The Role And Mechanism Of Protein Arginine Methyltransferase 2 In Intestinal Inflammation

Inflammatory bowel disease(IBD)is mainly characterized by recurrent intestinal inflammation,and the quality of life in IBD pantients is seriously affected.Protein arginine methyltransferase(PRMT),a group of common epigenetic-modifying enzymes,is widely involved in physiological and biochemical processes,such as signal transduction,cell activation,inflammation.However,there are few studies on the role of PRMT in IBD,which needs to be investigated,as well as its mechanism.In this study,the expression of PRMT was firstly studied in the colon of mice with dextran sulfate soduim(DSS)-induced colitis.The results showed that the expression of PRMT2 in intestinal epithelium was significantly elevated.Further studies showed that the expression of protein arginine methyltransferase 2(PRMT2)was also elevated in intestinal samples from patients with IBD.Next,the role and mechanism of PRMT2 in DSS-induced colitis and tumor necrosis factor-?(TNF-?)-induced intestinal epithelial cells were explored by lentivirus-mediated PRMT2 overexpression or knockdown.At last,the effect of PRMT2 on the expression of cathelicidin-related antimicrobial peptide(CRAMP)was investigated.Recombinant bacteria that expressing CRAMP were constructed with Lactococcus lactis NZ9000(L.lactis NZ9000),and its protective effect and mechanism were explored in mice with colitis.The main results are as follows:(1)Mice were treated with DSS to induce intestinal inflammation and the expression of PRMT1?PRMT7 was tested respectively.The results showed that the expression of PRMT2 and PRMT5 was obviously elevated in the colon of mice with DSS-induced colitis.The expression of PRMT2 was elevated more significantly.Further more,the study showed that the expression of PRMT2 was elevated in colonic epithelium of DSS-treated mice.In order to verify the correlation between PRMT2 and clinical IBD,the expression of PRMT2 was detected in intestinal samples from IBD patients and healthy people.The results showed that the expression of PRMT2 in intestinal epithelium from patients with UC and CD was higher than that of healthy people.(2)PRMT2 was overexpressed or knocked down by lentivirus in mice before DSS treatment,so as to reveal the role of PRMT2 in mice with DSS-induced colitis.The results showed that overexpression of PRMT2 aggravated the symptoms of colitis in mice,as evidenced by increased weight loss,elevated disease activity index,shortened colon length and increased intestinal immune cell infiltration in PRMT2 overexpression mice.Overexpression of PRMT2 increased the expression of TNF-?,C-X-C motif chemokine ligand 1(CXCL-1),C-X-C motif chemokine ligand 12(CXCL-12),C-C motif chemokine ligand 2(CCL-2),C-C motif chemokine ligand 5(CCL-5)and C-C motif chemokine ligand20(CCL-20)in the colon of mice.Overexpression of PRMT2 increased intestinal permeability in mice,decreased the expression of zona occludens(ZO-1)and claudin-1,increased the expression of claudin-2.In vitro cell experiments showed that PRMT2 enhanced TNF-?-induced expression of inflammatory cytokines in intestinal epithelial cells.Consistently,PRMT2 knockdown attenuated intestinal injury in mice with DSS-induced colitis and decreased the expression of inflammatory cytokines in intestinal epithelial cells induced by TNF-?.(3)Mice with DSS-induced colitis and TNF-? induced intestinal epithelial cells were used after lentivirus-mediated PRMT2 overexpression or knockdown,so as to reveal the mechanism of PRMT2 in intestinal inflammation.The results showed that PRMT2 catalyzed asymmetric dimethylation of arginine 8 in histone H3(H3R8me2a).The binding of H3R8me2 a to the promoter region of suppressor of cytokine signaling molecule 3(SOCS3)reduced the expression of SOCS3.Thus,the ubiquitination of tumor necrosis factor receptor-associated factor 5(TRAF5)midiated by SOCS3 was reduced,which led to the increasing of TRAF5 expression,as well as TRAF5 mediated nuclear factor-?B(NF-?B)and mitogen-activated protein kinase(MAPK)activation.(4)In mice with DSS-induced colitis and TNF-?-induced intestinal epithelial cells,PRMT2 was overexpressed or knocked down,and the expression of CRAMP was tested,so as to investigate whether PRMT2 can affect the expression of CRAMP.The results showed that PRMT2 overexpression decreased the expression of CRAMP,while PRMT2 knockdown increased the expression of PRMT2.In addition,CRAMP was recombined with L.lactis NZ9000,and the recombined strains were given to colitis mice by gavage.The results showed that CRAMP expressing L.lactis NZ9000 alleviated colitis in mice,as proved by reduced weight loss,decreased disease activity index,maintained colon and reduced inflammatory cell infiltration.CRAMP expressing L.lactis NZ9000 reduced the expression of proinflammatory cytokines interleukin-6(IL-6),interleukin-1?(IL-1?)and TNF-?.Meanwhile,the expression of regulatory cytokine interleukin-10(IL-10),ZO-1,zona occludens 2(ZO-2)and occludin were increased.Regarding the mechanism,CRAMP expressing L.lactis NZ9000 decreased the activation of mitogen-activated protein kinase p38(p38)and nuclear factor ?B p65(p65)in colon of mice.In conclusion,PRMT2 reduced SOCS3 expression by increasing H3R8 asymmetric methylation at SOCS3 promoter,which led to reduced SOCS3-mediated degradation of TRAF5 via ubiquitination.The expression of TRAF5 was elevated,which led to TRAF5-mediated NF-?B and MAPK activation.In addition,PRMT2 also aggravated colitis by reducing the expression of CRAMP.And CRAMP expressing L.lactis NZ9000 alleviated colitis by reducing the activation of p38 and p65.