Arginine Di-methylation of RIPK3 Safeguards Necroptosis for Intestinal Homeostasis

AbstractThe necroptosis mediated by RIPK3 is stringently regulated for intestinal homeostasis. Here we found that mice lackingPrmt5(Protein arginase methyltransferase 5) in intestinal epithelial cells (IECs) caused premature death with IECs necroptosis, villus atrophy and loss of Paneth cells. This pathology can be partially rescued by antibiotic treatment, germ-free breeding condition and pharmaceutical inhibition of RIPK1 and RIPK3, but aggravated for embryonic lethality byCaspase-8 deficiency, which demonstrating the importance of commensal bacteria and necroptosis for thePrmt5-IEC deficiency. Intriguingly, tumor-necrosis factor (TNF) receptor 1(Tnfr1) deficiency could not completely rescue the pathology, and mice deficit in Z- DNA binding protein 1(ZBP1) exhibited shorter lifespan compared withPrmt5null mice, suggestingPrmt5loss might trigger TNFR-RIPK1-depenfent and ZBP1- dependent necroptosis. Mechanically, we identified the 479-arginine residue of RIPK3 di-methylated by PRMT5 was an endogenous checkpoint for necroptosis. Furthermore, RIPK3-R479K mutation had higher affinity with both RIPK1 and ZBP1 by immunoprecipitation and STORM (Stochastic Optical Reconstruction Microscopy) analysis, which might explain the endogenous necroptosis triggered by mutated RIPK3 even without upstream stimuli. Moreover, the peptide of RIPK3-SDMA (Symmetric dimethylarginine of 479) could rescue lethality ofPrmt5 lacking mice through necrosome formation inhibition, which demonstrating the great potential for necroptosis-related disease treatment through RIPK3 dimethylation targeting.