Ectodomain shedding of CD99 within highly conserved regions is mediated by the metalloprotease meprin β and promotes transendothelial cell migration

Abstract

ABSTRACT:The adhesion molecule CD99 is essential for the transendothelial migration of leukocytes. In this study, we used biochemical and cellular assays to show that CD99 undergoes ectodomain shedding by the metalloprotease meprin b and subsequent intramembrane proteolysis by g-secretase. The cleavage site in CD99 was identified by mass spectrometry within an acidic region highly conserved through different vertebrate species. This finding fits perfectly to the unique cleavage specificity of meprin b with a strong preference for aspartate residues and suggests coevolution of protease and substrate. We hypothesized that limited CD99 cleavage by meprin b would alter cellular transendothelial migration (TEM) behavior in tissue remodeling processes, such as inflammation and cancer. Indeed, meprin b induced cell migration of Lewis lung carcinoma cells in an in vitro TEM assay. Accordingly, deficiency of meprin b in Mep1b 2/2 mice resulted in significantly increased CD99 protein levels in the lung. Therefore, meprin b could serve as a therapeutic target, given that in a proof-of-concept approach we showed accumulation of CD99 protein in lungs of meprin b inhibitortreated mice.-Bedau, T., Peters, F., Prox, J., Arnold, P., Schmidt, F., Finkernagel, M., Köllmann, S., Wichert, R., Otte, A., Ohler, A., Stirnberg, M., Lucius, R., Koudelka, T., Tholey, A., Biasin, V., Pietrzik, C. U., Kwapiszewska, G., Becker-Pauly, C. Ectodomain shedding of CD99 within highly conserved regions is mediated by the metalloprotease meprin b and promotes transendothelial cell migration. FASEB J. 31, 1226-1237 (2017). www.fasebj.orgRegulated intramembrane proteolysis (RIP) of cell adhesion molecules, such as junctional adhesion molecule (JAM)-A, intercellular adhesion molecule (ICAM)-1, and L-selectin, was shown to be essential for transendothelial migration (TEM) of inflammatory or cancer cells (1). Meprin b, a multidomain type I transmembrane metalloprotease, is an initiator of RIP, and structural studies revealed dimeric formation of the protease with the active site in proximity to the cell surface (2-4). In addition, meprin b can be shed from the cell surface by ADAM10/ 17, resulting in a soluble active protease, which for instance is important for mucus detachment in the small intestine (5). Meprin b is characterized by a unique cleavage specificity, with a preference for negatively charged amino acids (6). These structural features provide all requirements that meprin b must have to act as an ectodomain sheddase at the cell surface. Indeed, membrane-bound amyloid precursor protein (APP), for instance, is cleaved by meprin b, resulting in the release of sAPP-b fragments and neurotoxic Ab peptides (4,7,8). Many of the known substrates of meprin b have been identified by mass spectrometry (MS)-based proteomic approaches (9).

Meprin β cleaves TREM2 and controls its phagocytic activity on macrophages

“…Meprin β has been linked to inflammatory bowel disease 47 and it was shown to shed the IL‐6R and to induce pro‐inflammatory trans‐signaling 27 . Meprin β promotes transendothelial migration of cells through CD99 cleavage, an important step for immune cells to reach the site of inflammation 28 . Additionally, meprin β can cleave, and thereby inactivate IL‐6 48 and the important chemokine CCL2/MCP‐1 leading to a reduced monocyte attraction 49 …”

“…Meprin β belongs to the astacin family of metalloproteinases and exhibits striking cleavage specificity with a preference for negatively charged amino acids around the scissile bond 25 . Similar to ADAM10 and ADAM17, meprin β was found to cleave several cell surface proteins such as the amyloid precursor protein (APP) at the β‐secretase site, 26 the IL‐6 receptor on human granulocytes to induce IL‐6 trans‐signaling, 27 and CD99 on endothelial cells, thereby promoting transendothelial cell migration 28,29 …”

Proteolytic ectodomain shedding of membrane proteins in mammals—hardware, concepts, and recent developments

“…Several TM proteins were cleaved within their ectodomain at a large distance from the membrane, which is not seen as a shedding event, as a large part of the ectodomain remains. However, meprin β also acts as a sheddase and cleaves close to the TM domain in CD99 to promote transendothelial cell migration, and in APP, for which it acts as an alternative β‐secretase (Jefferson et al , ; Arolas et al , ; Bedau et al , ). To what extent meprin β may contribute to Alzheimer's disease still needs to be explored in more detail.…”

Cancer-associated mutations in the canonical cleavage site do not influence CD99 shedding by the metalloprotease meprin β but alter cell migration in vitro

“…Recently, CD99 was found to be specifically cleaved by the metalloprotease meprin β [ 8 , 9 ]. Importantly, ectodomain shedding of CD99 by meprin β and subsequent regulated intramembrane proteolysis by γ-secretase resulted in increased TEM of murine Lewis lung carcinoma (LLC) cells.…”