Journal: Nature Communications

Article Title: Targeting leucine-rich repeat kinase 2 overcomes resistance to oncolytic herpes simplex virus-based therapies in glioblastoma

doi: 10.1038/s41467-026-70132-9

Figure Lengend Snippet: a A scatter plot showing differential gene expression in oHSV-GFP-infected U-251 MG cells treated with or without LRRK2-IN-1 (LRRK2i). U-251 MG cells were pretreated with LRRK2i (2.5 µM) or left untreated for 4 h, followed by infection with oHSV-GFP (MOI = 0.25 PFU/cell) for 4 h. RNA-Seq was used to assess differentially expressed genes among the treated cells. Blue dots represent genes significantly downregulated by LRRK2i during infection, and red dots represent genes significantly upregulated by LRRK2i during infection. b A heatmap showing the expression patterns of differentially expressed genes belonging to the “response to virus” category. The normalized expression values (Log 2 (TPM + 1)) in the heatmap depict the mean across three biological replicates. c GO enrichment analysis showing the top eight most enriched pathways downregulated by LRRK2i in oHSV-GFP-infected U-251 MG cells. d U-251 MG and U-87 MG cell lines were treated with LRRK2-IN-1 (2.5 μM) for 4 h, followed by IFNα and anti-IFNR1 (α-IFNR1) for an additional 4 h, and then infected with oHSV-GFP (MOI = 0.25 PFU/cell). Cell viability was determined 48 h post-infection. e Western blot analysis of p-STAT1, STAT1, and β-actin in GBM cell lines infected with oHSV-GFP or mock-infected, either with or without LRRK2-IN-1 (2.5 µM), 48 h post-infection. IFNα served as a positive control. The samples derive from the same experiment but different gels for p-STAT1, another for STAT1 and another for actin were processed in parallel. f p-STAT1 and STAT1 expression levels were quantified by densitometric analysis of bands from three independent Western blot experiments corresponding to Fig. 4e. g RT‒qPCR analysis of the expression of the indicated IFN-stimulated gene transcript expression in GBM cell lines infected with oHSV-GFP or mock-infected, either with or without LRRK2-IN-1 (2.5 µM), 12 h post-infection. h Western blot analysis of the indicated IFN-stimulated proteins (IFI16 and IRF9) in GBM cell lines infected with oHSV-GFP or mock-infected, either with or without LRRK2-IN-1 (2.5 µM), 48 h post-infection. IFI16 and IRF9 expression levels were quantified by densitometric analysis of bands from three independent Western blot experiments. The samples derive from the same experiment but different gels for IFI16, another for IRF9 and another for actin were processed in parallel. The data are presented as the mean ± SD from two independent experiments ( d ) or three independent experiments ( d , f – h ), and P values were calculated using an ordinary one-way ANOVA ( d , f – h ).

Article Snippet: Recombinant human STAT1 protein (ABclonal, RP01251) was incubated with active, baculovirus-expressed LRRK2 protein (Thermo Fisher Scientific, A15197 ) in 1 × kinase buffer (Cell Signaling Technologies, 9802S) containing 200 μM ATP (Cell Signaling Technologies, 9804S) for 30 min at 30 °C.

Techniques: Gene Expression, Infection, RNA Sequencing, Expressing, Virus, Western Blot, Positive Control

Journal: Nature Communications

Article Title: Targeting leucine-rich repeat kinase 2 overcomes resistance to oncolytic herpes simplex virus-based therapies in glioblastoma

doi: 10.1038/s41467-026-70132-9

Figure Lengend Snippet: a Anti-myc immunoprecipitates prepared from 293 T cells transfected with the indicated plasmids were subjected to SDS‒PAGE and blotted with the indicated antibodies. The samples derive from the same experiment but different gels for Myc and another for STAT1 were processed in parallel. b Endogenous LRRK2 and STAT1 complexes in U-118 MG cells were isolated by co-immunoprecipitation (co-IP) and detected using anti-LRRK2 antibodies. The samples derive from the same experiment but different gels for LRRK2 and another for STAT1 were processed in parallel. c The interactions between STAT1-Flag and Myc-LRRK2 mutants were detected by a co-IP assay. FL, full-length. The samples derive from the same experiment but different gels for Myc and another for Flag were processed in parallel. d , e The interactions between Myc-LRRK2 and STAT1-HA truncations were detected by a co-IP assay. The samples derive from the same experiment but different gels for Myc and another for HA were processed in parallel. f STAT1-His proteins were incubated with human FL LRRK2 proteins in the presence of ATP. The reaction products were analyzed with the indicated antibodies. CD300A proteins served as a negative control. The samples derive from the same experiment but different gels for Flag and another for His were processed in parallel. g , h 293 T cells were cotransfected with Flag-STAT1 and LRRK2 in the presence or absence of LRRK2-IN-1, and the phosphorylation of STAT1 was analyzed by western blot analysis. The samples derive from the same experiment but different gels for STAT1, another for p-STAT1 701 and another for p-STAT1 727 were processed in parallel. i 293 T cells were cotransfected with Flag-STAT1 and each LRRK2 mutant, either in the presence or absence of LRRK2-IN-1, and STAT1 phosphorylation was assessed via western blot analysis. The samples derive from the same experiment but different gels for STAT1, another for p-STAT1 701 and another for GAPDH were processed in parallel. j Western blotting was performed to assess and quantify phosphorylated STAT1 and HSV-derived GFP in U-87 MG cells treated with fludarabine (10 µM) for 4 h, either in the presence or absence of LRRK2-IN-1, followed by infection with oHSV-GFP (MOI = 1 PFU/cell). The samples derive from the same experiment but different gels for STAT1 and GFP, another for p-STAT1 701 and another for actin were processed in parallel. k Western blotting was performed to assess and quantify phosphorylated STAT1 and HSV-derived GFP in GBM-1 cells. Cells transfected with EV, LRRK2 or LRRK2 D1994A were infected with oHSV-GFP (MOI = 0.5 PFU/cell), with or without LRRK2-IN-1. The samples derive from the same experiment but different gels for STAT1 and GFP, another for p-STAT1 701 and another for actin were processed in parallel. l Western blotting was performed to assess and quantify phosphorylated TBK1, phosphorylated LRRK2, phosphorylated Rab10, and phosphorylated STAT1 in BX795 (10 µM)-treated U-87 cells following infection with oHSV-GFP (MOI = 0.5 PFU/cell). The samples derive from the same experiment but different gels for p-TBK1, another for p-LRRK2, another for LRRK2, another for p-Rab10 and p-STAT1, another for STAT1 and another for actin were processed in parallel. Western blot analyses were independently repeated twice for panels ( a – d and f )and three times for panels ( e , g and i – l ), with similar results. The data are presented as the mean ± SD from three independent experiments ( h – l ), and P values were calculated using an ordinary one-way ANOVA ( h – l ).

Article Snippet: Recombinant human STAT1 protein (ABclonal, RP01251) was incubated with active, baculovirus-expressed LRRK2 protein (Thermo Fisher Scientific, A15197 ) in 1 × kinase buffer (Cell Signaling Technologies, 9802S) containing 200 μM ATP (Cell Signaling Technologies, 9804S) for 30 min at 30 °C.

Techniques: Transfection, Isolation, Immunoprecipitation, Co-Immunoprecipitation Assay, Incubation, Negative Control, Phospho-proteomics, Western Blot, Mutagenesis, Derivative Assay, Infection

Journal: bioRxiv

Article Title: Integrating Artificial Intelligence and Bioinformatics Methods to Identify Disruptive STAT1 Variants Impacting Protein Stability and Function

doi: 10.1101/2024.10.04.616446

Figure Lengend Snippet: Selection of nsSNPs of STAT1 gene.

Article Snippet: Identification of nsSNPs in STAT1 protein domains The FASTA sequence of the STAT1 protein was submitted to the InterPro server ( https://www.ebi.ac.uk/interpro ).

Techniques: Selection

Journal: Endocrine-related cancer

Article Title: Signal transducer and activator of transcription 5A/B in prostate and breast cancers

doi: 10.1677/ERC-08-0013

Figure Lengend Snippet: Chromosomal mapping and sequence identity between human and mouse signal transducer and activator of transcriptions (STATs)

Article Snippet: In the mouse, Stat1 and Stat4 are located on chromosome 1 (band 1 C1.1) ( Schindler et al . 1992 , Yamamoto et al . 1994 ); Stat2 and Stat6 are on chromosome 10 (band 10 D3) ( Fu et al . 1992 , Quelle et al . 1995 ); and Stat3 , Stat5a , and Stat5b map to chromosome 11 (band 11 D) ( Zhong et al . 1994 a , Copeland et al . 1995 , Shi et al . 1996 , Levy et al . 1998 ; ). table ft1 table-wrap mode="anchored" t5 caption a7 STAT Human Mouse % Identity References Amino acid Chromosome Amino acid Chromosome STAT1 750 2 749 1 93 Schindler et al . (1992) and Haddad et al . (1998) STAT2 846 12 915 10 64 Fu et al . (1992) and Goureau et al . (2001) STAT3 769 17 769 11 99 Zhong et al . (1994 a ) and Choi et al . (1996) STAT4 748 2 748 1 94 Yamamoto et al . (1994 , 1997) STAT5A 794 17 793 11 96 Mui et al . (1995) and Lin et al . (1996) STAT5B 787 17 786 11 96 Mui et al . (1995) and Lin et al . (1996) STAT6 847 12 837 10 85 Quelle et al . (1995) and Leek et al . (1997) Open in a separate window Chromosomal mapping and sequence identity between human and mouse signal transducer and activator of transcriptions (STATs) STATs are proteins of 750–900 amino acids (90–115 kDa) with five structurally and functionally conserved domains that allow the transduction of ligand-specific signals ( Schindler & Darnell 1995 ).

Techniques: Sequencing