Journal: MedComm

Article Title: LINC02774 inhibits glycolysis in glioma to destabilize HIF‐1α dependent on transcription factor RP58

doi: 10.1002/mco2.364

Figure Lengend Snippet: RP58 in RIEMR‐associated LINC02774 regulates the PHD3 and suppresses the hypoxia signaling activity. (A) RIP assay was used to detect the level of LINC02774 binding to RP58 under endogenous or exogenous LINC02774 derived from U251 transfected with LINC02774 or vector. (B) RNA pull‐down analysis of FLAG‐tagged RP58 (Full‐length, 1−276aa vs 277−531aa) pull‐down. (C) Western blot to detect the expression level of HIF‐1α in U251 cells after overexpressing RP58. (D) The level of PHD3, HIF‐1α‐OH, GLUT1 and VEGF in U251 cells overexpressing RP58. (E, F) The transcription level of PHD3 and HIF‐1α following the overexpression of RP58 were identified in the U251 cell line as well as culture in 21% (normoxia)(E) or 1%O2 (hypoxia)(F) for 24 h. (G) ChIP analysis of RP58 binding to PHD3 promoters in U251cell transfected with LINC02774 or vector. (H) The expression of PHD3 in LINC02774‐overexpressing U251 cells transiently transfected with RP58‐knockdown shRNA plasmid, and cell lysates were harvested at 72 h. (I, J) The xenograft tumors were shown in nude mice. Tumor images (I) and weights (J) were recorded ( n = 7 for each group). (K) The expression levels of RP58, PHD3, HIF‐1α, VEGF, and GLUT1 identified by IHC in xenograft tumors.

Article Snippet: Then, antibodies used were as follows: HIF‐1α (Novus, # NB100‐105, 1:40), PHD3 (Novus, #NB100‐139, 1:500), GLUT1 (Proteintech, # 66290‐1‐Ig, 1:400), RP58 (Proteintech, #12714‐1‐AP, 1:4000), and VEGF (Proteintech, # 19003‐1‐AP, 1:400), 4°C overnight, then incubated with the secondary antibody.

Techniques: Activity Assay, Binding Assay, Derivative Assay, Transfection, Plasmid Preparation, Western Blot, Expressing, Over Expression, Knockdown, shRNA

Journal: MedComm

Article Title: LINC02774 inhibits glycolysis in glioma to destabilize HIF‐1α dependent on transcription factor RP58

doi: 10.1002/mco2.364

Figure Lengend Snippet: RIEMR‐associated LINC02774 is a biomarker for prognosis and progression in glioma patients. (A) Immunohistochemistry staining detection of PHD3, HIF‐1α, GLUT1 and VEGF proteins in glioma along with different WHO grades (I, II, III and IV). (B) Kaplan–Meier analysis was conducted to evaluate the OS curves of glioma patients based on the expression levels of LINC02774. The TCGA database was utilized, and the specimens were divided into two groups: low LINC02774 expression ( n = 342) and high LINC02774 expression ( n = 343). (C) Kaplan–Meier analysis was used to assess the OS in glioma patients based on the expression levels of both LINC02774 and RP58. (D, E) Clinical follow‐up was conducted on 56 glioma patients, the OS(D) and RFS (E) associated with LINC02774. (F) A schematic model illustrating the major molecular mechanisms of the “LINC02774/RP58/PHD3/HIF‐1α” axis in glioma is shown. OS: overall survival, RFS: recurrence‐free survival.

Article Snippet: Then, antibodies used were as follows: HIF‐1α (Novus, # NB100‐105, 1:40), PHD3 (Novus, #NB100‐139, 1:500), GLUT1 (Proteintech, # 66290‐1‐Ig, 1:400), RP58 (Proteintech, #12714‐1‐AP, 1:4000), and VEGF (Proteintech, # 19003‐1‐AP, 1:400), 4°C overnight, then incubated with the secondary antibody.

Techniques: Biomarker Discovery, Immunohistochemistry, Staining, Expressing

Journal: bioRxiv

Article Title: The structural basis of Indisulam-mediated recruitment of RBM39 to the DCAF15-DDB1-DDA1 E3 ligase complex

doi: 10.1101/737510

Figure Lengend Snippet: a , Bioinformatics workflow for identifying proteins bearing putative degron motif required for DCAF15-Indisulam recruitment (left) and structures of DCAF15-bound Indisulam (orange) and RBM39(RRM2) (magenta) with central alpha helix highlighted (right). The RBM39 residues found to be most critical for DCAF15-Indisulam recruitment are labeled (M265, G268, E271, and P272). 6,475 proteins with known structures were identified in the Swiss-Prot database, of which 3,425 had a glycine in an alpha helix. 3,112 of the glycine-containing alpha helices aligned to an RBM39(RRM2) structure (2JRS) with RMSD <2.0 Å. Among these matches, only RBM23 and RBM39 helices had a sequence matching the required X 1 XXM 4 XXG 7 XXEP motif. RMSD values, PDB IDs, and gene names shown in bottom table. b , Volcano plot summary of expression proteomics experiments comparing lysates from HCT-116 cells treated for 4 h with 10 µM Indisulam or DMSO. Significant downregulated proteins (p value < 1E-2, Log 2 fold-change) are labeled. Data represents two (n=2) biological replicates per treatment condition in a single experiment. c , Western blots showing levels of RBM39, RBM23, and ZNF277 in HCT116 cells following 4 h treatment with varied concentrations of Indisulam or DMSO. Data shown from one individual, representative experiment from three independent repeats.

Article Snippet: Blots were incubated with primary antibody solutions made in TBS-T with 5% milk for RBM39 (Sigma, Cat #HPA001591, 1:2500) GAPDH (CST, Cat #2118L, 1:1000), Vinculin (Cell Signaling Technology, Cat# 13901S, 1:1000), ZNF277 (Pro-Sci, Cat #46-616, 1:1000), RBM23 (Invitrogen, Cat# PA5-52060, 1:1000), or DDA1 (Proteintech, Cat #14995-1-AP, 1:1000) overnight at 4°C.

Techniques: Labeling, Sequencing, Expressing, Western Blot

Journal: Scientific Reports

Article Title: Global analysis of differential gene expression related to long-term sperm storage in oviduct of Chinese Soft-Shelled Turtle Pelodiscus sinensis

doi: 10.1038/srep33296

Figure Lengend Snippet: DEGs involved in the apoptosis pathway during sperm storage.

Article Snippet: ENSPSIG00000004908 , −1.1619 , MECOM , Zinc finger, C2H2-like.

Techniques: