Journal: Cell Death and Differentiation

Article Title: Anti-apoptotic A1 is not essential for lymphoma development in Eµ-Myc mice but helps sustain transplanted Eµ-Myc tumour cells

doi: 10.1038/s41418-017-0045-8

Figure Lengend Snippet: Expression of BCL-2 protein family members in A1 +/+ Eµ -Myc and A1 −/− Eµ -Myc lymphomas and premalignant cells. Western blots were performed on a pro/pre-B cell lymphomas from lymph nodes of sick A1 +/+ Eµ -Myc (lanes 1–4) and A1 − / − Eµ -Myc mice (lanes 5–8) and pre-malignant pro/pre-B cells (MACS-sorted CD19 + bone marrow (BM) samples) from healthy A1 +/+ Eµ -Myc and A1 − / − Eµ -Myc mice euthanased at 28–30 days of age (lanes 9–12); b B cell lymphomas from lymph nodes of A1 +/+ Eµ -Myc (lanes 1–4) and A1 −/− Eµ -Myc mice (lanes 5–8). The same premalignant samples and control lysates as in ( a ) were run to enable comparison between ( a ) and ( b ). Lane labels indicate the individual mouse number. WEHI-231 cells served as a positive control for A1 protein, tumour 1/8094 as a positive control for p53 and p19ARF proteins and β-ACTIN as a loading control. Independent blots are separated by a horizontal black line and molecular weight markers (kD) are indicated. Additional lymphomas were analysed (data not shown), in total: 8 pro/pre-B and 8 B A1 +/+ Eµ -Myc ; 7 pro/pre-B and 6 B A1 −/− Eµ -Myc

Article Snippet: Membranes were subsequently probed with the following antibodies: A1 (clone 6D6, WEHI mAB lab), BCL-2 (clone 7, BD Biosciences), BCL-X L (clone 44, BD Biosciences), MCL-1 (clone 19C4–15, WEHI mAb lab), PUMA (polyclonal, Abcam), BIM (clone 3C5, WEHI mAb lab), p53 (FL-393, Santa Cruz Biotechnology, Santa Cruz, CA, USA), p19ARF (p19ARF exon 2, Rockland, Gilbertsville, PA, USA), c-MYC (D84C12, Cell Signaling Techology, Danvers, MA, USA) and β-ACTIN (clone AC-74, Sigma-Aldrich).

Techniques: Expressing, Western Blot, Control, Comparison, Positive Control, Molecular Weight

Journal: Journal of Clinical & Cellular Immunology

Article Title: Tumor Suppressor Gene P29ing4 is Overexpressed and Induces a CD8 T Effector Cell Response in Human Renal Cell Carcinoma

doi: 10.4172/2155-9899.1000511

Figure Lengend Snippet: Figure 1: Expression of p33ING1b and p29ING4 in renal cell carcinoma. (a) mRNA expression of p33ING1b and p29ING4 was increased at early and late stages of disease with higher expression of both genes at late stages of disease compared to early stages (Robson stages III/IV vs. I/II) (p33ING1b and p29ING4: stages I/II and III/IV vs. normal tissue p<0.001 and p<0.0001, respectively; p33ING1b and p29ING4: stages III/IV vs. I/II p<0.01 and p<0.001, respectively). (b) Results of the real time RT-PCR were normalized to kidney normal tissue and expressed as x-fold difference (2- ∆∆Ct). Expression of p33ING1b and p29ING4 by renal carcinoma cells was confirmed (Cy3 red; DAPI blue (nuclear counterstaining)). (c) Sections are representative of n=30 patients/group. Representative western blots of p33ING1b and p29ING4 (loading control ß-actin).

Article Snippet: Analysis of single staining was performed for p33ING1 (Santa Cruz Biotechnology, CA, USA), p29ING4 (Rockland/Biomol, Hamburg, Germany) CD4, CD25, Foxp3, CD8, (Abcam, Cambridge, UK), IFN-γ (BD Pharmingen, Heidelberg, Germany), IL-2, and IL-10 (R&D Systems, Minneapolis, USA).

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

Journal: Journal of Clinical & Cellular Immunology

Article Title: Tumor Suppressor Gene P29ing4 is Overexpressed and Induces a CD8 T Effector Cell Response in Human Renal Cell Carcinoma

doi: 10.4172/2155-9899.1000511

Figure Lengend Snippet: Figure 4: p33ING1b- and p29ING4-specific T cell reactivity. Four peptides resulted in significantly increased expression of IFN-γ compared to controls, independent of tumor stage (p33ING1b (aa109-118) and aa259-268, and p29ING4 (aa149-158) and aa239-248; p<0.001) Stages I/ II (a) and Stages III/IV (b). Elispot analysis was measured as spots/5 x 105 cells/patient.

Article Snippet: Analysis of single staining was performed for p33ING1 (Santa Cruz Biotechnology, CA, USA), p29ING4 (Rockland/Biomol, Hamburg, Germany) CD4, CD25, Foxp3, CD8, (Abcam, Cambridge, UK), IFN-γ (BD Pharmingen, Heidelberg, Germany), IL-2, and IL-10 (R&D Systems, Minneapolis, USA).

Techniques: Expressing, Enzyme-linked Immunospot

Journal: Journal of Clinical & Cellular Immunology

Article Title: Tumor Suppressor Gene P29ing4 is Overexpressed and Induces a CD8 T Effector Cell Response in Human Renal Cell Carcinoma

doi: 10.4172/2155-9899.1000511

Figure Lengend Snippet: Figure 5: p33ING1b- and p29ING4-specific T cell reactivity. Two peptides resulted in increased expression of IL-10 compared to controls; peptide p33ING1b (aa109-118) at all stages and p29ING4 (aa239-248) at late stages (p<0.001). Stages I/ II (a) and Stages III/IV (b). Luminex analysis was measured as pg/ml.

Article Snippet: Analysis of single staining was performed for p33ING1 (Santa Cruz Biotechnology, CA, USA), p29ING4 (Rockland/Biomol, Hamburg, Germany) CD4, CD25, Foxp3, CD8, (Abcam, Cambridge, UK), IFN-γ (BD Pharmingen, Heidelberg, Germany), IL-2, and IL-10 (R&D Systems, Minneapolis, USA).

Techniques: Expressing, Luminex

Journal: Journal of Clinical & Cellular Immunology

Article Title: Tumor Suppressor Gene P29ing4 is Overexpressed and Induces a CD8 T Effector Cell Response in Human Renal Cell Carcinoma

doi: 10.4172/2155-9899.1000511

Figure Lengend Snippet: Figure 6: p33ING1b- and p29ING4-specific T cell reactivity. Evaluation of IL-2 expression revealed that only a single epitope (p29ING4 (aa149-158)) resulted in high responsiveness of T cells from RCC patients after stimulation with the peptide (stages I/II vs. III/IV p<0.001). PBMCs from healthy volunteers (n=30) had no significant IL-2 expression after stimulation with all peptide epitopes. Luminex analysis for IL-2 expression was measured as pg/ml and presented as percent stimulation index.

Article Snippet: Analysis of single staining was performed for p33ING1 (Santa Cruz Biotechnology, CA, USA), p29ING4 (Rockland/Biomol, Hamburg, Germany) CD4, CD25, Foxp3, CD8, (Abcam, Cambridge, UK), IFN-γ (BD Pharmingen, Heidelberg, Germany), IL-2, and IL-10 (R&D Systems, Minneapolis, USA).

Techniques: Expressing, Luminex

Journal: Journal of Clinical & Cellular Immunology

Article Title: Tumor Suppressor Gene P29ing4 is Overexpressed and Induces a CD8 T Effector Cell Response in Human Renal Cell Carcinoma

doi: 10.4172/2155-9899.1000511

Figure Lengend Snippet: Figure 7: p29ING4 presentation by MHC class I (a) and II (b) molecules. p29ING4 presentation by major histocompatibility complex (MHC) class I and II antigens was shown to be increased in RCC but not in healthy kidneys.

Article Snippet: Analysis of single staining was performed for p33ING1 (Santa Cruz Biotechnology, CA, USA), p29ING4 (Rockland/Biomol, Hamburg, Germany) CD4, CD25, Foxp3, CD8, (Abcam, Cambridge, UK), IFN-γ (BD Pharmingen, Heidelberg, Germany), IL-2, and IL-10 (R&D Systems, Minneapolis, USA).

Techniques: Immunopeptidomics

Journal: NPJ Precision Oncology

Article Title: Y chromosome-linked EIF1AY deletion drives sex differences in multiple myeloma

doi: 10.1038/s41698-026-01317-0

Figure Lengend Snippet: A Global incidence rates of multiple myeloma (MM) in males and females retrieved from the Global Burden of Disease (GBD) 2021 database. B Global mortality rates of MM in males and females from the same database. C Kaplan–Meier survival curves were generated using data from the Cancer Genome Atlas (TCGA) to compare overall survival (OS) between male and female MM patients. D Volcano plot showing differential gene expression between male and female MM patients from the GSE6401 dataset. Red dots indicate genes upregulated in males, blue dots indicate downregulated genes, and gray dots represent nonsignificant changes. E Heatmap visualizing the same dataset, with red and blue indicating male and female samples, respectively. F A heatmap showing the correlation between gene co-expression modules and clinical traits in male and female MM samples. Each cell displays the correlation coefficient and corresponding p -value between a module eigengene and a clinical trait. G A protein–protein interaction (PPI) network was constructed using STRING (v12.0) based on the intersection of differentially expressed genes and genes from the MEgrey module. Nodes represent proteins, and edges indicate predicted interactions. Hub genes were identified via maximum clique centrality scoring, suggesting their central regulatory roles. H Kaplan–Meier analysis of OS in male MM patients stratified by EIF1AY expression using the Kaplan–Meier plotter database. I Kaplan–Meier analysis of OS in male MM patients stratified by RPS4Y1 expression. J Kaplan–Meier analysis of OS in male MM patients stratified by KDM5D expression. Data were analyzed using the log-rank test. K mRNA expression levels of EIF1AY in peripheral blood samples from 30 male healthy donors (HDs) and 54 male patients. L mRNA expression levels of RPS4Y1 in the same cohort. M Protein expression levels of RPS4Y1 and EIF1AY in HDs and MM patients. N Dual-color fluorescence in situ hybridization (FISH) targeting EIF1AY (Yq11.223, orange) and centromere X (CenX, green) was performed on bone marrow cells from HDs and MM patients. O Representative images show the presence of EIF1AY-positive cells in HDs and loss of EIF1AY in MM samples.

Article Snippet: Primary antibodies included GAPDH (10494-1-AP), beta tubulin (10094-1-AP), EIF1AY (11193-1-AP), RPS4Y1 (17296-1-AP), CD134 (20006-1-AP), and secondary antibody (RGAR004) (all from Proteintech).

Techniques: Generated, Gene Expression, Expressing, Construct, Fluorescence, In Situ Hybridization

Journal: NPJ Precision Oncology

Article Title: Y chromosome-linked EIF1AY deletion drives sex differences in multiple myeloma

doi: 10.1038/s41698-026-01317-0

Figure Lengend Snippet: A Cell proliferation assay showing the growth rate of RPMI 8226 cells overexpressing EIF1AY. B Tumor weight of male MM xenografts following EIF1AY overexpression. C Representative images of xenograft tumors from the same experiment. D Tumor growth curves of xenografts over 30 days. E Hematoxylin and eosin (HE) staining and immunohistochemical (IHC) detection of EIF1AY and Ki-67 expression in xenograft tissues. Scale bar: 80 μm. F Comparative analysis of the fractional distribution of 22 immune cell types in male versus female MM samples from the GSE164701 dataset. G Phorbol 12-myristate 13-acetate (PMA)-stimulated THP-1 cells were differentiated into M0 macrophages (THP-1-Mφ). qRT-PCR analysis of M2 macrophage markers (IL-10, TGF-β, ARG1, and CD206) in THP-1-Mφ exposed to conditioned medium (CM) from EIF1AY-overexpressing MM cells. H CD206 protein expression in THP-1-Mφ treated with CM from EIF1AY-overexpressing MM cells was evaluated by Western blot analysis. I Flow cytometric analysis of CD206 expression in THP-1-Mφ across the indicated groups. J IHC analysis of CD206 expression in xenograft tumor tissues. Scale bar: 80 μm. K Chemotaxis assay assessing migration of THP-1-Mφ in response to CM from EIF1AY-overexpressing U266 cells. L Chemotaxis assay assessing migration of THP-1-Mφ in response to CM from EIF1AY-overexpressing RPMI-8226 cells under the same experimental conditions. M Proliferation of MM cells cultured with CM from M2-polarized THP-1-Mφ induced by EIF1AY-overexpressing U266 cells. N Proliferation of MM cells cultured with CM from M2-polarized THP-1-Mφ induced by EIF1AY-overexpressing RPMI-8226 cells under the same experimental conditions.

Article Snippet: Primary antibodies included GAPDH (10494-1-AP), beta tubulin (10094-1-AP), EIF1AY (11193-1-AP), RPS4Y1 (17296-1-AP), CD134 (20006-1-AP), and secondary antibody (RGAR004) (all from Proteintech).

Techniques: Proliferation Assay, Over Expression, Staining, Immunohistochemical staining, Expressing, Quantitative RT-PCR, Western Blot, Chemotaxis Assay, Migration, Cell Culture

Journal: NPJ Precision Oncology

Article Title: Y chromosome-linked EIF1AY deletion drives sex differences in multiple myeloma

doi: 10.1038/s41698-026-01317-0

Figure Lengend Snippet: A Volcano plot of differentially expressed genes between normal macrophages (MΦs) and tumor-associated macrophages (TAMs) from GSE143583 . B Heatmap of the 381 differentially expressed genes between MΦs and TAMs; color intensity indicates expression. C WGCNA module-trait correlation heatmap; rows = module eigengenes, columns = macrophage subtypes. D Venn diagram showing overlap among differentially expressed genes, MEturquoise module genes, and membrane proteins from the Membranome database. E PPI network of genes in ( D ) generated via STRING; edges indicate predicted interactions. F DDR1 expression levels in MΦs and TAMs from GSE143583 . G DDR1 mRNA in THP-1-Mφ cultured with or without CM from MM cells, measured by qRT-PCR. H qRT-PCR analysis of IL-10, TGF-β, ARG1, and CD206 mRNA levels in THP-1-Mφ cultured with CM from MM cells overexpressing DDR1. I Flow cytometry analysis of CD206 expression in the indicated experimental groups. J qRT-PCR analysis of DDR1 mRNA expression in THP-1-Mφ cultured with or without CM from EIF1AY-overexpressing RPMI-8226 cells. K qRT-PCR analysis of DDR1 mRNA expression in THP-1-Mφ cultured with or without CM from EIF1AY-overexpressing U266 cells under the same experimental conditions. L Immunofluorescence (IF) analysis of DDR1 expression in THP-1-Mφ cultured with or without CM from EIF1AY-overexpressing MM cells. M Migration assay of THP-1-Mφ transduced to overexpress DDR1, cultured in CM from MM cells with or without EIF1AY overexpression (CM Vector or CM oe-EIF1AY). Recruited cells in the lower chamber were quantified to assess whether DDR1 overexpression in THP-1-Mφ rescues the migration impairment induced by EIF1AY expression in MM cells. N THP-1-Mφ pretreated with CM from EIF1AY-overexpressing or control MM cells were subsequently transduced with DDR1 and indirectly cocultured with MM cells in a transwell system. MM cell proliferation was then assessed to evaluate the effect of macrophage-derived signals. O qRT-PCR analysis of IL-4 and IL-13 mRNA levels in RPMI 8226 cells following EIF1AY overexpression. P ELISA analysis of IL-4 and IL-13 secretion in culture supernatants from EIF1AY-overexpressing MM cells. Q qRT-PCR analysis of DDR1 mRNA expression in THP-1-Mφ stimulated with or without IL-4 and IL-13.

Article Snippet: Primary antibodies included GAPDH (10494-1-AP), beta tubulin (10094-1-AP), EIF1AY (11193-1-AP), RPS4Y1 (17296-1-AP), CD134 (20006-1-AP), and secondary antibody (RGAR004) (all from Proteintech).

Techniques: Expressing, Membrane, Generated, Cell Culture, Quantitative RT-PCR, Flow Cytometry, Immunofluorescence, Migration, Over Expression, Plasmid Preparation, Control, Transduction, Derivative Assay, Enzyme-linked Immunosorbent Assay

Journal: NPJ Precision Oncology

Article Title: Y chromosome-linked EIF1AY deletion drives sex differences in multiple myeloma

doi: 10.1038/s41698-026-01317-0

Figure Lengend Snippet: A Scatterplot showing positive correlation between RPS4Y1 and EIF1AY in the GSE6401 dataset. B Correlation validated in male MM patient samples. C qRT-PCR of EIF1AY mRNA in RPMI-8226 cells after RPS4Y1 knockdown. D qRT-PCR of EIF1AY mRNA in RPMI-8226 cells after RPS4Y1 overexpression. E Western blot of EIF1AY protein in RPMI-8226 cells after RPS4Y1 knockdown. F Western blot of EIF1AY protein in RPMI-8226 cells after RPS4Y1 overexpression. G qRT-PCR of IL-10, TGF-β, ARG1, and CD206 in THP-1-Mφ cultured with CM from RPS4Y1-overexpressing MM cells. H Western blot of CD206 in THP-1-Mφ treated as in ( G ). I qRT-PCR of DDR1 mRNA in THP-1-Mφ cultured with CM from RPS4Y1-overexpressing RPMI-8226 cells. J IF staining of DDR1 in RPS4Y1-overexpressing RPMI-8226 cells. K qRT-PCR of IL-4 mRNA in RPMI-8226 cells overexpressing RPS4Y1. L qRT-PCR of IL-13 mRNA in RPMI-8226 cells treated as in ( K ). M ELISA quantification of IL-4 in culture supernatants after RPS4Y1 overexpression. N ELISA quantification of IL-13 in culture supernatants after RPS4Y1 overexpression. O Structure-based modeling of the RPS4Y1–EIF1AY complex showing key interaction residues and predicted binding affinity. P Co-IP in RPMI-8226 cells with anti-EIF1AY to detect RPS4Y1. Q Co-IP in RPMI-8226 cells with anti-RPS4Y1 to detect EIF1AY. R Co-IP in U266 cells with anti-EIF1AY to detect RPS4Y1. S Co-IP in U266 cells with anti-RPS4Y1 to detect EIF1AY. T Co-IP in Flag-RPS4Y1 cells to detect EIF1AY. U Co-IP in Flag-EIF1AY cells to detect RPS4Y1. V THP-1-Mφ chemotactic migration assays were performed using CM derived from RPS4Y1-overexpressing MM cells, with or without EIF1AY knockdown. W The number of tumor cells was quantified following incubation with CM from polarized THP-1-Mφ induced by either RPS4Y1-overexpressing MM cells or RPS4Y1-overexpressing MM cells with EIF1AY knockdown.

Article Snippet: Primary antibodies included GAPDH (10494-1-AP), beta tubulin (10094-1-AP), EIF1AY (11193-1-AP), RPS4Y1 (17296-1-AP), CD134 (20006-1-AP), and secondary antibody (RGAR004) (all from Proteintech).

Techniques: Quantitative RT-PCR, Knockdown, Over Expression, Western Blot, Cell Culture, Staining, Enzyme-linked Immunosorbent Assay, Binding Assay, Co-Immunoprecipitation Assay, Migration, Derivative Assay, Incubation

Journal: NPJ Precision Oncology

Article Title: Y chromosome-linked EIF1AY deletion drives sex differences in multiple myeloma

doi: 10.1038/s41698-026-01317-0

Figure Lengend Snippet: A Correlation between CD134 and RPS4Y1 expression in male MM samples. B Correlation between CD134 and EIF1AY expression in male MM samples. C qRT-PCR analysis of CD134 mRNA in RPMI-8226 cells after RPS4Y1 knockdown. D qRT-PCR analysis of CD134 mRNA in RPMI-8226 cells after RPS4Y1 overexpression. E qRT-PCR analysis of CD134 mRNA in RPMI-8226 cells after EIF1AY knockdown. F qRT-PCR analysis of CD134 mRNA in RPMI-8226 cells after EIF1AY overexpression. G – J Corresponding protein expression levels of CD134 in RPMI 8226 cells upon RPS4Y1 or EIF1AY knockdown and overexpression. G Western blot analysis of CD134 protein in RPMI-8226 cells after RPS4Y1 knockdown. H Western blot analysis of CD134 protein in RPMI-8226 cells after RPS4Y1 overexpression. I Western blot analysis of CD134 protein in RPMI-8226 cells after EIF1AY knockdown. J Western blot analysis of CD134 protein in RPMI-8226 cells after EIF1AY overexpression. K CD134 protein expression in RPMI 8226 cells was analyzed by Western blot following overexpression of RPS4Y1 alone or combined overexpression of RPS4Y1 and shEIF1AY. L Chemotactic migration of THP-1-Mφ in response to CM derived from MM cells treated with siCD134, oe-RPS4Y1 + siCD134, or oe-EIF1AY + siCD134. M Quantification of MM cell numbers cultured with CM derived from THP-1-Mφ polarized using the same treatment conditions described in ( L ). N RNA pull-down assay in RPMI-8226 cells using CD134-specific probes to detect binding to RPS4Y1 and EIF1AY. O RNA pull-down assay in U266 cells using CD134-specific probes to detect binding to RPS4Y1 and EIF1AY. P RNA immunoprecipitation (RIP) assay in Flag-RPS4Y1–overexpressing RPMI-8226 and U266 cells using anti-Flag antibodies. CD134 mRNA enrichment was quantified by qRT-PCR relative to IgG control (2 −ΔΔCt ). Q RIP assay in Flag-EIF1AY-overexpressing RPMI-8226 and U266 cells using anti-Flag antibodies. R CD134 mRNA stability in RPMI-8226 cells following RPS4Y1 knockdown, assessed by actinomycin D chase and qRT-PCR. S CD134 mRNA stability in RPMI-8226 cells following EIF1AY knockdown, assessed by actinomycin D chase and qRT-PCR.

Article Snippet: Primary antibodies included GAPDH (10494-1-AP), beta tubulin (10094-1-AP), EIF1AY (11193-1-AP), RPS4Y1 (17296-1-AP), CD134 (20006-1-AP), and secondary antibody (RGAR004) (all from Proteintech).

Techniques: Expressing, Quantitative RT-PCR, Knockdown, Over Expression, Western Blot, Migration, Derivative Assay, Cell Culture, Pull Down Assay, Binding Assay, RNA Immunoprecipitation, Control

Journal: NPJ Precision Oncology

Article Title: Y chromosome-linked EIF1AY deletion drives sex differences in multiple myeloma

doi: 10.1038/s41698-026-01317-0

Figure Lengend Snippet: EIF1AY, a Y chromosome-encoded protein, forms a complex with RPS4Y1 to directly bind and stabilize CD134 mRNA, thereby sustaining CD134 expression in MM cells. CD134 signaling suppresses the secretion of IL-4 and IL-13, cytokines that normally induce DDR1 expression on macrophages and promote their polarization toward the tumor-supportive M2 phenotype. The RPS4Y1-EIF1AY-CD134 axis inhibits M2 macrophage polarization and recruitment, limiting MM cell proliferation. Loss of EIF1AY disrupts this axis, resulting in increased IL-4 and IL-13 secretion, upregulated DDR1 expression on macrophages, enhanced M2 polarization, and accelerated tumor progression. This feed-forward regulatory loop reveals a Y chromosome-linked immune mechanism underlying sex differences in MM and identifies EIF1AY as a potential target for precision immunotherapy in male patients.

Article Snippet: Primary antibodies included GAPDH (10494-1-AP), beta tubulin (10094-1-AP), EIF1AY (11193-1-AP), RPS4Y1 (17296-1-AP), CD134 (20006-1-AP), and secondary antibody (RGAR004) (all from Proteintech).

Techniques: Expressing