Journal: bioRxiv

Article Title: Ecological determinants of disease and immunity in myelodysplastic syndromes

doi: 10.64898/2026.05.05.720208

Figure Lengend Snippet: a, Bar plots of relative median fluorescence intensity (MFI) of granzyme B and perforin from intracellular cytokine staining of primary untreated MDS CD8 T cells after exposure to increasing doses of TGFβ. Relative MFI values are shown as mean ± SD. Data from n=6 MDS patients. Mann-Whitney U test used for statistical analysis (*p<0.05, **p<0.01). b, Top, volcano plot of differentially expressed genes in CD8 memory T cells close to (≤20µm) versus far from (>50µm) megakaryocytes. Bottom, schematic of CD8 memory T cells based on their distance to nearest megakaryocytes from spatial data. c, Relative percent spliced in (dPSI) value of differential 3’ splice site in mRNAs from SF3B1 mutant MDS patient bulk RNA-seq versus SF3B1 wild-type MDS. Highlighted names indicate mRNAs encoding proteins involved in TGFβ signaling. d, Diagram of proteins (in green) involved in TGFβ signaling which undergo aberrant RNA splicing in SF3B1 mutant MDS. e, Protein diagram of TGFBR1 with red indicating insertion of four amino acids (GPFS) encoded by the long mRNA isoform promoted in SF3B1 mutant cells. f, Alpha fold model of mutant TGFBR1 (green) overlaid with published crystal structure of wild-type TGFBR1 (gray). The GPFS amino acid insertion seen in SF3B1 mutant cells is indicated in red in the inset. g, Percentage (%) of phospho-SMAD2 Serine 465/467 (pS465/467) in K562 cells with the indicated genetic alterations in SF3B1 or TGFBR1. Mean ± SD. Two-way ANOVA. ***p<0.001, ****p<0.0001. h , Representative flow cytometry histograms of p-SMAD2 S465/467 from (g). i, Schematic of an SF3B1-mutant megakaryocyte with increased TGFβ production suppressing cytotoxic activity of nearby CD8 + T cells (cell-extrinsic effect); mutant cell simultaneously exhibits impaired TGFβ sensing (cell-intrinsic effect).

Article Snippet: The K562 human myeloid leukemia cell line was purchased from American Type Culture Collection (ATCC; #CCL-243).

Techniques: Fluorescence, Staining, MANN-WHITNEY, Mutagenesis, RNA Sequencing, Flow Cytometry, Activity Assay

Journal: bioRxiv

Article Title: Ecological determinants of disease and immunity in myelodysplastic syndromes

doi: 10.64898/2026.05.05.720208

Figure Lengend Snippet: (a) Sashimi plots of bulk RNA-sequencing data of an aberrant 3’ splice site usage in TGFBR1, MAP3K7 and SMURF2 mRNA in SF3B1 mutant acute myeloid leukemia (AML) (top; n=76 patients), SF3B1 wild-type (WT) AML (middle; n=739 patients), and normal bone marrow (bottom; n=26 patients). Red lines indicate SF3B1 mutant-specific junctions while black lines represent junction spanning reads in wild-type cells. The number of reads is listed, and the frequency of reads is in parentheses. b , Crystal structure of the short-isoform of TGFBR1 (grey) overlaid with that of the alpha-fold predicted model of SF3B1 mutant induced long-isoform (green). c, RT-PCR analysis of aberrant 3’ splice events in TGFBR1, MAP3K7, and SMURF2 in human isogenic K562 cells with knockin of SF3B1 K700E mutation. d , RT-PCR analysis of endogenous TGFBR1 , MAP3K7, and SMURF2 splicing in primary samples in healthy bone marrow control patients (n=5), patients with SF3B1 K700E mutant MDS (n=5) and non-splicing factor mutant patients with MDS (n=5). e , Sanger sequencing electropherogram of the top and bottom PCR products from gel-purified TGFBR1 RT-PCRs from MDS SF3B1 mutant patients shown in ( c ). The red box highlights the alternatively spliced sequence in the top band, which includes a 12-nucleotide insertion predominantly observed in SF3B1 K700E mutant MDS patients. The bottom band sequence is displayed below, showing the canonical exonic sequence. f , Western blot of K562 cells with SF3B1 mutation, TGFBR1 knockout (KO), or TGFBR1 KO with addback of TGFBR1 cDNA encoding the long or short isoform. g, Western blot of cytoplasmic, membrane, and soluble nuclear fractions of K562 cells with TGFBR1 KO alone or with overexpression of TGFBR1 cDNA encoding the long or short isoform.

Article Snippet: The K562 human myeloid leukemia cell line was purchased from American Type Culture Collection (ATCC; #CCL-243).

Techniques: RNA Sequencing, Mutagenesis, Reverse Transcription Polymerase Chain Reaction, Knock-In, Control, Sequencing, Purification, Western Blot, Knock-Out, Membrane, Over Expression

Journal: Oncology Reports

Article Title: Knockdown of CCT2 inhibits the malignant progression of hepatocellular carcinoma cells by impairing STAT3 activation

doi: 10.3892/or.2026.9086

Figure Lengend Snippet: Knockdown of CCT2 inhibits STAT3 signaling activation in hepatocellular carcinoma cells. The protein levels of STAT3, p-STAT3, MCL1, MMP2 and SOX2 in (A) Huh-7 and (B) HCCLM3 cells were measured by western blotting. The protein levels of (C) p-STAT3, (D) MCL1, (E) MMP2 and (F) SOX2 in subcutaneous tumor tissue were detected by immunohistochemical staining. *P<0.05, **P<0.01 vs. sh-NC. CCT2, chaperonin containing TCP1 subunit 2; sh, short hairpin; NC, negative control; p, phosphorylated; MCL1, myeloid cell leukemia sequence 1; SOX2, SRY-box transcription factor 2.

Article Snippet: The primary antibodies were as follows: CCT2 (cat. no. 24896-1-AP), β-actin (cat. no. 66009-1-Ig), MMP2 (cat. no. 10373-2-AP), myeloid cell leukemia sequence 1 (MCL1; cat. no. 16225-1-AP) and SRY-box transcription factor 2 (SOX2; cat. no. 11064-1-AP; all Proteintech Group, Inc.) and STAT3 (cat. no. 4904) and phosphorylated (p-)STAT3 (Tyr705; cat. no. 4113; both Cell Signaling Technology, Inc.) The membranes were washed three times in TBST (0.1% Tween-20) for 5 min each at room temperature.

Techniques: Knockdown, Activation Assay, Western Blot, Immunohistochemical staining, Staining, Negative Control, Sequencing