The role of ADAM10 in the regulation of T cell responses in Mus musculus

Abstract

ADAM10 (A Disintegrin And Metalloproteinase 10) is an ubiquitously expressed membrane-bound metalloproteinase involved in ectodomain shedding of about 100 so far identified membrane proteins, including cytokines, receptors as well as growth factors and adhesion molecules. Being the principal sheddase of Notch receptors, ADAM10 orchestrates the Notch-signaling pathway, and thus plays a crucial role in embryonic development, tissue homeostasis and cell fate decisions. ADAM10 is constitutively expressed in T cells and was found to cleave a number of proteins, in addition to Notch1 and Notch2, that might be relevant for T cells. Although these substrates indicate a role of ADAM10 in T cell function, the relevance of ADAM10 for T cell responses is still largely unclear. To study the role of ADAM10 in vivo, Adam10fl/flxCD4Cre (A10/CD4Cre) mice with a T cell restricted deletion of Adam10 were used for the assessment of known and identification of novel substrates of ADAM10, as well as a phenotypical and functional analysis of ADAM10-deficient conventional CD4 and CD8 T cells, regulatory T cells (Tregs) and Natural Killer (NK) T cells under homeostasis and after infection.

It was possible to prove CD40L, CD44, FasL, ICOS-L, IL-23R, IL-2Rα, LAG3, NRP1, PD-L1 and Tim-3 as ADAM10 targets on T cells. In addition, CD27 and ST2 were identified as potential novel ADAM10 targets on T cells. Characterization of T cell subsets in A10/CD4Cre mice revealed a general shift towards a more activated phenotype in conventional CD4 and CD8 T cells as well as Tregs and NK T cells under homeostasis, marked by an enhanced expression of activation-associated markers and effector cytokines. A consistent shift towards a type 3/Th17 phenotype was observed in all T cell subsets. In addition, a shift towards CD4neg NK T cells and an enlarged Treg population were detected. Yet, despite this bias to more activated T cells and to Th17 cells, ADAM10-deficient mice were similar or even less efficient in the control of Candida albicans and Staphylococcus aureus, two pathogens for which protection depends on Th17 cells. The infection with Listeria monocytogenes not only allowed to analyze the general immune response, but also to detect de novo-generated listeria-specific Th1 cells and CD8 T cells. A10/CD4Cre mice showed a defective listeria-specific T cell response, marked by a lower percentage of listeria-specific CD4 and CD8 T cells. A lower absolute number of cytokine+ NK T cells and a reduced serum cytokine level was also observed in A10/CD4Cre mice after injection of the NK T cell antigen α-galactosylceramide (α-GalCer). Still, A10/CD4Cre mice developed a significantly aggravated α-GalCer-induced liver inflammation, as indicated by large areas of liver damage visible in histology and significantly higher AST and ALT serum levels. Analysis of NK T cells revealed enhanced surface expression of FasL on NK T cells but also on conventional T cells in A10/CD4Cre mice treated with α-GalCer. As FasL directly kills Fas expressing hepatocytes, the observed enhanced FasL expression could be responsible for the aggravated liver damage in A10/CD4Cre mice.

As ADAM10 is the major sheddase responsible for the cleavage of Notch1 and Notch2, T cells of Notch1/2/CD4Cre mice were compared to those of A10/CD4Cre mice. Likewise, Notch1/2/CD4Cre mice revealed a shift towards CD4neg cells in NK T cells and an enlarged Treg population. However, a generally more activated phenotype and the prominent shift towards a type 3 phenotype in all T cell subsets was not observed in Notch1/2/CD4Cre mice. Thus, impaired Notch signaling can only be partially responsible for the altered function of ADAM10-deficient T cells and other, so far unknown, ADAM10 substrates are involved in regulation of T cells.

In conclusion, ADAM10 was found to cleave numerous T cell-relevant surface proteins and was shown to play a pivotal role for the function of conventional CD4 and CD8 T cells as well as Tregs and NK T cells.