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

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 RNF31/HOIP and RBCK/HOIL1 in humans and Sharpin in mice lead to autoinflammation and immunodeficiency, but the mechanism underlying the immune dysregulation remains unclear. We now show that the phenotype of the Sharpin cpdm/cpdm 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 Sharpin cpdm/cpdm mice were fully reversed in Sharpin cpdm/cpdm Cyld -/- mice. We observed enhanced association of RIPK1 with the death-signaling Complex II following TNF stimulation in Sharpin cpdm/cpdm cells, a finding dependent on CYLD since we observed reversal in Sharpin cpdm/cpdm Cyld -/- cells. Enhanced RIPK1 recruitment to Complex II in Sharpin cpdm/cpdm cells correlated with impaired phosphorylation of CYLD at serine 418, a modification reported to inhibit its enzymatic activity. The dermatitis in the Sharpin cpdm/cpdm mice was also ameliorated by the conditional deletion of Cyld using LysM-cre or Cx3cr1-cre 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 Sharpin cpdm/cpdm phenotype illustrates the pathological consequences when CYLD inhibition fails.