IF (1:150), WB (1:1000) Chotiner et al. eLife 2023;12:RP92195.

    Techniques:

    Figure 6. Localization of TRIP13 to the synaptonemal complex (SC) is independent of individual axial element components. (A) Immunofluorescent analysis of TRIP13 in Hormad1-/- spermatocytes from 2-month-old mice. (B) Immunofluorescent analysis of TRIP13 in Rec8-/- spermatocytes from 2-month- old mice. (C) Immunofluorescent analysis of TRIP13 in Sycp2-/- spermatocytes from 2-month-old mice. (D) Immunofluorescent analysis of TRIP13 in Skp1cKO spermatocytes from 2-month-old mice. Scale bars, 10 µm.

    Journal: eLife

    Article Title: TRIP13 localizes to synapsed chromosomes and functions as a dosage-sensitive regulator of meiosis

    doi: 10.7554/elife.92195

    Figure Lengend Snippet: Figure 6. Localization of TRIP13 to the synaptonemal complex (SC) is independent of individual axial element components. (A) Immunofluorescent analysis of TRIP13 in Hormad1-/- spermatocytes from 2-month-old mice. (B) Immunofluorescent analysis of TRIP13 in Rec8-/- spermatocytes from 2-month- old mice. (C) Immunofluorescent analysis of TRIP13 in Sycp2-/- spermatocytes from 2-month-old mice. (D) Immunofluorescent analysis of TRIP13 in Skp1cKO spermatocytes from 2-month-old mice. Scale bars, 10 µm.

    Article Snippet: or resource Designation Source or reference Identifiers Additional information Gene (Mus musculus) Trip13 GenBank Gene ID: 69716 Genetic reagent (M. musculus) Trip13tm1.1(KOMP)Vlcg/JMmucd MMRCC MMRRC_050223- UCD Genetic reagent (M. musculus) Hormad1 knockout PMID:21079677; Shin et al., 2010 Rajkovic lab Genetic reagent (M. musculus) Sycp2 knockout PMID:16717126; Yang et al., 2006 Wang lab Genetic reagent (M. musculus) Rec8 knockout PMID:32232159; Guan et al., 2020 Wang lab Genetic reagent (M. musculus) 3×FLAG-Trip13 This paper Wang Lab Genetic reagent (M. musculus) Trip13-3×FLAG This paper Wang Lab Antibody Anti- ACTB (mouse monoclonal) Sigma Cat# A5441, RRID:AB_476744 WB (1:2000) Antibody Anti- centromere (CREST) (human polyclonal) Antibodies Incorporated Cat# 15- 234, RRID:AB_2687472 IF (1:500) Antibody Anti- SYCP1 (rabbit polyclonal) Abcam Cat# ab15090, RRID:AB_301636 IF (1:300) Antibody Anti- SYCP2 (guinea pig polyclonal) PMID:16717126 Custom made IF (1:150) Antibody Anti- SYCP3 (mouse monoclonal) Abcam Cat# ab97672, RRID:AB_10678841 IF (1:500) Antibody Anti- SYCP3 (rabbit polyclonal) ProteinTech Group Cat# 23024- 1- AP, RRID:AB_11232426 IF (1:500), WB (1:2000) Antibody Anti- HORMAD1 (rabbit polyclonal) ProteinTech Group Cat# 13917- 1- AP, RRID:AB_2120844 IF (1:300) Antibody Anti- HORMAD2 (rabbit polyclonal) PMID:19851446; Wojtasz et al., 2009 A gift from Toth lab IF (1:500) Antibody Anti- REC114 (rabbit polyclonal) PMID:31003867; Boekhout et al., 2019 A gift from Keeney Lab IF (1:100) Antibody Anti- REC8 (rabbit polyclonal) Custom made A gift from Mengcheng Luo lab IF (1:200) Antibody Anti- FLAG (mouse monoclonal) Sigma Cat# F3165, RRID:AB_259529 IF (1:300), WB (1:4000) Antibody Anti- SKP1 (rabbit polyclonal) Cell Signaling Cat# 12248S, RRID:AB_2754993 IF (1:200), WB (1:1000) Antibody Anti- TRIP13 (rabbit polyclonal) ProteinTech Group Cat# 19602- 1- AP IF (1:150), WB (1:1000) Chotiner et al. eLife 2023;12:RP92195.

    Techniques:

    Figure 7. FLAG-tagged TRIP13 proteins localize correctly and are functional. (A) Western blot analysis of tagged and untagged TRIP13 proteins in testes from P20 wild type (no tag), heterozygous-tagged, and homozygous-tagged males. (B) Immunofluorescence of FLAG-tagged TRIP13 in pachytene spermatocytes from P20 homozygous testes. N-terminal tag, 3×FLAG-Trip13; C-terminal tag, Trip13−3×FLAG. Scale bar, 10 μm. (C) A schematic illustration of TRIP13, SKP1, HORMAD1/2, and the synaptonemal complex. Relative locations of TRIP13 and SKP1 within the synaptonemal complex (SC) are depicted. HORAMD1/2 are retained in synapsed regions in Trip13-deficient or Skp1-deficient spermatocytes.

    Journal: eLife

    Article Title: TRIP13 localizes to synapsed chromosomes and functions as a dosage-sensitive regulator of meiosis

    doi: 10.7554/elife.92195

    Figure Lengend Snippet: Figure 7. FLAG-tagged TRIP13 proteins localize correctly and are functional. (A) Western blot analysis of tagged and untagged TRIP13 proteins in testes from P20 wild type (no tag), heterozygous-tagged, and homozygous-tagged males. (B) Immunofluorescence of FLAG-tagged TRIP13 in pachytene spermatocytes from P20 homozygous testes. N-terminal tag, 3×FLAG-Trip13; C-terminal tag, Trip13−3×FLAG. Scale bar, 10 μm. (C) A schematic illustration of TRIP13, SKP1, HORMAD1/2, and the synaptonemal complex. Relative locations of TRIP13 and SKP1 within the synaptonemal complex (SC) are depicted. HORAMD1/2 are retained in synapsed regions in Trip13-deficient or Skp1-deficient spermatocytes.

    Article Snippet: or resource Designation Source or reference Identifiers Additional information Gene (Mus musculus) Trip13 GenBank Gene ID: 69716 Genetic reagent (M. musculus) Trip13tm1.1(KOMP)Vlcg/JMmucd MMRCC MMRRC_050223- UCD Genetic reagent (M. musculus) Hormad1 knockout PMID:21079677; Shin et al., 2010 Rajkovic lab Genetic reagent (M. musculus) Sycp2 knockout PMID:16717126; Yang et al., 2006 Wang lab Genetic reagent (M. musculus) Rec8 knockout PMID:32232159; Guan et al., 2020 Wang lab Genetic reagent (M. musculus) 3×FLAG-Trip13 This paper Wang Lab Genetic reagent (M. musculus) Trip13-3×FLAG This paper Wang Lab Antibody Anti- ACTB (mouse monoclonal) Sigma Cat# A5441, RRID:AB_476744 WB (1:2000) Antibody Anti- centromere (CREST) (human polyclonal) Antibodies Incorporated Cat# 15- 234, RRID:AB_2687472 IF (1:500) Antibody Anti- SYCP1 (rabbit polyclonal) Abcam Cat# ab15090, RRID:AB_301636 IF (1:300) Antibody Anti- SYCP2 (guinea pig polyclonal) PMID:16717126 Custom made IF (1:150) Antibody Anti- SYCP3 (mouse monoclonal) Abcam Cat# ab97672, RRID:AB_10678841 IF (1:500) Antibody Anti- SYCP3 (rabbit polyclonal) ProteinTech Group Cat# 23024- 1- AP, RRID:AB_11232426 IF (1:500), WB (1:2000) Antibody Anti- HORMAD1 (rabbit polyclonal) ProteinTech Group Cat# 13917- 1- AP, RRID:AB_2120844 IF (1:300) Antibody Anti- HORMAD2 (rabbit polyclonal) PMID:19851446; Wojtasz et al., 2009 A gift from Toth lab IF (1:500) Antibody Anti- REC114 (rabbit polyclonal) PMID:31003867; Boekhout et al., 2019 A gift from Keeney Lab IF (1:100) Antibody Anti- REC8 (rabbit polyclonal) Custom made A gift from Mengcheng Luo lab IF (1:200) Antibody Anti- FLAG (mouse monoclonal) Sigma Cat# F3165, RRID:AB_259529 IF (1:300), WB (1:4000) Antibody Anti- SKP1 (rabbit polyclonal) Cell Signaling Cat# 12248S, RRID:AB_2754993 IF (1:200), WB (1:1000) Antibody Anti- TRIP13 (rabbit polyclonal) ProteinTech Group Cat# 19602- 1- AP IF (1:150), WB (1:1000) Chotiner et al. eLife 2023;12:RP92195.

    Techniques: Functional Assay, Western Blot, Immunofluorescence

    Figure 1. TRIP13 localizes to the synaptonemal complex and telomeres in spermatocytes. (A) Western blot analysis of TRIP13 in adult mouse tissues. Heart and skeletal muscle lack ACTB. (B) Immunofluorescence of TRIP13 in sections of 3-month-old wild type and Trip13-/- testes. Lep, leptotene; Zyg, zygotene; Pac, pachytene; eS, elongating spermatids; ES, elongated spermatids. (C) Immunofluorescence of TRIP13 in spread nuclei of spermatocytes from wild type P20 testes. (D) Super-resolution localization of TRIP13 to the central element (CE) but not lateral element (LE) of the synaptonemal complex at early pachytene stage. The enlarged view of the boxed chromosome is shown at the bottom.

    Journal: eLife

    Article Title: TRIP13 localizes to synapsed chromosomes and functions as a dosage-sensitive regulator of meiosis

    doi: 10.7554/elife.92195

    Figure Lengend Snippet: Figure 1. TRIP13 localizes to the synaptonemal complex and telomeres in spermatocytes. (A) Western blot analysis of TRIP13 in adult mouse tissues. Heart and skeletal muscle lack ACTB. (B) Immunofluorescence of TRIP13 in sections of 3-month-old wild type and Trip13-/- testes. Lep, leptotene; Zyg, zygotene; Pac, pachytene; eS, elongating spermatids; ES, elongated spermatids. (C) Immunofluorescence of TRIP13 in spread nuclei of spermatocytes from wild type P20 testes. (D) Super-resolution localization of TRIP13 to the central element (CE) but not lateral element (LE) of the synaptonemal complex at early pachytene stage. The enlarged view of the boxed chromosome is shown at the bottom.

    Article Snippet: or resource Designation Source or reference Identifiers Additional information Gene (Mus musculus) Trip13 GenBank Gene ID: 69716 Genetic reagent (M. musculus) Trip13tm1.1(KOMP)Vlcg/JMmucd MMRCC MMRRC_050223- UCD Genetic reagent (M. musculus) Hormad1 knockout PMID:21079677; Shin et al., 2010 Rajkovic lab Genetic reagent (M. musculus) Sycp2 knockout PMID:16717126; Yang et al., 2006 Wang lab Genetic reagent (M. musculus) Rec8 knockout PMID:32232159; Guan et al., 2020 Wang lab Genetic reagent (M. musculus) 3×FLAG-Trip13 This paper Wang Lab Genetic reagent (M. musculus) Trip13-3×FLAG This paper Wang Lab Antibody Anti- ACTB (mouse monoclonal) Sigma Cat# A5441, RRID:AB_476744 WB (1:2000) Antibody Anti- centromere (CREST) (human polyclonal) Antibodies Incorporated Cat# 15- 234, RRID:AB_2687472 IF (1:500) Antibody Anti- SYCP1 (rabbit polyclonal) Abcam Cat# ab15090, RRID:AB_301636 IF (1:300) Antibody Anti- SYCP2 (guinea pig polyclonal) PMID:16717126 Custom made IF (1:150) Antibody Anti- SYCP3 (mouse monoclonal) Abcam Cat# ab97672, RRID:AB_10678841 IF (1:500) Antibody Anti- SYCP3 (rabbit polyclonal) ProteinTech Group Cat# 23024- 1- AP, RRID:AB_11232426 IF (1:500), WB (1:2000) Antibody Anti- HORMAD1 (rabbit polyclonal) ProteinTech Group Cat# 13917- 1- AP, RRID:AB_2120844 IF (1:300) Antibody Anti- HORMAD2 (rabbit polyclonal) PMID:19851446; Wojtasz et al., 2009 A gift from Toth lab IF (1:500) Antibody Anti- REC114 (rabbit polyclonal) PMID:31003867; Boekhout et al., 2019 A gift from Keeney Lab IF (1:100) Antibody Anti- REC8 (rabbit polyclonal) Custom made A gift from Mengcheng Luo lab IF (1:200) Antibody Anti- FLAG (mouse monoclonal) Sigma Cat# F3165, RRID:AB_259529 IF (1:300), WB (1:4000) Antibody Anti- SKP1 (rabbit polyclonal) Cell Signaling Cat# 12248S, RRID:AB_2754993 IF (1:200), WB (1:1000) Antibody Anti- TRIP13 (rabbit polyclonal) ProteinTech Group Cat# 19602- 1- AP IF (1:150), WB (1:1000) Chotiner et al. eLife 2023;12:RP92195.

    Techniques: Western Blot, Immunofluorescence

    Figure 2. Loss of Trip13 leads to meiotic arrest in males. (A) Image of testes from 2- to 3-month-old mice. (B) Western blot analysis of TRIP13 in P21 testes. SYCP3 serves as a meiosis-specific marker. ACTB serves as a loading control. (C) Testis to body weight ratio of 2–3-month-old mice. n = 3 males. Statistics, one-way ANOVA. (D) Sperm count of 2–3-month-old Trip13+/+ and Trip13+/- males. n = 3 males. Statistics, one-way ANOVA. (E) Histological analysis of 2-month-old testes. Sertoli, Sertoli cell; Zyg, zygotene; Pa-like, pachytene-like; Dip, diplotene; eS, elongating spermatids.

    Journal: eLife

    Article Title: TRIP13 localizes to synapsed chromosomes and functions as a dosage-sensitive regulator of meiosis

    doi: 10.7554/elife.92195

    Figure Lengend Snippet: Figure 2. Loss of Trip13 leads to meiotic arrest in males. (A) Image of testes from 2- to 3-month-old mice. (B) Western blot analysis of TRIP13 in P21 testes. SYCP3 serves as a meiosis-specific marker. ACTB serves as a loading control. (C) Testis to body weight ratio of 2–3-month-old mice. n = 3 males. Statistics, one-way ANOVA. (D) Sperm count of 2–3-month-old Trip13+/+ and Trip13+/- males. n = 3 males. Statistics, one-way ANOVA. (E) Histological analysis of 2-month-old testes. Sertoli, Sertoli cell; Zyg, zygotene; Pa-like, pachytene-like; Dip, diplotene; eS, elongating spermatids.

    Article Snippet: or resource Designation Source or reference Identifiers Additional information Gene (Mus musculus) Trip13 GenBank Gene ID: 69716 Genetic reagent (M. musculus) Trip13tm1.1(KOMP)Vlcg/JMmucd MMRCC MMRRC_050223- UCD Genetic reagent (M. musculus) Hormad1 knockout PMID:21079677; Shin et al., 2010 Rajkovic lab Genetic reagent (M. musculus) Sycp2 knockout PMID:16717126; Yang et al., 2006 Wang lab Genetic reagent (M. musculus) Rec8 knockout PMID:32232159; Guan et al., 2020 Wang lab Genetic reagent (M. musculus) 3×FLAG-Trip13 This paper Wang Lab Genetic reagent (M. musculus) Trip13-3×FLAG This paper Wang Lab Antibody Anti- ACTB (mouse monoclonal) Sigma Cat# A5441, RRID:AB_476744 WB (1:2000) Antibody Anti- centromere (CREST) (human polyclonal) Antibodies Incorporated Cat# 15- 234, RRID:AB_2687472 IF (1:500) Antibody Anti- SYCP1 (rabbit polyclonal) Abcam Cat# ab15090, RRID:AB_301636 IF (1:300) Antibody Anti- SYCP2 (guinea pig polyclonal) PMID:16717126 Custom made IF (1:150) Antibody Anti- SYCP3 (mouse monoclonal) Abcam Cat# ab97672, RRID:AB_10678841 IF (1:500) Antibody Anti- SYCP3 (rabbit polyclonal) ProteinTech Group Cat# 23024- 1- AP, RRID:AB_11232426 IF (1:500), WB (1:2000) Antibody Anti- HORMAD1 (rabbit polyclonal) ProteinTech Group Cat# 13917- 1- AP, RRID:AB_2120844 IF (1:300) Antibody Anti- HORMAD2 (rabbit polyclonal) PMID:19851446; Wojtasz et al., 2009 A gift from Toth lab IF (1:500) Antibody Anti- REC114 (rabbit polyclonal) PMID:31003867; Boekhout et al., 2019 A gift from Keeney Lab IF (1:100) Antibody Anti- REC8 (rabbit polyclonal) Custom made A gift from Mengcheng Luo lab IF (1:200) Antibody Anti- FLAG (mouse monoclonal) Sigma Cat# F3165, RRID:AB_259529 IF (1:300), WB (1:4000) Antibody Anti- SKP1 (rabbit polyclonal) Cell Signaling Cat# 12248S, RRID:AB_2754993 IF (1:200), WB (1:1000) Antibody Anti- TRIP13 (rabbit polyclonal) ProteinTech Group Cat# 19602- 1- AP IF (1:150), WB (1:1000) Chotiner et al. eLife 2023;12:RP92195.

    Techniques: Western Blot, Marker, Control

    Figure 4. Normal localization of centromere and telomere markers in Trip13-deficient spermatocytes from juvenile mice. (A–D) Immunofluorescent analysis of centromere and telomere markers in Trip13+/+ and Trip13-/- pachytene spermatocytes: CREST (A), CENPC (B), TRF1 (C), and MAJIN (D). SYCP3 labels the lateral elements of the synaptonemal complex. Scale bars, 10 µm.

    Journal: eLife

    Article Title: TRIP13 localizes to synapsed chromosomes and functions as a dosage-sensitive regulator of meiosis

    doi: 10.7554/elife.92195

    Figure Lengend Snippet: Figure 4. Normal localization of centromere and telomere markers in Trip13-deficient spermatocytes from juvenile mice. (A–D) Immunofluorescent analysis of centromere and telomere markers in Trip13+/+ and Trip13-/- pachytene spermatocytes: CREST (A), CENPC (B), TRF1 (C), and MAJIN (D). SYCP3 labels the lateral elements of the synaptonemal complex. Scale bars, 10 µm.

    Article Snippet: or resource Designation Source or reference Identifiers Additional information Gene (Mus musculus) Trip13 GenBank Gene ID: 69716 Genetic reagent (M. musculus) Trip13tm1.1(KOMP)Vlcg/JMmucd MMRCC MMRRC_050223- UCD Genetic reagent (M. musculus) Hormad1 knockout PMID:21079677; Shin et al., 2010 Rajkovic lab Genetic reagent (M. musculus) Sycp2 knockout PMID:16717126; Yang et al., 2006 Wang lab Genetic reagent (M. musculus) Rec8 knockout PMID:32232159; Guan et al., 2020 Wang lab Genetic reagent (M. musculus) 3×FLAG-Trip13 This paper Wang Lab Genetic reagent (M. musculus) Trip13-3×FLAG This paper Wang Lab Antibody Anti- ACTB (mouse monoclonal) Sigma Cat# A5441, RRID:AB_476744 WB (1:2000) Antibody Anti- centromere (CREST) (human polyclonal) Antibodies Incorporated Cat# 15- 234, RRID:AB_2687472 IF (1:500) Antibody Anti- SYCP1 (rabbit polyclonal) Abcam Cat# ab15090, RRID:AB_301636 IF (1:300) Antibody Anti- SYCP2 (guinea pig polyclonal) PMID:16717126 Custom made IF (1:150) Antibody Anti- SYCP3 (mouse monoclonal) Abcam Cat# ab97672, RRID:AB_10678841 IF (1:500) Antibody Anti- SYCP3 (rabbit polyclonal) ProteinTech Group Cat# 23024- 1- AP, RRID:AB_11232426 IF (1:500), WB (1:2000) Antibody Anti- HORMAD1 (rabbit polyclonal) ProteinTech Group Cat# 13917- 1- AP, RRID:AB_2120844 IF (1:300) Antibody Anti- HORMAD2 (rabbit polyclonal) PMID:19851446; Wojtasz et al., 2009 A gift from Toth lab IF (1:500) Antibody Anti- REC114 (rabbit polyclonal) PMID:31003867; Boekhout et al., 2019 A gift from Keeney Lab IF (1:100) Antibody Anti- REC8 (rabbit polyclonal) Custom made A gift from Mengcheng Luo lab IF (1:200) Antibody Anti- FLAG (mouse monoclonal) Sigma Cat# F3165, RRID:AB_259529 IF (1:300), WB (1:4000) Antibody Anti- SKP1 (rabbit polyclonal) Cell Signaling Cat# 12248S, RRID:AB_2754993 IF (1:200), WB (1:1000) Antibody Anti- TRIP13 (rabbit polyclonal) ProteinTech Group Cat# 19602- 1- AP IF (1:150), WB (1:1000) Chotiner et al. eLife 2023;12:RP92195.

    Techniques:

    Figure 5. HORMAD1 and HORMAD2 accumulate on the lateral elements of synapsed autosomes in Trip13-

    Journal: eLife

    Article Title: TRIP13 localizes to synapsed chromosomes and functions as a dosage-sensitive regulator of meiosis

    doi: 10.7554/elife.92195

    Figure Lengend Snippet: Figure 5. HORMAD1 and HORMAD2 accumulate on the lateral elements of synapsed autosomes in Trip13-

    Article Snippet: or resource Designation Source or reference Identifiers Additional information Gene (Mus musculus) Trip13 GenBank Gene ID: 69716 Genetic reagent (M. musculus) Trip13tm1.1(KOMP)Vlcg/JMmucd MMRCC MMRRC_050223- UCD Genetic reagent (M. musculus) Hormad1 knockout PMID:21079677; Shin et al., 2010 Rajkovic lab Genetic reagent (M. musculus) Sycp2 knockout PMID:16717126; Yang et al., 2006 Wang lab Genetic reagent (M. musculus) Rec8 knockout PMID:32232159; Guan et al., 2020 Wang lab Genetic reagent (M. musculus) 3×FLAG-Trip13 This paper Wang Lab Genetic reagent (M. musculus) Trip13-3×FLAG This paper Wang Lab Antibody Anti- ACTB (mouse monoclonal) Sigma Cat# A5441, RRID:AB_476744 WB (1:2000) Antibody Anti- centromere (CREST) (human polyclonal) Antibodies Incorporated Cat# 15- 234, RRID:AB_2687472 IF (1:500) Antibody Anti- SYCP1 (rabbit polyclonal) Abcam Cat# ab15090, RRID:AB_301636 IF (1:300) Antibody Anti- SYCP2 (guinea pig polyclonal) PMID:16717126 Custom made IF (1:150) Antibody Anti- SYCP3 (mouse monoclonal) Abcam Cat# ab97672, RRID:AB_10678841 IF (1:500) Antibody Anti- SYCP3 (rabbit polyclonal) ProteinTech Group Cat# 23024- 1- AP, RRID:AB_11232426 IF (1:500), WB (1:2000) Antibody Anti- HORMAD1 (rabbit polyclonal) ProteinTech Group Cat# 13917- 1- AP, RRID:AB_2120844 IF (1:300) Antibody Anti- HORMAD2 (rabbit polyclonal) PMID:19851446; Wojtasz et al., 2009 A gift from Toth lab IF (1:500) Antibody Anti- REC114 (rabbit polyclonal) PMID:31003867; Boekhout et al., 2019 A gift from Keeney Lab IF (1:100) Antibody Anti- REC8 (rabbit polyclonal) Custom made A gift from Mengcheng Luo lab IF (1:200) Antibody Anti- FLAG (mouse monoclonal) Sigma Cat# F3165, RRID:AB_259529 IF (1:300), WB (1:4000) Antibody Anti- SKP1 (rabbit polyclonal) Cell Signaling Cat# 12248S, RRID:AB_2754993 IF (1:200), WB (1:1000) Antibody Anti- TRIP13 (rabbit polyclonal) ProteinTech Group Cat# 19602- 1- AP IF (1:150), WB (1:1000) Chotiner et al. eLife 2023;12:RP92195.

    Techniques:

    Figure 6. Localization of TRIP13 to the synaptonemal complex (SC) is independent of individual axial element components. (A) Immunofluorescent analysis of TRIP13 in Hormad1-/- spermatocytes from 2-month-old mice. (B) Immunofluorescent analysis of TRIP13 in Rec8-/- spermatocytes from 2-month- old mice. (C) Immunofluorescent analysis of TRIP13 in Sycp2-/- spermatocytes from 2-month-old mice. (D) Immunofluorescent analysis of TRIP13 in Skp1cKO spermatocytes from 2-month-old mice. Scale bars, 10 µm.

    Journal: eLife

    Article Title: TRIP13 localizes to synapsed chromosomes and functions as a dosage-sensitive regulator of meiosis

    doi: 10.7554/elife.92195

    Figure Lengend Snippet: Figure 6. Localization of TRIP13 to the synaptonemal complex (SC) is independent of individual axial element components. (A) Immunofluorescent analysis of TRIP13 in Hormad1-/- spermatocytes from 2-month-old mice. (B) Immunofluorescent analysis of TRIP13 in Rec8-/- spermatocytes from 2-month- old mice. (C) Immunofluorescent analysis of TRIP13 in Sycp2-/- spermatocytes from 2-month-old mice. (D) Immunofluorescent analysis of TRIP13 in Skp1cKO spermatocytes from 2-month-old mice. Scale bars, 10 µm.

    Article Snippet: or resource Designation Source or reference Identifiers Additional information Gene (Mus musculus) Trip13 GenBank Gene ID: 69716 Genetic reagent (M. musculus) Trip13tm1.1(KOMP)Vlcg/JMmucd MMRCC MMRRC_050223- UCD Genetic reagent (M. musculus) Hormad1 knockout PMID:21079677; Shin et al., 2010 Rajkovic lab Genetic reagent (M. musculus) Sycp2 knockout PMID:16717126; Yang et al., 2006 Wang lab Genetic reagent (M. musculus) Rec8 knockout PMID:32232159; Guan et al., 2020 Wang lab Genetic reagent (M. musculus) 3×FLAG-Trip13 This paper Wang Lab Genetic reagent (M. musculus) Trip13-3×FLAG This paper Wang Lab Antibody Anti- ACTB (mouse monoclonal) Sigma Cat# A5441, RRID:AB_476744 WB (1:2000) Antibody Anti- centromere (CREST) (human polyclonal) Antibodies Incorporated Cat# 15- 234, RRID:AB_2687472 IF (1:500) Antibody Anti- SYCP1 (rabbit polyclonal) Abcam Cat# ab15090, RRID:AB_301636 IF (1:300) Antibody Anti- SYCP2 (guinea pig polyclonal) PMID:16717126 Custom made IF (1:150) Antibody Anti- SYCP3 (mouse monoclonal) Abcam Cat# ab97672, RRID:AB_10678841 IF (1:500) Antibody Anti- SYCP3 (rabbit polyclonal) ProteinTech Group Cat# 23024- 1- AP, RRID:AB_11232426 IF (1:500), WB (1:2000) Antibody Anti- HORMAD1 (rabbit polyclonal) ProteinTech Group Cat# 13917- 1- AP, RRID:AB_2120844 IF (1:300) Antibody Anti- HORMAD2 (rabbit polyclonal) PMID:19851446; Wojtasz et al., 2009 A gift from Toth lab IF (1:500) Antibody Anti- REC114 (rabbit polyclonal) PMID:31003867; Boekhout et al., 2019 A gift from Keeney Lab IF (1:100) Antibody Anti- REC8 (rabbit polyclonal) Custom made A gift from Mengcheng Luo lab IF (1:200) Antibody Anti- FLAG (mouse monoclonal) Sigma Cat# F3165, RRID:AB_259529 IF (1:300), WB (1:4000) Antibody Anti- SKP1 (rabbit polyclonal) Cell Signaling Cat# 12248S, RRID:AB_2754993 IF (1:200), WB (1:1000) Antibody Anti- TRIP13 (rabbit polyclonal) ProteinTech Group Cat# 19602- 1- AP IF (1:150), WB (1:1000) Chotiner et al. eLife 2023;12:RP92195.

    Techniques:

    Figure 7. FLAG-tagged TRIP13 proteins localize correctly and are functional. (A) Western blot analysis of tagged and untagged TRIP13 proteins in testes from P20 wild type (no tag), heterozygous-tagged, and homozygous-tagged males. (B) Immunofluorescence of FLAG-tagged TRIP13 in pachytene spermatocytes from P20 homozygous testes. N-terminal tag, 3×FLAG-Trip13; C-terminal tag, Trip13−3×FLAG. Scale bar, 10 μm. (C) A schematic illustration of TRIP13, SKP1, HORMAD1/2, and the synaptonemal complex. Relative locations of TRIP13 and SKP1 within the synaptonemal complex (SC) are depicted. HORAMD1/2 are retained in synapsed regions in Trip13-deficient or Skp1-deficient spermatocytes.

    Journal: eLife

    Article Title: TRIP13 localizes to synapsed chromosomes and functions as a dosage-sensitive regulator of meiosis

    doi: 10.7554/elife.92195

    Figure Lengend Snippet: Figure 7. FLAG-tagged TRIP13 proteins localize correctly and are functional. (A) Western blot analysis of tagged and untagged TRIP13 proteins in testes from P20 wild type (no tag), heterozygous-tagged, and homozygous-tagged males. (B) Immunofluorescence of FLAG-tagged TRIP13 in pachytene spermatocytes from P20 homozygous testes. N-terminal tag, 3×FLAG-Trip13; C-terminal tag, Trip13−3×FLAG. Scale bar, 10 μm. (C) A schematic illustration of TRIP13, SKP1, HORMAD1/2, and the synaptonemal complex. Relative locations of TRIP13 and SKP1 within the synaptonemal complex (SC) are depicted. HORAMD1/2 are retained in synapsed regions in Trip13-deficient or Skp1-deficient spermatocytes.

    Article Snippet: or resource Designation Source or reference Identifiers Additional information Gene (Mus musculus) Trip13 GenBank Gene ID: 69716 Genetic reagent (M. musculus) Trip13tm1.1(KOMP)Vlcg/JMmucd MMRCC MMRRC_050223- UCD Genetic reagent (M. musculus) Hormad1 knockout PMID:21079677; Shin et al., 2010 Rajkovic lab Genetic reagent (M. musculus) Sycp2 knockout PMID:16717126; Yang et al., 2006 Wang lab Genetic reagent (M. musculus) Rec8 knockout PMID:32232159; Guan et al., 2020 Wang lab Genetic reagent (M. musculus) 3×FLAG-Trip13 This paper Wang Lab Genetic reagent (M. musculus) Trip13-3×FLAG This paper Wang Lab Antibody Anti- ACTB (mouse monoclonal) Sigma Cat# A5441, RRID:AB_476744 WB (1:2000) Antibody Anti- centromere (CREST) (human polyclonal) Antibodies Incorporated Cat# 15- 234, RRID:AB_2687472 IF (1:500) Antibody Anti- SYCP1 (rabbit polyclonal) Abcam Cat# ab15090, RRID:AB_301636 IF (1:300) Antibody Anti- SYCP2 (guinea pig polyclonal) PMID:16717126 Custom made IF (1:150) Antibody Anti- SYCP3 (mouse monoclonal) Abcam Cat# ab97672, RRID:AB_10678841 IF (1:500) Antibody Anti- SYCP3 (rabbit polyclonal) ProteinTech Group Cat# 23024- 1- AP, RRID:AB_11232426 IF (1:500), WB (1:2000) Antibody Anti- HORMAD1 (rabbit polyclonal) ProteinTech Group Cat# 13917- 1- AP, RRID:AB_2120844 IF (1:300) Antibody Anti- HORMAD2 (rabbit polyclonal) PMID:19851446; Wojtasz et al., 2009 A gift from Toth lab IF (1:500) Antibody Anti- REC114 (rabbit polyclonal) PMID:31003867; Boekhout et al., 2019 A gift from Keeney Lab IF (1:100) Antibody Anti- REC8 (rabbit polyclonal) Custom made A gift from Mengcheng Luo lab IF (1:200) Antibody Anti- FLAG (mouse monoclonal) Sigma Cat# F3165, RRID:AB_259529 IF (1:300), WB (1:4000) Antibody Anti- SKP1 (rabbit polyclonal) Cell Signaling Cat# 12248S, RRID:AB_2754993 IF (1:200), WB (1:1000) Antibody Anti- TRIP13 (rabbit polyclonal) ProteinTech Group Cat# 19602- 1- AP IF (1:150), WB (1:1000) Chotiner et al. eLife 2023;12:RP92195.

    Techniques: Functional Assay, Western Blot, Immunofluorescence