Journal: Leukemia

Article Title: Distinct characteristics of VEXAS-causative UBA1 M41 and recurrent functional non-M41 mutations

doi: 10.1038/s41375-025-02775-4

Figure Lengend Snippet: A Schematic representation of the UBA1 gene, highlighting its functional domains and pinpointing the exact locations of the identified UBA1 variants. The variants were classified into Tier-1 (pathogenic: P), Tier-2 (likely pathogenic: LP) and Tier-3 (variant of uncertain significance: VUS), as depicted in the outer pie chart on the left. UBA1 variants are displayed on top of the gene if the locus occurred recurrently in our screen. Combination of variants detected in the same patients are also shown. Variants detected only as combinations are italicized. M41 and other tier assignments are color-coded. Bold underlined variants were tested for functional significance. B Chart summarizing UBA1 variants and their functional status, indicating the presence or absence of ubiquitylation. Quantification of polyubiquitin levels and mono-ubiquitylated histone H2A/B (Ub-H2A, Ub-H2B) or E2 enzymes (UBE2D3-Ub, UBE2L3-Ub) were normalized within each sample to β-actin levels and scaled to WT transfection. E2 enzyme ubiquitylation was quantified as the ratio of charged form to uncharged form and scaled to WT. Data represent n = 3–6 biological replicates, shown as mean −/+ s.d., significance determined by unpaired t-test with Welch’s correction (*p < 0.05, **p < 0.01).

Article Snippet: Primary antibodies for Poly-ubiquitin (Cell Signaling, 3936S), UBE2D3 (Cell-Signaling, 4330), UBE2L3 (R&D Systems, E2-640), ubiquitylated-H2A (Cell Signaling, 8240), H2A (Cell Signaling, 12349), ubiquitylated-H2B (Cell Signaling, 5546S), H2B (Cell Signaling, 12364), and β-actin (Cell Signaling, 4970) were used at a concentration of 1:1000 and visualized using HRP-conjugated secondary antibodies (anti-rabbit [Cell Signaling, 7074S] or anti-mouse [Cell Signaling, 7076S]) at a concentration of 1:3000.

Techniques: Functional Assay, Variant Assay, Transfection

Journal: American Journal of Human Genetics

Article Title: UBE2L3 Polymorphism Amplifies NF-κB Activation and Promotes Plasma Cell Development, Linking Linear Ubiquitination to Multiple Autoimmune Diseases

doi: 10.1016/j.ajhg.2014.12.024

Figure Lengend Snippet: rs140490 Is Associated with SLE and a Cell-Specific Increase in UBE2L3 Production (A) Locuszoom plot showing SNPs around UBE2L3 imputed to 1000 Genomes level. Recombination rate was calculated from HapMap data. (B) rs140490 is associated with increased expression in microarray data (expressed as log base 2, bars show median and interquartile range) from EBV-transformed lymphoblastoid cell lines, as well as increased mRNA expression in CD19 + B cells and CD14 + monocytes isolated from PBMCs. (C) Semiquantitative analysis of UBE2L3 protein level from immunoblot densitometry compared to actin, stratified by rs140490 genotype in CD19 + B cells and CD4 + T cells isolated from PBMCs from healthy individuals, with representative immunoblots shown from individuals for each rs140490 genotype.

Article Snippet: The following plasmids on the pCMV6 vector were purchased from Origene: untagged UBE2L3, HOIP, UBE2D1, UBE2D2, UBE2D3, Sharpin-Myc-DDK, and empty vector (pCMV6-AC). pCMV6-HOIL-1-Myc-DDK (Origene) was subcloned into pCMV6AC to remove the C terminus Myc-DDK tag. pcDNA3.1-V5-His-HOIP and pcDNA3.1-V5-His-HOIP (p.Cys885Ser) were kind gifts of H. Walczak. pFlagCMV2-UbcH8 (UBE2L6) was obtained from Addgene.

Techniques: Expressing, Microarray, Transformation Assay, Isolation, Western Blot

Journal: American Journal of Human Genetics

Article Title: UBE2L3 Polymorphism Amplifies NF-κB Activation and Promotes Plasma Cell Development, Linking Linear Ubiquitination to Multiple Autoimmune Diseases

doi: 10.1016/j.ajhg.2014.12.024

Figure Lengend Snippet: UBE2L3 Exerts Rate-Limiting Control over LUBAC-Mediated NF-κB Activation (A) Basal NF-κB luciferase activity in HEK293 cells in response to increased production of combinations of HOIL-1, HOIP, Sharpin, and UBE2L3. UBE2L3 enhances NF-κB activation due to increased levels of HOIL-1 and HOIP. Error bars represent standard error of the mean. (B) Dominant-negative variant p.Cys86Ser UBE2L3 suppresses NF-κB activity due to LUBAC. (C) UBE2L3 is the preferred E2 ubiquitin-conjugating enzyme for LUBAC in vivo. Luciferase assay time course over 48 hr in HEK293-NF-κB reporter cells transiently transfected different E2 ubiquitin-conjugating enzymes in combination with EV, HOIP alone, HOIL-1+HOIP, or Sharpin+HOIP, comparing the basal NF-κB response after increased production of individual E2 enzymes with LUBAC. (D and E) Comparison of NF-κB p65 translocation measured by Imagestream analysis in HEK293 cells transiently transfected with empty vector (EV) or HOIL-1+V5-HOIP+UBE2L3. p65 nuclear translocation is quantified by Imagestream similarity feature correlating fluorescence co-localization of AlexaFluor488-p65 with nuclear DAPI. Similarity histograms show levels of p65 translocation in EV, compared to unstimulated HEK293 cells transfected with HOIL-1+HOIP+UBE2L3. EV-transfected cells stimulated with TNF 10 ng/ml for 1 hr acted as a positive control. AlexaFluor647 anti-V5-tag and PE anti-UBE2L3 were used to gate cells with high levels of HOIP(V5) and UBE2L3, showing high p65 translocation in this group compared to cells with normal HOIP and UBE2L3 levels. Representative histograms are shown from one of three separate experiments. (D) Representative Imagestream images of HEK293 cells with median similarity level of p65 translocation as measured in (E) showing AlexaFluor488-p65 (green), DAPI nuclear dye (pseudocolored red), PE-UBE2L3 (yellow), and AlexaFluor647-V5(HOIP) (purple). Merged images show p65/DAPI overlap in yellow.

Article Snippet: The following plasmids on the pCMV6 vector were purchased from Origene: untagged UBE2L3, HOIP, UBE2D1, UBE2D2, UBE2D3, Sharpin-Myc-DDK, and empty vector (pCMV6-AC). pCMV6-HOIL-1-Myc-DDK (Origene) was subcloned into pCMV6AC to remove the C terminus Myc-DDK tag. pcDNA3.1-V5-His-HOIP and pcDNA3.1-V5-His-HOIP (p.Cys885Ser) were kind gifts of H. Walczak. pFlagCMV2-UbcH8 (UBE2L6) was obtained from Addgene.

Techniques: Control, Activation Assay, Luciferase, Activity Assay, Dominant Negative Mutation, Variant Assay, Ubiquitin Proteomics, In Vivo, Transfection, Comparison, Translocation Assay, Plasmid Preparation, Fluorescence, Positive Control

Journal: American Journal of Human Genetics

Article Title: UBE2L3 Polymorphism Amplifies NF-κB Activation and Promotes Plasma Cell Development, Linking Linear Ubiquitination to Multiple Autoimmune Diseases

doi: 10.1016/j.ajhg.2014.12.024

Figure Lengend Snippet: UBE2L3 and LUBAC Augment Late-Phase NF-κB Activation in Response to TNF (A) Increased production of LUBAC in combination with UBE2L3 augments NF-κB activation after 10 hr of TNF stimulation. This augmentation of the TNF response is abrogated by variant p.Cys86Ser UBE2L3 or variant p.Cys885Ser HOIP. (B) Time course showing NF-κB activation after TNF stimulation in response to increased synthesis of LUBAC and UBE2L3. Maximal dysregulation of NF-κB occurs in the late-phase response to TNF. Error bars represent standard error of the mean.

Article Snippet: The following plasmids on the pCMV6 vector were purchased from Origene: untagged UBE2L3, HOIP, UBE2D1, UBE2D2, UBE2D3, Sharpin-Myc-DDK, and empty vector (pCMV6-AC). pCMV6-HOIL-1-Myc-DDK (Origene) was subcloned into pCMV6AC to remove the C terminus Myc-DDK tag. pcDNA3.1-V5-His-HOIP and pcDNA3.1-V5-His-HOIP (p.Cys885Ser) were kind gifts of H. Walczak. pFlagCMV2-UbcH8 (UBE2L6) was obtained from Addgene.

Techniques: Activation Assay, Variant Assay

Journal: American Journal of Human Genetics

Article Title: UBE2L3 Polymorphism Amplifies NF-κB Activation and Promotes Plasma Cell Development, Linking Linear Ubiquitination to Multiple Autoimmune Diseases

doi: 10.1016/j.ajhg.2014.12.024

Figure Lengend Snippet: Inhibition of UBE2L3 Regulates NF-κB Signaling and Target Gene Transcription (A) Luciferase assay in HEK293-NF-κB reporter cell line showing that siRNA inhibition of HOIL-1, HOIP, Sharpin, and UBE2L3 inhibits NF-κB activation in response to TNF. Error bars represent standard error of the mean. (B) Immunoblot showing siRNA knockdown of HOIL-1, HOIP, Sharpin, and UBE2L3. (C) Inhibition of UBE2L3 or HOIP in HEK293 cells leads to basal increase in IκBα levels and impaired phosphorylation of IκBα in response to TNF, and subsequent IκBα degradation is reduced. UBE2L3 does not impair JNK or ERK phosphorylation. (D) Inhibition of UBE2L3 or HOIP reduces transcription of NF-κB target genes measured by real-time qPCR. HEK293 cells transfected with control, UBE2L3, or HOIP siRNA were stimulated with 10 ng/ml TNF.

Article Snippet: The following plasmids on the pCMV6 vector were purchased from Origene: untagged UBE2L3, HOIP, UBE2D1, UBE2D2, UBE2D3, Sharpin-Myc-DDK, and empty vector (pCMV6-AC). pCMV6-HOIL-1-Myc-DDK (Origene) was subcloned into pCMV6AC to remove the C terminus Myc-DDK tag. pcDNA3.1-V5-His-HOIP and pcDNA3.1-V5-His-HOIP (p.Cys885Ser) were kind gifts of H. Walczak. pFlagCMV2-UbcH8 (UBE2L6) was obtained from Addgene.

Techniques: Inhibition, Luciferase, Activation Assay, Western Blot, Knockdown, Phospho-proteomics, Transfection, Control

Journal: American Journal of Human Genetics

Article Title: UBE2L3 Polymorphism Amplifies NF-κB Activation and Promotes Plasma Cell Development, Linking Linear Ubiquitination to Multiple Autoimmune Diseases

doi: 10.1016/j.ajhg.2014.12.024

Figure Lengend Snippet: UBE2L3 Genotype Affects Basal and Stimulated NF-κB Translocation in Primary Human B Cells and Monocytes Translocation of NF-κB p65 was quantified by similarity score on Imagestream analysis of CD19 + B cells and CD14 + monocytes isolated from PBMCs from healthy individuals, stratified by genotype at rs140490. (A) Representative similarity histograms measuring co-localization of p65 and nuclear dye DRAQ5 for B cells after 30 min of stimulation with CD40L and for monocytes after 30 min of TNF stimulation for each rs140490 genotype. Translocated cells were defined as similarity score > 1.5. (B) Imagestream images for each rs140490 genotype showing CD40L-stimulated B cells and TNF-stimulated monocytes of median similarity levels of p65 translocation from histograms in (A) showing p65 (Alexa488), DRAQ5 nuclear dye, and merged images. (C) Graphs show percent of NF-κB p65 nuclear translocated cells in unstimulated CD19 + B cells and CD14 + monocytes from genotyped individuals. (D) Graphs show 60 min time course of p65 translocation in response to stimulation of B cells with 0.1 μg/ml CD40L and monocytes with 10 ng/ml TNF, stratified by rs140490 genotype. Error bars represent standard error of the mean.

Article Snippet: The following plasmids on the pCMV6 vector were purchased from Origene: untagged UBE2L3, HOIP, UBE2D1, UBE2D2, UBE2D3, Sharpin-Myc-DDK, and empty vector (pCMV6-AC). pCMV6-HOIL-1-Myc-DDK (Origene) was subcloned into pCMV6AC to remove the C terminus Myc-DDK tag. pcDNA3.1-V5-His-HOIP and pcDNA3.1-V5-His-HOIP (p.Cys885Ser) were kind gifts of H. Walczak. pFlagCMV2-UbcH8 (UBE2L6) was obtained from Addgene.

Techniques: Translocation Assay, Isolation

Journal: American Journal of Human Genetics

Article Title: UBE2L3 Polymorphism Amplifies NF-κB Activation and Promotes Plasma Cell Development, Linking Linear Ubiquitination to Multiple Autoimmune Diseases

doi: 10.1016/j.ajhg.2014.12.024

Figure Lengend Snippet: UBE2L3 Is Highly Abundant in Peripheral Blood Plasmablasts and Plasma Cells PBMCs isolated from healthy individuals and SLE-affected individuals were subject to ten-color flow cytometry to analyze UBE2L3 levels in B cell subsets. (A) Gating strategy for B cell subsets. (B) Histograms of B cell subsets from a representative SLE-affected individual demonstrating clear shift in UBE2L3 protein levels in CD19 mid CD20 − CD27 hi CD38 hi IgD − plasmablast and CD20 − CD27 hi CD38 hi IgD − CD138 hi plasma cell populations (isotype control light gray). (C) UBE2L3 abundance was highly increased in plasmablasts and plasma cells compared to other B cell subsets in both SLE-affected individuals and controls. (D and E) UBE2L3 levels were significantly higher in (D) Ki-67 + proliferating B cells and (E) CD95 + activated B cells in SLE-affected individuals. Histograms: isotype control, light gray; blue, Ki-67 − or CD95 − ; red, Ki-67 + or CD95 + . Error bars represent standard error of the mean.

Article Snippet: The following plasmids on the pCMV6 vector were purchased from Origene: untagged UBE2L3, HOIP, UBE2D1, UBE2D2, UBE2D3, Sharpin-Myc-DDK, and empty vector (pCMV6-AC). pCMV6-HOIL-1-Myc-DDK (Origene) was subcloned into pCMV6AC to remove the C terminus Myc-DDK tag. pcDNA3.1-V5-His-HOIP and pcDNA3.1-V5-His-HOIP (p.Cys885Ser) were kind gifts of H. Walczak. pFlagCMV2-UbcH8 (UBE2L6) was obtained from Addgene.

Techniques: Clinical Proteomics, Isolation, Flow Cytometry, Control

Journal: American Journal of Human Genetics

Article Title: UBE2L3 Polymorphism Amplifies NF-κB Activation and Promotes Plasma Cell Development, Linking Linear Ubiquitination to Multiple Autoimmune Diseases

doi: 10.1016/j.ajhg.2014.12.024

Figure Lengend Snippet: UBE2L3 Genotype Influences Peripheral Blood Plasmablast Proliferation in SLE (A) The UBE2L3 risk allele (rs140490 genotype) was associated with a significant expansion of peripheral blood plasmablasts and plasma cells (expressed as percent of live B cells) from SLE-affected individuals (n = 29), whereas this expansion was not seen in healthy controls (n = 25). Error bars represent standard error of the mean. (B) Representative flow cytometry plots showing percentages of plasmablasts and plasma cells (expressed as percent of live B cells) from SLE-affected individuals for each homozygous genotype.

Article Snippet: The following plasmids on the pCMV6 vector were purchased from Origene: untagged UBE2L3, HOIP, UBE2D1, UBE2D2, UBE2D3, Sharpin-Myc-DDK, and empty vector (pCMV6-AC). pCMV6-HOIL-1-Myc-DDK (Origene) was subcloned into pCMV6AC to remove the C terminus Myc-DDK tag. pcDNA3.1-V5-His-HOIP and pcDNA3.1-V5-His-HOIP (p.Cys885Ser) were kind gifts of H. Walczak. pFlagCMV2-UbcH8 (UBE2L6) was obtained from Addgene.

Techniques: Clinical Proteomics, Flow Cytometry

Journal: Journal of Virology

Article Title: The membrane-associated ubiquitin ligase MARCHF8 stabilizes the human papillomavirus oncoprotein E7 by degrading CUL1 and UBE2L3 in head and neck cancer

doi: 10.1128/jvi.01726-23

Figure Lengend Snippet: CUL1 and UBE2L3 overexpression increases the ubiquitination and decreases HPV16 E7 protein level in HPV+ HNC cells. ( A ) The schematic model of HPV16 E7 degradation through CUL1- and UBE2L3-mediated ubiquitination. Figure generated with a Biorender icon. CUL1 or UBE2L3 is overexpressed in HPV+ HNC (SCC152) ( B–C ) and SCC2 ( D–E ) cells using lentiviral transduction. CUL1, UBE2L3, HPV16 E7, and MARCHF8 proteins were detected by western blotting. The relative band intensities were quantified using NIH ImageJ in SCC152 ( C ) and SCC2 ( E ). β-Actin was used as an internal control. ( F ) SCC152 cells with CUL1 or UBE2L3 overexpression were treated with MG132 (10 µM). Ubiquitinated proteins were pulled down from the cell lysate using anti-ubiquitin antibody-conjugated magnetic beads, and HPV16 E7 protein was detected by western blotting. The data shown are means ± SD of three independent experiments. Student’s t -test was used to determine P -values. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: UBE2L3 , Human/Mouse , Proteintech , 14115–1-AP , AB_2210891 , IP-WB.

Techniques: Over Expression, Ubiquitin Proteomics, Generated, Transduction, Western Blot, Control, Magnetic Beads

Journal: Journal of Virology

Article Title: The membrane-associated ubiquitin ligase MARCHF8 stabilizes the human papillomavirus oncoprotein E7 by degrading CUL1 and UBE2L3 in head and neck cancer

doi: 10.1128/jvi.01726-23

Figure Lengend Snippet: CUL1 and UBE2L3 protein levels are low in HPV+ HNC cells. CUL1 and UBE2L3 protein levels in normal (N/Tert-1), HPV- HNC (SCC1, SCC9, and SCC19), and HPV+ HNC (SCC2, SCC90, and SCC152) cells were determined by western blotting ( A ). Relative band intensities were quantified using NIH ImageJ ( B ). HPV16 E7 and β-actin were used as viral and internal controls, respectively. CUL1 and UBE2L3 mRNA expression levels in normal (N/Tert-1), HPV+ HNC (SCC2, SCC90, and SCC152), and HPV− HNC (SCC1, SCC9, and SCC19) cells were quantified by RT-qPCR ( C ). The data shown are normalized by the glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNA level as an internal control. The mRNA expression levels of CUL1 and UBE2L3 were analyzed using our previous gene expression data (GSE6791) and shown as fluorescence intensity (log2) from microdissected human tissue samples from HPV+ ( n = 16) and HPV− ( n = 26) HNC patients and normal individuals ( n = 12) ( D ). CUL1, UBE2L3, and E7 protein levels in SCC152 cells treated with MG132 (10 µM) or dimethyl sulfoxide (DMSO) ( E ). Relative band intensities were quantified using NIH ImageJ ( F ). CUL1 and UBE2L3 proteins were detected in N/Tert-1 cells expressing HPV16 E6, E7, or E6 and E7 by western blotting ( G ). The size of HPV16 E7 in N/Tert-1 E7 cells is about 22 kDa due to HA tagging, while the size of native HPV16 E7 in N/Tert-1 E6E7 cells is about 17 kDa. Relative band intensities were quantified using NIH ImageJ ( H ). Total RNA was extracted from N/Tert-1 containing an empty vector and N/Tert-1 cells expressing HPV16 E6, E7, or E6 and E7 (E6E7). The HPV16 E6 and E7 mRNA levels were quantified by RT-qPCR ( I ). The data shown are normalized by the GAPDH mRNA level as an internal control. All experiments were repeated at least three times, and the data shown are means ± SD of three independent experiments. Student’s t -test determined P -values. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: UBE2L3 , Human/Mouse , Proteintech , 14115–1-AP , AB_2210891 , IP-WB.

Techniques: Western Blot, Expressing, Quantitative RT-PCR, Control, Gene Expression, Fluorescence, Plasmid Preparation

Journal: Journal of Virology

Article Title: The membrane-associated ubiquitin ligase MARCHF8 stabilizes the human papillomavirus oncoprotein E7 by degrading CUL1 and UBE2L3 in head and neck cancer

doi: 10.1128/jvi.01726-23

Figure Lengend Snippet: Knockdown of MARCHF8 expression increases CUL1 and UBE2L3 protein levels and decreases HPV16 E7 protein levels in HPV+ HNC cells. SCC152 (A and C) and SCC2 (B and D) cells were transduced with five and three lentiviral shR-MARCHF8, respectively, or shR-scr as a control. MARCHF8, CUL1, UBE2L3, HPV16 E7, and pRb proteins in SCC152 ( A ) and SCC2 ( B ) cells were detected by western blotting. The relative band intensities were quantified using NIH ImageJ ( C and D ). β-Actin was used as an internal control. The mRNA levels of CUL1, UBE2L3, and HPV16 E7 were quantified by RT-qPCR in SCC152 ( E ) and SCC2 ( F ) cells transduced with five and three lentiviral shR-MARCHF8, respectively, or shR-scr as a control. The data shown are normalized by the GAPDH mRNA level as an internal control. The data shown are means ± SD of three independent experiments. Student’s t -test determined P -values. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: UBE2L3 , Human/Mouse , Proteintech , 14115–1-AP , AB_2210891 , IP-WB.

Techniques: Knockdown, Expressing, Transduction, Control, Western Blot, Quantitative RT-PCR

Journal: Journal of Virology

Article Title: The membrane-associated ubiquitin ligase MARCHF8 stabilizes the human papillomavirus oncoprotein E7 by degrading CUL1 and UBE2L3 in head and neck cancer

doi: 10.1128/jvi.01726-23

Figure Lengend Snippet: MARCHF8 protein interacts with and ubiquitinates CUL1 and UBE2L3 proteins. MARCHF8 ( A ), CUL1 ( B ), and UBE2L3 ( C ) were pulled down from the lysate of SCC-152 cells treated with MG132 (10 µM) using anti-MARCHF8 ( A ), anti-CUL1 ( B ), and anti-UBE2L3 ( C ) antibodies, respectively. CUL1, UBE2L3, HPV16 E7, and MARCHF8 were detected from the immunoprecipitated proteins by western blotting. ( D–F ) Ubiquitinated proteins were pulled down from the lysate of SCC152 cells with shR-scr or shRNA against MARCHF8 (shR-MARCHF8 clone 3) treated with MG132 (10 µM) using anti-ubiquitin antibody-conjugated magnetic beads. CUL1 ( D ), UBE2L3 ( E ), and HPV16 E7 ( F ) proteins were detected in the immunoprecipitated proteins by western blotting. All experiments were repeated at least three times. The data shown are means ± SD of three independent experiments. Student’s t -test determined P -values. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: UBE2L3 , Human/Mouse , Proteintech , 14115–1-AP , AB_2210891 , IP-WB.

Techniques: Immunoprecipitation, Western Blot, shRNA, Ubiquitin Proteomics, Magnetic Beads

Journal: Journal of Virology

Article Title: The membrane-associated ubiquitin ligase MARCHF8 stabilizes the human papillomavirus oncoprotein E7 by degrading CUL1 and UBE2L3 in head and neck cancer

doi: 10.1128/jvi.01726-23

Figure Lengend Snippet: Marchf8 knockout restores CUL1 and UBE2L3 protein levels in HPV+ mouse oral cancer cells. CUL1 and UBE2L3 protein levels in mouse normal immortalized (NiMOE), HPV− transformed (HPV− MOE), and HPV+ transformed (mEERL) oral epithelial cells were determined by western blotting ( A ). β-actin was used as an internal control. The relative band intensities were quantified using NIH ImageJ ( B ). mEERL cells were transduced with lentiviruses containing Cas9 and one of two sgRNAs against Marchf8 (sgR- Marchf 8-2 and sgR- Marchf 8-3) or scrambled sgRNA (sgR-scr). MARCHF8, CUL1, UBE2L3, and HPV16 E7 proteins were detected by western blotting ( C ). Relative band intensities were quantified using NIH ImageJ ( D ). The data shown are means ± SD of three independent experiments. All experiments were repeated at least three times, and the data shown are means ± SD. Student’s t -test determined P -values. * P < 0.05, *** P < 0.001.

Article Snippet: UBE2L3 , Human/Mouse , Proteintech , 14115–1-AP , AB_2210891 , IP-WB.

Techniques: Knock-Out, Transformation Assay, Western Blot, Control, Transduction

Journal: Journal of Virology

Article Title: The membrane-associated ubiquitin ligase MARCHF8 stabilizes the human papillomavirus oncoprotein E7 by degrading CUL1 and UBE2L3 in head and neck cancer

doi: 10.1128/jvi.01726-23

Figure Lengend Snippet: CUL1 and UBE2L3 overexpression suppresses HPV+ HNC tumor growth in vivo . mEERL cells overexpressing Cul1 or Ube2l3 were generated by lentiviral transduction of the Cul1 or Ube2l3 genes , respectively, and blasticidin selection. CUL1, UBE2L3, and HPV16 E7 proteins were detected by western blotting ( A ). Relative band intensities were quantified using NIH ImageJ ( B ). β-actin was used as an internal control. The data shown are means ± SD of three independent experiments. Student’s t -test determined P -values. * P < 0.05, ** P < 0.01, *** P < 0.001. mEERL/vector ( C ), mEERL/ Cul1 ( D ), or mEERL/ Ube2l3 ( E ) cells were injected into the rear right flank of C57BL/6J mice ( n = 10 per group). Tumor volume was measured twice a week ( C–F ). Survival rates of mice were analyzed using a Kaplan-Meier estimator ( G ). The time to event was determined for each group, with the event defined as a tumor size larger than 2,000 mm 3 . The data shown are means ± SD. P -values of mice injected with mEERL/ Cul1 and mEERL/ Ube2l3 cells compared with mice injected with mEERL/vector cells were determined for tumor growth ( F ) and survival ( G ) by two-way analysis of variance. Shown are representative of two independent experiments.

Article Snippet: UBE2L3 , Human/Mouse , Proteintech , 14115–1-AP , AB_2210891 , IP-WB.

Techniques: Over Expression, In Vivo, Generated, Transduction, Selection, Western Blot, Control, Plasmid Preparation, Injection

Journal: Journal of Virology

Article Title: The membrane-associated ubiquitin ligase MARCHF8 stabilizes the human papillomavirus oncoprotein E7 by degrading CUL1 and UBE2L3 in head and neck cancer

doi: 10.1128/jvi.01726-23

Figure Lengend Snippet: The schematic model of MARCHF8-mediated HPV16 E7 stabilization by degrading CUL1 and UBE2L3. The HPV16 oncoprotein E6 activates the MARCHF8 promoter activity through the MYC/MAX transcription factor complex and upregulates MARCHF8 in the HPV+ HNC cells . MARCHF8 protein ubiquitinates and degrades CUL1 and UBE2L3 proteins, leading to the prevention of HPV16 E7 protein degradation.

Article Snippet: UBE2L3 , Human/Mouse , Proteintech , 14115–1-AP , AB_2210891 , IP-WB.

Techniques: Activity Assay

Journal: Journal of Virology

Article Title: The membrane-associated ubiquitin ligase MARCHF8 stabilizes the human papillomavirus oncoprotein E7 by degrading CUL1 and UBE2L3 in head and neck cancer

doi: 10.1128/jvi.01726-23

Figure Lengend Snippet: List of the oligonucleotides ^a

Article Snippet: UBE2L3 , Human/Mouse , Proteintech , 14115–1-AP , AB_2210891 , IP-WB.

Techniques: Sequencing, Cloning

Journal: Journal of Virology

Article Title: The membrane-associated ubiquitin ligase MARCHF8 stabilizes the human papillomavirus oncoprotein E7 by degrading CUL1 and UBE2L3 in head and neck cancer

doi: 10.1128/jvi.01726-23

Figure Lengend Snippet: List of the antibodies

Article Snippet: UBE2L3 , Human/Mouse , Proteintech , 14115–1-AP , AB_2210891 , IP-WB.

Techniques: Western Blot, Ubiquitin Proteomics