Fig. 8: SIRT6 promotes the Tyr641 phosphorylation of STAT6 via inhibiting SOCS3 expression.

a–c Western blot (a) and quantification (b) and qPCR (c) analyses of SOCS3 expression in the jejunal IECs from 2-month-old male C57BL/6J mice on indicated days post-H. poly infection. n = 3 mice/group in (b) and 5 mice/group in (c); p = 0.0499 (6 dpi) in (b); p = 0.0002 (6 dpi), <0.0001 (9 dpi), 0.0058 (14 dpi) in (c). d–f Western blot (d) and quantification (e) and qPCR (f) analyses of SOCS3 expression in the jejunal IECs from 2–3-month-old naive and H. poly-infected LoxP and IEC-KO mice. n = 3 mice/group in (e) and 4 mice/group in (f); from left to right, #p = 0.0012 in (e); *p = 0.0493, 0.0378; #p = 0.0233, 0.0241 in (f). g–i Western blot (g) and quantification (h) and qPCR (i) analyses of SOCS3 expression in the jejunal IECs from 3-month-old male WT and IEC-Tg mice infected with H. poly. n = 6 mice/group in (h) and 7 mice/group in (i); p = 0.0073 in (h). j Schematic illustration of the primer amplicons (Regions 1–3) used for scanning ChIP analysis of the proximal SOCS3 promoter. k, l The enrichment of SIRT6 (k) and the effects of SIRT6 overexpression or knockdown on H3K56 acetylation (l) within the SOCS3 proximal promoter were analyzed using ChIP in rIL13-treated (10 ng/ml, 6  h) NCM460 cells. In (k), n = 6 (Region-1&−3) and 5 (Region-2) biological replicates/group; p = 0.0257 (Region-1), 0.0002 (Region-2); In (l), n = 6 biological replicates/group; GFP vs SIRT6, p = 0.0136 (Region-2); shGFP vs shSIRT6, p = 0.0011 (Region-1), 0.0101 (Region-2). m, n The effects of SOCS3 overexpression on STAT6 (Y641) phosphorylation were examined by western blot in rIL13-treated (10 ng/ml, 6 h) HEK293T (m) and NCM460 (n) cells. o Quantification analysis of P-STAT6 (Y641) levels shown in (m, n). n = 3 biological replicates/group; p < 0.0001 (HEK293T), 0.0107 (NCM460). p–s Western blot (p, r) and quantification (q, s) analyses of P-STAT6 (Y641), SIRT6 and SOCS3 in NCM460 cells transfected with shSIRT6 and/or shSOCS3 constructs (p, q) or SIRT6 and/or SOCS3 constructs (r, s), followed with rIL13 stimulation (10 ng/ml, 6 h). n = 3 biological replicates/group; In (q), from left to right, *p = 0.0039; #p = 0.0017 (P-STAT6); *p = 0.0491, 0.0263, 0.0306; #p = 0.0011 (SOCS3); In (s), from left to right, *p = 0.0466, 0.0101, 0.0096; #p = 0.0141 (P-STAT6); #p = 0.0457 (SOCS3). t, u p2xSTAT6-Luc2P luciferase reporter assays in HEK293T cells transfected with shSIRT6 and/or shSOCS3 constructs (t) or SIRT6 and/or SOCS3 constructs (u), followed with vehicle or rIL13 stimulation (10 ng/ml, 6 h). n = 4 biological replicates/group. In (t), from left to right, p = 0.0098, 0.0075, 0.005, 0.0343, 0.0027, respectively. In (u), from left to right, p = 0.0459, 0.0416, 0.0003, 0.003, respectively. v Schematic illustration of the role of IEC SIRT6 in regulation of helminth-induced epithelial remodeling. Data are presented as mean ± SEM. All p values were generated by two-tailed unpaired t test. *p < 0.05; In (b, c), *p < 0.05 vs naive; In (e, f), *p < 0.05 vs LoxP-naive, #p < 0.05 vs LoxP-H. poly; In (q), *p < 0.05 vs shGFP, #p < 0.05 vs shSOCS3; In (s), *p < 0.05 vs GFP, #p < 0.05 vs SOCS3-FLAG. Source data are provided as a Source Data file.