anti cpsf2  (Santa Cruz Biotechnology)

Journal: Nature Communications

Article Title: LENG8 granule regulates alternative polyadenylation in mammals

doi: 10.1038/s41467-026-77634-6

Figure Lengend Snippet: a , b Heatmap showing total ( a ) and LENG8 target genes ( b ) with shortened or lengthened 3′UTR in three types of germ cells. Line plot on the top showing the mean PDUI in spermatogonia (SG), spermatocytes (SC), and round spermatids (RS). c – f Immunofluorescence images of the relationship between CPSF4 (green) and LENG8 (magenta) ( c ), LENG8 (green) and FIP1L1 (magenta) ( d ), LENG8 (magenta) and YTHDC1 (green) ( e ), LENG8 (magenta) and PABPN1(green) ( f ) in spermatocytes (top) and round spermatids (bottom). Nuclei were counterstained with DAPI (blue). Middle: magnified views of the areas enclosed by dashed boxes. Right: Pearson correlation coefficient showing co-localization patterns between LENG8 and CPSF4, FIP1L1, YTHDC1, and PABPN1. Data are presented as mean ± SEM, n = 20 LENG8 granules from three biologically independent experiments. P -values were determined by two-sided unpaired Student’s t test. g Heatmap illustrating the RNA level, ribosome-protected fragments (RPFs, which correlate with translational efficiency) and protein abundance of targets in four clusters across spermatogonia, spermatocytes and round spermatids. h GO-term enrichment analysis of LENG8 targets in testes, Statistical significance was determined by a modified Fisher′s exact test (EASE score). i Model for LENG8 granule-mediated APA regulation. LENG8 granules mediate the recruitment of CPA factors and facilitate CPA complex assembly at pPASs in a YTHDC1-m ⁶ A dependent manner, ultimately enhancing the usage of these sites. Source data are provided as a Source Data file.

Article Snippet: Antibodies were used for western blot, IF, and IP: anti-LENG8 (Signalway Antibody, 39875, 1:100 in IF), anti-LENG8 (Bethyl, A304-947A, 1:1000 in WB), anti-LENG8 (our lab, 1:100 in IF and IP), anti-PABPN1 (Beyotime Biotechnology, AG3616, 1:2000 in WB, 1:200 in IF), anti-YTHDC1(Santa Cruz, sc-136428, 1:200 in IF), anti-CPSF4 (Santa Cruz, sc-390643, 1:10 in IF), anti-SYCP3(Santa Cruz, sc-74569, 1:300 in IF), anti-NONO (Santa Cruz, sc-166702, 1:10 in IF), anti-DDX25 (Santa Cruz, sc-166289, 1:200 in IF), anti-CPSF4 (Proteintech, 15023-1-AP, 1:1000 in WB), anti-COIL (Proteintech, 67218-1-Ig, 1:100 in IF), anti-YTHDC1 (ABclonal, A22992, 1:1000 in WB), anti-FIP1L1(ABclonal, A7138, 1:1000 in WB, 1:100 in IF), anti-GFP (Abways, AB0005, 1:2000 in WB), anti-β-actin (Abways, Ab2001, 1:10000 in WB), anti-Myc (Abways, AB0001, 1:2000 in WB), anti-Flag (Abways, AB0008, 1:2000 in WB), anti-HA (Cell Signaling Technology, 3724, 1:2000 in WB), anti-SC35 (Abcam, Ab11826, 1:200 in IF), anti-CPSF2 (ABclonal, A9297, 1:1000 in WB, 1:100 in IF), anti-CPSF6(Santa Cruz, sc-376228, 1:100 in IF), anti-SC35 (Santa Cruz, sc-53518, 1:100 in IF), Mouse polyclonal anti-IgG (Merck, 12-371), Rabbit polyclonal anti-IgG (Merck, 12-370), HRP, Goat Anti-Mouse IgG (Abbkine, A21010, 1:10000 in WB), HRP, Goat Anti-Mouse IgG LCS (Abbkine, A25012, 1:10000 in WB), HRP, Goat Anti-Rabbit IgG (Abbkine, A21020, 1:10000 in WB), Alexa Fluor 488 AffiniPure Donkey Anti-MOUSE (JACKSON ImmunoResearch, 715-545-150, 1:500 in IF), Alexa Fluor 594 AffiniPure Donkey Anti-Rabbit (JACKSON ImmunoResearch, 711-585-152, 1:500 in IF), IFKine Red Donkey Anti-Mouse (Abbkine, A24411, 1:500 in IF) and IFKine Green Donkey Anti-Rabbit (Abbkine, A24221, 1:500 in IF), ABflo 647-conjugated Goat anti-Mouse IgG (ABclonal, AS059, 1:500 in IF).

Techniques: Immunofluorescence, Quantitative Proteomics, Analysis, Modification, Cell Differentiation, Significance Assay

Journal: Nucleic Acids Research

Article Title: eIF2D promotes 40S ribosomal subunit recycling during intrinsic ribosome destabilization

doi: 10.1093/nar/gkaf1322

Figure Lengend Snippet: Decreased IRD-inducing protein levels and altered splicing patterns in eIF2D-deficient cells. ( A ) Cumulative fraction of the fold changes in protein levels in eIF2D KO cells relative to the control cells ( n = 4 biologically independent samples). Protein levels were analyzed via DIA-MS, and the results are shown for all detected, short nascent chain, and long nascent chain proteins. P -values were calculated using the two-tailed Mann–Whitney U test. ( B ) Differences in the median log 2 fold changes in protein levels between the indicated cell lines and control cells. ** P < .01 and *** P < .001; NS, not significant (two-tailed Mann–Whitney U test). ( C ) Gene Ontology (GO) term analysis of IRD target genes with reduced protein levels in eIF2D KO cells. FDR, false discovery rate. ( D ) Volcano plots showing the differences in the PSI scores of skipped exons between the eIF2D KO and control cells ( n = 2 biologically independent samples). Gray dashed lines indicate a ∆PSI of ±0.05 and an FDR of 0.05. ( E ) Numbers of skipped and retained introns in the eIF2D KO, eIF2A KO, and MCTS1 KO cells compared to those isplicing, and transcripn the control cells. Alternative splicing events not consistently altered in two independent clones were excluded. ( F ) Cleavage and polyadenylation specific factor 2 ( CPSF2 ) gene structure and Sashimi plots showing the differential inclusion of CPSF2 exon 14. Representative data of two replicates are shown. ( G ) Immunoblotting analysis of WT, eIF2D KO, and MCTS1 KO cells using the indicated antibodies. HSP90 was used a loading control.

Article Snippet: Primary antibodies against MCTS1 (#GTX117793; GeneTex), DENR (203057-T46; Sino biologicals), eIF2D (12840-1-AP; Proteintech), CPSF2 (A301-581A; Bethyl Laboratories), HSP90 (610 419; BD Biosciences), and V5 epitope (R960CUS; Thermo Fisher Scientific) were used in this study.

Techniques: Control, Two Tailed Test, MANN-WHITNEY, Alternative Splicing, Clone Assay, Western Blot

Journal: Cell reports

Article Title: Non-canonical isoforms of the mRNA polyadenylation factor WDR33 regulate STING-mediated immune responses

doi: 10.1016/j.celrep.2024.113886

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Mouse anti-CPSF2 , Santa Cruz Biotechnology , Cat# sc-165983; RRID: AB_2084371.

Techniques: Virus, Recombinant, Negative Control, Software