Immune dysregulation in SHARPIN-deficient mice is dependent on CYLD-mediated cell death.

Abstract

SHARPIN, together with RNF31/HOIP and RBCK1/HOIL1, form the linear ubiquitin chain assembly complex (LUBAC) E3 ligase that catalyzes M1-linked polyubiquitination. Mutations in <i>RNF31/HOIP</i> and <i>RBCK/HOIL1</i> in humans and <i>Sharpin</i> in mice lead to autoinflammation and immunodeficiency, but the mechanism underlying the immune dysregulation remains unclear. We now show that the phenotype of the <i>Sharpin^cpdm/cpdm</i> mice is dependent on CYLD, a deubiquitinase previously shown to mediate removal of K63-linked polyubiquitin chains. Dermatitis, disrupted splenic architecture, and loss of Peyer's patches in the <i>Sharpin^cpdm/cpdm</i> mice were fully reversed in <i>Sharpin^cpdm/cpdm Cyld^-/-</i> mice. We observed enhanced association of RIPK1 with the death-signaling Complex II following TNF stimulation in <i>Sharpin^cpdm/cpdm</i> cells, a finding dependent on CYLD since we observed reversal in <i>Sharpin^cpdm/cpdm Cyld^-/-</i> cells. Enhanced RIPK1 recruitment to Complex II in <i>Sharpin^cpdm/cpdm</i> cells correlated with impaired phosphorylation of CYLD at serine 418, a modification reported to inhibit its enzymatic activity. The dermatitis in the <i>Sharpin^cpdm/cpdm</i> mice was also ameliorated by the conditional deletion of <i>Cyld</i> using <i>LysM-cre</i> or <i>Cx3cr1-cre</i> indicating that CYLD-dependent death of myeloid cells is inflammatory. Our studies reveal that under physiological conditions, TNF- and RIPK1-dependent cell death is suppressed by the linear ubiquitin-dependent inhibition of CYLD. The <i>Sharpin^cpdm/cpdm</i> phenotype illustrates the pathological consequences when CYLD inhibition fails.