Ubiquitin Ligase MARCH8 attenuates Graft versus Host Disease via Regulation of Gut Epithelial Cell Surface MHC II Expression.

Abstract

Allogeneic stem cell transplantation (SCT) is the preferred therapy for the majority of haematological malignancies, but its use is limited by graft-versus-host disease (GVHD), the primary life threatening complication associated with this procedure. Donor T cells, which are primed against host tissue antigens, are critical mediators of GVHD. Recently we reported that class II dependent priming of donor CD4+ T cells by host non-haematopoietic cells is sufficient to elicit GVHD and that early after SCT the intestinal tract represents the primary site of alloantigen presentation. The membrane-associated RING-CH (MARCH) family of E3 ubiquitin ligases function as immune regulators by limiting the expression of key immune receptors. Notably, March8 limits the expression of MHC II on non-haematopoietic cells by ubiquitinating MHC II and thus targeting it for degradation through endosomal-proteases. This process involves inclusion of the ubiquitinated proteins in intraluminal vesicles via the ESCRT system. In this study we demonstrate constitutive, non-haematopoietic-restricted expression of March8 mRNA throughout the intestinal tract, and elevated cell surface MHC II expression on intestinal epithelial cells (IEC) from MARCH8 deficient (MARCH8-/-) mice compared to wild type (WT) mice. Histological comparison of the gut between MARCH8-/- and WT mice appeared normal. Further to this, colonic crypts were cultured from both MARCH8-/- and WT mice suggesting no difference in their capacity for growth. Using the BALB/c into B6 model of GVHD, MARCH8-/- recipients exhibited significantly increased clinical scores and reduced survival with 100% of MARCH8-/- recipients succumbing to disease by D9 post-transplant while more than 60% WT recipient mice survive beyond D40 (p= 0.0062). Examination of the gut at day 7 after SCT demonstrated significantly increased pathology in March8-/- recipients associated with a marked increase in donor T cell expansion. This was donor T cell dependent, as March8-/- recipients of T cell depleted grafts survive long term (D60+). Taken together our data identifies a critical role for MHC II antigen presentation by IEC in the initiation of GVHD, and an immunomodulatory role for MARCH8 for the control of IEC MHC II expression and GVHD severity.