Journal: Nature Communications

Article Title: USP24 induces IL-6 in tumor-associated microenvironment by stabilizing p300 and β-TrCP and promotes cancer malignancy

doi: 10.1038/s41467-018-06178-1

Figure Lengend Snippet: USP24 level in M2 macrophages. a Representative images of immunohistochemistry staining of USP24 in lung cancer specimens by using anti-USP24 antibody. Leukocytes were circled with red dashed lines. Scale bar represents 200 μm. b Immunofluorescence staining of USP24, CD68, and DAPI in one human lung cancer specimen. Scale bar represents 50 μm. c Morphological change of M2 macrophages derived from THP-1 monocytes. Scale bar represents 200 μm. d , e USP24 protein ( n = 3) ( d ) and RNA ( n = 3) ( e ) level in THP-1 and M2 macrophages were analyzed by western blotting and RT-PCR. Results were normalized with tubulin or GAPDH level and expressed as fold of control. Data are shown as mean ± SEM, two-tailed unpaired Student’s t -test, * P < 0.05 and ** P < 0.01

Article Snippet: A549 cells were overexpressed with myc-ubi and treated MG132 (Sigma-Aldrich) for 12 h. β-TrCP protein in A549 cell lysates were immunoprecipitated by incubating with anti-β-TrCP antibody for 4 h, and then incubated with protein A agarose (Millipore) for 1 h. After washing, substrate was mixed with human recombinant USP24 protein (50 μg per ml) (Origene) in deubiquitination buffer (50 mM Tris PH 8.0, 10 mM DTT and 5 μM MG132) for 2 h in 37 °C.

Techniques: Immunohistochemistry, Staining, Immunofluorescence, Derivative Assay, Western Blot, Reverse Transcription Polymerase Chain Reaction, Two Tailed Test

Journal: Nature Communications

Article Title: USP24 induces IL-6 in tumor-associated microenvironment by stabilizing p300 and β-TrCP and promotes cancer malignancy

doi: 10.1038/s41467-018-06178-1

Figure Lengend Snippet: The metastasis-related effects of conditioned medium derived from M2 macrophages. a A549 cells were treated with RPMI or conditioned medium derived from THP-1 or M2 macrophages for 24 h, and transwell migration assay was performed to analyze the migratory ability of lung cancer cells (DAPI staining, n = 9). Scale bar represents 60 μm. b – d A549 cells were treated with RPMI or conditioned medium derived from USP24 knockdown or USP24 overexpression M2 macrophages for 24 h, and transwell migration assay ( b ) (DAPI staining, n = 9, scale bar: 100 μm; Giemsa staining, n = 4, scale bar: 60 μm), chemotactic assay ( c ) (DAPI staining, n = 3, scale bar: 100 μm; Giemsa staining, n = 4, scale bar: 60 μm) and wound-healing migration assay ( d ) were performed. Wound edges were indicated with red dashed lines and wound width was presented as solid red lines. Scale bar represents 200 μm. e HMEC-1 cells were treated with RPMI or conditioned medium derived from scramble- or USP24-knockdown M2 macrophages for 6 h. Cells were stained with F-actin and photographed ( n = 6). Results were normalized with control and expressed as fold of control. Scale bar represents 200 μm. Data are shown as mean ± SEM, two-tailed unpaired Student’s t -test, * P < 0.05, ** P < 0.01, and *** P < 0.005

Article Snippet: A549 cells were overexpressed with myc-ubi and treated MG132 (Sigma-Aldrich) for 12 h. β-TrCP protein in A549 cell lysates were immunoprecipitated by incubating with anti-β-TrCP antibody for 4 h, and then incubated with protein A agarose (Millipore) for 1 h. After washing, substrate was mixed with human recombinant USP24 protein (50 μg per ml) (Origene) in deubiquitination buffer (50 mM Tris PH 8.0, 10 mM DTT and 5 μM MG132) for 2 h in 37 °C.

Techniques: Derivative Assay, Transwell Migration Assay, Staining, Over Expression, Chemotaxis Assay, Migration, Two Tailed Test

Journal: Nature Communications

Article Title: USP24 induces IL-6 in tumor-associated microenvironment by stabilizing p300 and β-TrCP and promotes cancer malignancy

doi: 10.1038/s41467-018-06178-1

Figure Lengend Snippet: IL-6 regulated by USP24 increases lung cancer metastasis. a – j Western blot and Q-PCR was utilized to analyze the protein level of USP24 ( a , f ) and mRNA levels of USP24 , IL-6 , IL-8 , and IL-10 in M2 macrophages after USP24 knockdown ( b – e ) or overexpression ( g – j ) ( n = 3). k , l IL-6 mRNA level in A549 cells was analyzed by RT-PCR ( k ) and Q-PCR ( l ) after USP24 knockdown ( n = 3). Results of Q-PCR were normalized with GAPDH and expressed as fold of control. m , n IL-6 secretion was analyzed in conditioned medium derived from M2 macrophages ( m ) and A549 cells ( n ) after USP24 knockdown ( n = 12). o – q IL-6 was replenished in USP24-knockdown M2 macrophages derived conditioned medium, and migration assay ( o ) ( n = 6), chemotaxis assay ( p ) ( n = 3) (DAPI staining, scale bar: 100 μm), and angiogenesis assay ( q ) ( n = 12) (scale bar: 200 μm.) were analyzed. Results were normalized with control and expressed as fold of control. r The levels of IL-6, Thrombospondin-1, TNF-α, VEGF, Angiopoietin-2 and CD40L were studied by using protein array. s CL1–5 cells were treated with DMEM ( n = 5) or conditioned medium derived from scramble knockdown ( n = 4), USP24 knockdown ( n = 4) or IL-6 replenished USP24 knockdown ( n = 5) M2 macrophages for 24 h and injected into SCID mice through tail vein injection. The white arrowheads indicate the metastatic nodules (scale bar: 1 cm). Data are shown as mean ± SEM, two-tailed unpaired Student’s t -test, ns for not significant, * P < 0.05, ** P < 0.01, and *** P < 0.005

Article Snippet: A549 cells were overexpressed with myc-ubi and treated MG132 (Sigma-Aldrich) for 12 h. β-TrCP protein in A549 cell lysates were immunoprecipitated by incubating with anti-β-TrCP antibody for 4 h, and then incubated with protein A agarose (Millipore) for 1 h. After washing, substrate was mixed with human recombinant USP24 protein (50 μg per ml) (Origene) in deubiquitination buffer (50 mM Tris PH 8.0, 10 mM DTT and 5 μM MG132) for 2 h in 37 °C.

Techniques: Western Blot, Over Expression, Reverse Transcription Polymerase Chain Reaction, Derivative Assay, Migration, Chemotaxis Assay, Staining, Angiogenesis Assay, Protein Array, Injection, Two Tailed Test

Journal: Nature Communications

Article Title: USP24 induces IL-6 in tumor-associated microenvironment by stabilizing p300 and β-TrCP and promotes cancer malignancy

doi: 10.1038/s41467-018-06178-1

Figure Lengend Snippet: USP24-stabilized p300 increases NF-κB and IL-6 expression in M2 macrophages. a Western blotting was used to analyze the indicated protein levels in scramble- and USP24-knockdown M2 macrophages by using two different shRNA clones. b Q-PCR was used to analyze IL-6 RNA level in scramble- and p300-knockdown M2 macrophages. c IL-6 level in conditioned medium derived from scramble- and p300-knockdown M2 macrophage was studied by ELISA ( n = 4). d The recruitment of acetyl histone H3 to the promoters of IL-6 and NF-κB in USP24-knockdown M2 macrophages was analyzed by chromatin immunoprecipitation (IP) assay. e The lysates were harvested from M2 macrophages for immunoprecipitation with anti-USP24 antibodies. IP samples were used to study the USP24 and p300 levels by western blotting with anti-USP24 and anti-p300 antibodies. f The lysates were harvested from scramble- and USP24-knockdown M2 macrophage for immunoprecipitation with anti-p300 antibodies. IP samples were used to study the ubiquitinated p300 by western blotting with anti-ubiquitin antibody. g The lysates collected from scramble- or USP24-knockdown M2 macrophages with MG132 treatment were used to study the indicated protein levels by western blotting. h Scramble and USP24-knockdown M2 macrophages were treated with CHX and collected at indicated time points. p300 protein stability was analyzed by western blotting. i , j NF-κB protein and mRNA levels were measured in scramble- and USP24-knockdown M2 macrophages by western blotting, RT-PCR ( i ) and Q-PCR ( j ). k , l Western blotting ( k ) and Q-PCR ( l ) were used to analyze indicated protein and mRNA levels in scramble- and p300-knockdown M2 macrophages. m The lysates were harvested from A549 cells with or without USP24 knockdown or p300 overexpression and analyzed by using the western blotting and RT-PCR. Protein and mRNA levels were quantified after independent experiments ( n = 3). Results of western blot and Q-PCR were normalized with actin, tubulin or GAPDH and expressed as fold of control. Data are shown as mean ± SEM, two-tailed unpaired Student’s t -test, * P < 0.05, ** P < 0 .01, and *** P < 0.005

Article Snippet: A549 cells were overexpressed with myc-ubi and treated MG132 (Sigma-Aldrich) for 12 h. β-TrCP protein in A549 cell lysates were immunoprecipitated by incubating with anti-β-TrCP antibody for 4 h, and then incubated with protein A agarose (Millipore) for 1 h. After washing, substrate was mixed with human recombinant USP24 protein (50 μg per ml) (Origene) in deubiquitination buffer (50 mM Tris PH 8.0, 10 mM DTT and 5 μM MG132) for 2 h in 37 °C.

Techniques: Expressing, Western Blot, shRNA, Clone Assay, Derivative Assay, Enzyme-linked Immunosorbent Assay, Chromatin Immunoprecipitation, Immunoprecipitation, Reverse Transcription Polymerase Chain Reaction, Over Expression, Two Tailed Test

Journal: Nature Communications

Article Title: USP24 induces IL-6 in tumor-associated microenvironment by stabilizing p300 and β-TrCP and promotes cancer malignancy

doi: 10.1038/s41467-018-06178-1

Figure Lengend Snippet: Methylation status of IL-6 promoter and DNMT1 level in USP24-knockdown A549 cells. a Reporter assay was performed to analyze IL-6 promoter activity in A549 cells after scramble knockdown or USP24 knockdown ( n = 6). b Western blotting was used to analyze indicated protein level in scramble and USP24-knockdown A549 cells by using two different shRNA clones ( n = 3). c Recruitment of acetyl histone H3 to the promoter of IL-6 in scramble- and USP24-knockdown A549 cells was analyzed by chromatin immunoprecipitation assay. Anti-acetyl histone H3 antibody was used and IL-6 promoter was analyzed by RT-PCR and Q-PCR ( n = 3). d Bisulfite sequencing was used to analyze methylation sites in between −375 and −19 region. PCR products were inserted into yT&A vectors and amplified with competent cells. Sequences of the colonies derived from these competent cells were analyzed by Mission Biotech. Methylation status of each site analyzed by bisulfite sequencing in A549 cells was represented in percentage. e DNMT1, DNMT3a, and DNMT3a/b levels were analyzed in scramble- and USP24-knockdown A549 cells by western blotting ( n = 3). f Scramble- and USP24-knockdown A549 cells were treated with CHX and collected at indicated time points. DNMT1 protein stability was analyzed by western blotting ( n = 3). g USP24 was knockdown in A549 cells, and then cells were harvested for immunoprecipitation assay with anti-DNMT1 and anti-Ubi antibodies. IP samples were analyzed by western blotting with antibodies against the indicated proteins. h Samples collected from A549 cells with or without knockdown of USP24 and DNMT1 were used to study the mRNA levels of USP24 , IL-6 , and GAPDH by using RT-PCR. Results of western blot and Q-PCR were normalized with actin, tubulin or reads of scramble-IgG and expressed as fold of control. Data are shown as mean ± SEM, two-tailed unpaired Student’s t -test, ns for not significant, * P < 0.05, ** P < 0.01, and *** P < 0.005

Article Snippet: A549 cells were overexpressed with myc-ubi and treated MG132 (Sigma-Aldrich) for 12 h. β-TrCP protein in A549 cell lysates were immunoprecipitated by incubating with anti-β-TrCP antibody for 4 h, and then incubated with protein A agarose (Millipore) for 1 h. After washing, substrate was mixed with human recombinant USP24 protein (50 μg per ml) (Origene) in deubiquitination buffer (50 mM Tris PH 8.0, 10 mM DTT and 5 μM MG132) for 2 h in 37 °C.

Techniques: Methylation, Reporter Assay, Activity Assay, Western Blot, shRNA, Clone Assay, Chromatin Immunoprecipitation, Reverse Transcription Polymerase Chain Reaction, Methylation Sequencing, Amplification, Derivative Assay, Immunoprecipitation, Two Tailed Test

Journal: Nature Communications

Article Title: USP24 induces IL-6 in tumor-associated microenvironment by stabilizing p300 and β-TrCP and promotes cancer malignancy

doi: 10.1038/s41467-018-06178-1

Figure Lengend Snippet: USP24 stabilizes β-TrCP in A549 cells. a Western blotting was used to analyze β-TrCP level in scramble knockdown and USP24 knockdown A549 cells by using two different shRNA clones ( n = 3). b Scramble- and USP24-knockdown A549 cells were treated with CHX and β-TrCP protein stability was analyzed by western blotting ( n = 3). c β-TrCP overexpressed A549 cells were collected for immunoprecipitation with anti-USP24 antibody and indicated proteins were analyzed by western blotting. d Scramble-knockdown and USP24-knockdown A549 cells were overexpressed with myc-ubi and collected for immunoprecipitation with anti-β-TrCP antibody. Indicated protein was analyzed by western blotting. e A549 cells were overexpressed with myc-ubi and harvested for immunoprecipitation with anti-β-TrCP antibody. Human purified USP24 protein was mixed with IP sample and analyzed by western blotting. f Indicated protein levels of samples from scramble or USP24 knockdown A549 cells with or without MG132 treatment were analyzed by western blotting. Results of western blot were normalized with actin and expressed as fold of control. Data are shown as mean ± SEM, two-tailed unpaired Student’s t -test, *** P < 0.005

Article Snippet: A549 cells were overexpressed with myc-ubi and treated MG132 (Sigma-Aldrich) for 12 h. β-TrCP protein in A549 cell lysates were immunoprecipitated by incubating with anti-β-TrCP antibody for 4 h, and then incubated with protein A agarose (Millipore) for 1 h. After washing, substrate was mixed with human recombinant USP24 protein (50 μg per ml) (Origene) in deubiquitination buffer (50 mM Tris PH 8.0, 10 mM DTT and 5 μM MG132) for 2 h in 37 °C.

Techniques: Western Blot, shRNA, Clone Assay, Immunoprecipitation, Purification, Two Tailed Test

Journal: Nature Communications

Article Title: USP24 induces IL-6 in tumor-associated microenvironment by stabilizing p300 and β-TrCP and promotes cancer malignancy

doi: 10.1038/s41467-018-06178-1

Figure Lengend Snippet: The role of β-TrCP in USP24 knockdown increased DNMT1 and IκB. a Indicated protein levels were examined in scramble knockdown and USP24-knockdown A549 cells by western blotting ( n = 3). b Scramble- and USP24-knockdown A549 cells were treated with CHX and IκB protein stability was analyzed by western blotting ( n = 3). c Scramble- and USP24-knockdown A549 cells were overexpressed with myc-ubi and cells were collected for immunoprecipitation with anti-myc antibody. IP samples were analyzed by western blotting. d Scramble-knockdown and USP24-knockdown A549 cells were overexpressed with β-TrCP and collected for analyzing indicated proteins by western blotting ( n = 3). Results of western blot were normalized with actin and expressed as fold of control. e Indicated proteins and mRNAs levels in A549 cells with or without USP24 knockdown or β-TrCP overexpression were analyzed by western blotting and RT-PCR. f GFP and GFP-USP24 overexpressed A549 cells were fixed for immunofluorescence assay with indicated antibodies. GFP or GFP-USP24 overexpressed cells were indicated with white arrowheads. Yellow arrowheads indicate GFP-USP24 negatively expressed cells in GFP-USP24 overexpression group. Scale bar represents 10 μm. g Luciferase activity was measured by transfecting NF-κB response element (RE) containing vector into scramble knockdown, USP24 knockdown, GFP overexpressed, and GFP-USP24 overexpressed A549 cells ( n = 3). Results were normalized with control and expressed as fold of control. h , i The intersections between 452 NF-κB target genes (blue circle), 125 USP24 upregulated genes (red circle) and 251 USP24 downregulated genes (green circle) were analyzed ( h ). Heatmaps of intersected were shown, and metastatic genes were indicated by asterisks ( i ). Data are shown as mean ± SEM, two-tailed unpaired Student’s t -test, ns for not significant, * P < 0.05 and *** P < 0.005

Article Snippet: A549 cells were overexpressed with myc-ubi and treated MG132 (Sigma-Aldrich) for 12 h. β-TrCP protein in A549 cell lysates were immunoprecipitated by incubating with anti-β-TrCP antibody for 4 h, and then incubated with protein A agarose (Millipore) for 1 h. After washing, substrate was mixed with human recombinant USP24 protein (50 μg per ml) (Origene) in deubiquitination buffer (50 mM Tris PH 8.0, 10 mM DTT and 5 μM MG132) for 2 h in 37 °C.

Techniques: Western Blot, Immunoprecipitation, Over Expression, Reverse Transcription Polymerase Chain Reaction, Immunofluorescence, Luciferase, Activity Assay, Plasmid Preparation, Two Tailed Test

Journal: Nature Communications

Article Title: USP24 induces IL-6 in tumor-associated microenvironment by stabilizing p300 and β-TrCP and promotes cancer malignancy

doi: 10.1038/s41467-018-06178-1

Figure Lengend Snippet: Correlation between M2 macrophage marker, USP24, p300, NF-κB, β-TrCP, and DNMT1 in lung cancer specimens. a Eight weeks old of EGFR L858R transgenic mice were treated with doxycycline for 3 months or 5 months to induce the formation of lung cancer, and then sacrificed at indicated time points. Lungs were harvested and immunohistochemistry was performed to analyze the expression of indicated proteins. YM-1 positive area was marked with blue dashed lines. Scale bar represents 50 μm. b Immunohistochemistry staining was used to analyze indicated protein in 50 human lung cancer specimens. Each slide was given with different scores to indicate the intensity of protein signals. Samples with score 1 and 2 were considered as low protein expression, and score 3 and 4 were considered as high protein expression. Scale bar represents 50 μm. In slides stained with anti-USP24 antibody, 30 samples were identified as low expression, and 20 samples were identified as high expression. Expression level of indicated proteins in low USP24 expression groups or high USP24 expression groups were shown in percentage. Correlation between USP24 expression and p300, β-TrCP, and DNMT1 were examined by Fisher’s exact test. P value of the correlation of USP24 to p300, USP24 to β-TrCP, and USP24 to DNMT1 were 0.1825, 0.0002, and 0.0421, respectively

Article Snippet: A549 cells were overexpressed with myc-ubi and treated MG132 (Sigma-Aldrich) for 12 h. β-TrCP protein in A549 cell lysates were immunoprecipitated by incubating with anti-β-TrCP antibody for 4 h, and then incubated with protein A agarose (Millipore) for 1 h. After washing, substrate was mixed with human recombinant USP24 protein (50 μg per ml) (Origene) in deubiquitination buffer (50 mM Tris PH 8.0, 10 mM DTT and 5 μM MG132) for 2 h in 37 °C.

Techniques: Marker, Transgenic Assay, Immunohistochemistry, Expressing, Staining

Journal: Nature Communications

Article Title: USP24 induces IL-6 in tumor-associated microenvironment by stabilizing p300 and β-TrCP and promotes cancer malignancy

doi: 10.1038/s41467-018-06178-1

Figure Lengend Snippet: Schematic summary of the role of USP24 in IL-6 mediated metastasis and angiogenesis. USP24 promotes IL-6 expression by increasing the levels of NF-κB and histone H3 acetylation through stabilizes p300 in the M2 macrophages. In lung cancer cells, USP24 increases p300 and β-TrCP levels which promotes histone H3 acetylation, NF-κB nuclear translocation and decreases IL-6 promoter methylation, thus resulting in the upregulation of IL-6 level in tumor-associated microenvironment and lung cancer malignancy

Article Snippet: A549 cells were overexpressed with myc-ubi and treated MG132 (Sigma-Aldrich) for 12 h. β-TrCP protein in A549 cell lysates were immunoprecipitated by incubating with anti-β-TrCP antibody for 4 h, and then incubated with protein A agarose (Millipore) for 1 h. After washing, substrate was mixed with human recombinant USP24 protein (50 μg per ml) (Origene) in deubiquitination buffer (50 mM Tris PH 8.0, 10 mM DTT and 5 μM MG132) for 2 h in 37 °C.

Techniques: Expressing, Translocation Assay, Methylation

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: SNPs Analyzed [Note]

Article Snippet: 227 , rs1165226 , {"type":"entrez-nucleotide","attrs":{"text":"AK127075","term_id":"34533820","term_text":"AK127075"}} AK127075 , C_11732134_10 , 53895603 , 54977923 , 38.1 , .457 , .708.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Results of single-locus association tests of AAO in PD. Two methods were used to assess association with AAO in the overall PD data set: the MKM (triangles) and the OM (diamonds). The SNP numbers of the significant polymorphisms (i.e., those with P ⩽.01) are indicated. The candidate genes are EIF2B3, TESK2, FLJ14442, ELAVL4, C1orf8, USP24, and AK127075.

Article Snippet: 227 , rs1165226 , {"type":"entrez-nucleotide","attrs":{"text":"AK127075","term_id":"34533820","term_text":"AK127075"}} AK127075 , C_11732134_10 , 53895603 , 54977923 , 38.1 , .457 , .708.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Characterization of USP24L, with mRNA and predicted protein sequence of the USP24L transcript. The protein sequence in bold corresponds to the overlap with AK127075, and the underlined sequence matches the USP24 protein sequence. The DNA sequence underlined and in bold corresponds to the two additional exons of USP24L, in comparison with XM_371254.

Article Snippet: 227 , rs1165226 , {"type":"entrez-nucleotide","attrs":{"text":"AK127075","term_id":"34533820","term_text":"AK127075"}} AK127075 , C_11732134_10 , 53895603 , 54977923 , 38.1 , .457 , .708.

Techniques: Sequencing

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Summary of P Values Obtained by the OM and MKM for SNPs in EIF2B3 and USP24, in the Overall, Positive-Linkage, and Negative-Linkage Data Sets [Note]

Article Snippet: 227 , rs1165226 , {"type":"entrez-nucleotide","attrs":{"text":"AK127075","term_id":"34533820","term_text":"AK127075"}} AK127075 , C_11732134_10 , 53895603 , 54977923 , 38.1 , .457 , .708.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Summary of Haplotypes Showing Significant Association with AAO in the Overall PD Data Set [Note]

Article Snippet: 227 , rs1165226 , {"type":"entrez-nucleotide","attrs":{"text":"AK127075","term_id":"34533820","term_text":"AK127075"}} AK127075 , C_11732134_10 , 53895603 , 54977923 , 38.1 , .457 , .708.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: SNPs Analyzed [Note]

Article Snippet: 225 , rs567734 , {"type":"entrez-nucleotide","attrs":{"text":"AK127075","term_id":"34533820","term_text":"AK127075"}} AK127075 , C__998713_10 , 53861957 , 54944282 , 18.8 , .830 , .335.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Results of single-locus association tests of AAO in PD. Two methods were used to assess association with AAO in the overall PD data set: the MKM (triangles) and the OM (diamonds). The SNP numbers of the significant polymorphisms (i.e., those with P ⩽.01) are indicated. The candidate genes are EIF2B3, TESK2, FLJ14442, ELAVL4, C1orf8, USP24, and AK127075.

Article Snippet: 225 , rs567734 , {"type":"entrez-nucleotide","attrs":{"text":"AK127075","term_id":"34533820","term_text":"AK127075"}} AK127075 , C__998713_10 , 53861957 , 54944282 , 18.8 , .830 , .335.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Characterization of USP24L, with mRNA and predicted protein sequence of the USP24L transcript. The protein sequence in bold corresponds to the overlap with AK127075, and the underlined sequence matches the USP24 protein sequence. The DNA sequence underlined and in bold corresponds to the two additional exons of USP24L, in comparison with XM_371254.

Article Snippet: 225 , rs567734 , {"type":"entrez-nucleotide","attrs":{"text":"AK127075","term_id":"34533820","term_text":"AK127075"}} AK127075 , C__998713_10 , 53861957 , 54944282 , 18.8 , .830 , .335.

Techniques: Sequencing

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Summary of P Values Obtained by the OM and MKM for SNPs in EIF2B3 and USP24, in the Overall, Positive-Linkage, and Negative-Linkage Data Sets [Note]

Article Snippet: 225 , rs567734 , {"type":"entrez-nucleotide","attrs":{"text":"AK127075","term_id":"34533820","term_text":"AK127075"}} AK127075 , C__998713_10 , 53861957 , 54944282 , 18.8 , .830 , .335.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Summary of Haplotypes Showing Significant Association with AAO in the Overall PD Data Set [Note]

Article Snippet: 225 , rs567734 , {"type":"entrez-nucleotide","attrs":{"text":"AK127075","term_id":"34533820","term_text":"AK127075"}} AK127075 , C__998713_10 , 53861957 , 54944282 , 18.8 , .830 , .335.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: SNPs Analyzed [Note]

Article Snippet: 222 , rs667353 , USP24 , C_11289191_1_ , 53845130 , 54927458 , 36.8 , .880 , 1.000.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Results of single-locus association tests of AAO in PD. Two methods were used to assess association with AAO in the overall PD data set: the MKM (triangles) and the OM (diamonds). The SNP numbers of the significant polymorphisms (i.e., those with P ⩽.01) are indicated. The candidate genes are EIF2B3, TESK2, FLJ14442, ELAVL4, C1orf8, USP24, and AK127075.

Article Snippet: 222 , rs667353 , USP24 , C_11289191_1_ , 53845130 , 54927458 , 36.8 , .880 , 1.000.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Characterization of USP24L, with mRNA and predicted protein sequence of the USP24L transcript. The protein sequence in bold corresponds to the overlap with AK127075, and the underlined sequence matches the USP24 protein sequence. The DNA sequence underlined and in bold corresponds to the two additional exons of USP24L, in comparison with XM_371254.

Article Snippet: 222 , rs667353 , USP24 , C_11289191_1_ , 53845130 , 54927458 , 36.8 , .880 , 1.000.

Techniques: Sequencing

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Summary of P Values Obtained by the OM and MKM for SNPs in EIF2B3 and USP24, in the Overall, Positive-Linkage, and Negative-Linkage Data Sets [Note]

Article Snippet: 222 , rs667353 , USP24 , C_11289191_1_ , 53845130 , 54927458 , 36.8 , .880 , 1.000.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Summary of Haplotypes Showing Significant Association with AAO in the Overall PD Data Set [Note]

Article Snippet: 222 , rs667353 , USP24 , C_11289191_1_ , 53845130 , 54927458 , 36.8 , .880 , 1.000.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: SNPs Analyzed [Note]

Article Snippet: 230 , rs287235 , … , C__686425_10 , 53966079 , 55048417 , 23.0 , 1.000 , .735.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Results of single-locus association tests of AAO in PD. Two methods were used to assess association with AAO in the overall PD data set: the MKM (triangles) and the OM (diamonds). The SNP numbers of the significant polymorphisms (i.e., those with P ⩽.01) are indicated. The candidate genes are EIF2B3, TESK2, FLJ14442, ELAVL4, C1orf8, USP24, and AK127075.

Article Snippet: 230 , rs287235 , … , C__686425_10 , 53966079 , 55048417 , 23.0 , 1.000 , .735.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Characterization of USP24L, with mRNA and predicted protein sequence of the USP24L transcript. The protein sequence in bold corresponds to the overlap with AK127075, and the underlined sequence matches the USP24 protein sequence. The DNA sequence underlined and in bold corresponds to the two additional exons of USP24L, in comparison with XM_371254.

Article Snippet: 230 , rs287235 , … , C__686425_10 , 53966079 , 55048417 , 23.0 , 1.000 , .735.

Techniques: Sequencing

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Summary of P Values Obtained by the OM and MKM for SNPs in EIF2B3 and USP24, in the Overall, Positive-Linkage, and Negative-Linkage Data Sets [Note]

Article Snippet: 230 , rs287235 , … , C__686425_10 , 53966079 , 55048417 , 23.0 , 1.000 , .735.

Techniques:

Journal:

Article Title: Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

doi:

Figure Lengend Snippet: Summary of Haplotypes Showing Significant Association with AAO in the Overall PD Data Set [Note]

Article Snippet: 230 , rs287235 , … , C__686425_10 , 53966079 , 55048417 , 23.0 , 1.000 , .735.

Techniques:

Journal: Cell chemical biology

Article Title: A Rapid and Precise Mutation-Activated Fluorescence Reporter for Analyzing Acute Mutagenesis Frequency

doi: 10.1016/j.chembiol.2018.05.010

Figure Lengend Snippet: (A) Representative red-green flow diagrams of HCT116 cells carrying CherryOFF-GFP and USP24 KD or nonsilencing shRNA control, exposed to 10 or 40 J/m2 UV radiation, or untreated. UV irradiation increased mutagenesis in both cell lines, and higher mutagenesis was seen in USP24 KD.

Article Snippet: USP24 knockdown efficiency was quantified via western blot using a rabbit-anti-USP24 antibody (Origene TA5240) compared to tubulin control (Santa Cruz H-235) Genomic DNA Purification Genomic DNA was extracted from cells with Cyclo-Prep Genomic DNA isolation kit (Amresco) and proteinase K (New England Biolabs).

Techniques: shRNA, Irradiation, Mutagenesis

Journal: Cell chemical biology

Article Title: A Rapid and Precise Mutation-Activated Fluorescence Reporter for Analyzing Acute Mutagenesis Frequency

doi: 10.1016/j.chembiol.2018.05.010

Figure Lengend Snippet: (A) USP24 KD HCT116 cells treated with 10 J/m2 UV, and ATE1-KO MEF cells treated with 40 J/m2, were sorted and cultured. Representative picture shown with arrow indicating one cell exhibiting only GFP signal in the sorted HCT116 cells. Scale bar represents 100 µm. As shown in the table (right), we found less than 1% green-only cells in HCT cells that were sorted for red fluorescence, which may be due to sticky cell doublets during FACS. In the less self-adhesive MEF cells, a much lower percentage of green-only cells was found in the CherryOFF-GFP carrying cells that were sorted for red signal. Mean cell numbers are shown (±SEM) from three independent experiments.

Article Snippet: USP24 knockdown efficiency was quantified via western blot using a rabbit-anti-USP24 antibody (Origene TA5240) compared to tubulin control (Santa Cruz H-235) Genomic DNA Purification Genomic DNA was extracted from cells with Cyclo-Prep Genomic DNA isolation kit (Amresco) and proteinase K (New England Biolabs).

Techniques: Cell Culture, Fluorescence

Journal: Cell chemical biology

Article Title: A Rapid and Precise Mutation-Activated Fluorescence Reporter for Analyzing Acute Mutagenesis Frequency

doi: 10.1016/j.chembiol.2018.05.010

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: USP24 knockdown efficiency was quantified via western blot using a rabbit-anti-USP24 antibody (Origene TA5240) compared to tubulin control (Santa Cruz H-235) Genomic DNA Purification Genomic DNA was extracted from cells with Cyclo-Prep Genomic DNA isolation kit (Amresco) and proteinase K (New England Biolabs).

Techniques: shRNA, Recombinant, Isolation, SYBR Green Assay, Software

Journal: Frontiers in Molecular Neuroscience

Article Title: Proteomic analysis of spinal cord tissue in a rat model of cancer-induced bone pain

doi: 10.3389/fnmol.2022.1009615

Figure Lengend Snippet: The information of up and down-regulated differentially expressed mitochondrial proteins.

Article Snippet: After the deparaffinization and rehydration, the spinal cord sections were immersed in citrate antigen retrieval solution (Beyotime, China) for antigen repair, incubated with 3% hydrogen peroxide for 10 min blocked with Immunol Staining Blocking Buffer (Beyotime, China) for 60 min, incubated with anti-SNAP25 rabbit pAb (A2234, ABclonal), GATM rabbit pAb (1:1000, A6598), and NDUFA11 rabbit pAb (1:1000, A16239) overnight at 4°C and goat anti-rabbit IgG H&L (FITC) (ab6717, USA) at room temperature for 60 min.

Techniques:

Journal: Frontiers in Molecular Neuroscience

Article Title: Proteomic analysis of spinal cord tissue in a rat model of cancer-induced bone pain

doi: 10.3389/fnmol.2022.1009615

Figure Lengend Snippet: Validation of synaptic- and mitochondrial-related proteins using immunofluorescence and Western blot assay. (A,B) Representative immunofluorescence staining images (A) and quantitative analysis (B) for SNAP25, GATM, and NDUFA11 in the spinal cord. Scale bar = 20 μm. Data were expressed as the mean ± SD ( n = 3 mice/group). * p < 0.05 vs. sham group. (C,D) Representative Western blot bands (C) and quantitative analysis (D) of CPLX1, SNAP25, and SYT1 protein in the spinal cord. Data were presented as mean ± SD ( n = 3 mice/group). * p < 0.05 vs. sham group. (E,F) Representative western blot bands (E) and quantitative analysis (F) of ALDH1B, GATM, and NDUFA11 protein in spinal cord. Data were presented as mean ± SD ( n = 3 mice/group). * p < 0.05 vs. sham group.

Article Snippet: After the deparaffinization and rehydration, the spinal cord sections were immersed in citrate antigen retrieval solution (Beyotime, China) for antigen repair, incubated with 3% hydrogen peroxide for 10 min blocked with Immunol Staining Blocking Buffer (Beyotime, China) for 60 min, incubated with anti-SNAP25 rabbit pAb (A2234, ABclonal), GATM rabbit pAb (1:1000, A6598), and NDUFA11 rabbit pAb (1:1000, A16239) overnight at 4°C and goat anti-rabbit IgG H&L (FITC) (ab6717, USA) at room temperature for 60 min.

Techniques: Immunofluorescence, Western Blot, Staining