FAM49B Fragmentation by Asparagine Endopeptidase Promotes Vascular Smooth Muscle Cell Migration in Atherogenesis.

Background: The migration of vascular smooth muscle cells (VSMCs) is critical for the development of atherosclerosis. However, the underlying molecular mechanisms are not completely understood. Here, we detected FAM49B (family with sequence similarity 49 member B) fragments in atherosclerotic plaques and identified their roles in VSMC migration and atherogenesis.

Methods: Transgenic mice such as Aep (asparagine endopeptidase)^-/-, Aep^-/-Apoe^-/-, and VSMC-specific full-length FAM49B and FAM49B fragment overexpression by adenovirus gene transfer were used to determine the role of FAM49B fragments in atherosclerosis. In addition, the effects of compound 11, an AEP inhibitor, on the progression of atherosclerosis in Apoe^-/- mice were analyzed. FAM49B fragments were identified by mass spectrometry. Moreover, the expression of FAM49B fragments in atherosclerotic plaques from mice and patients was analyzed by immunofluorescence and immunoblotting.

Results: The levels of FAM49B are increased in atherosclerotic lesions. Interestingly, FAM49B is cleaved by the cysteine protease AEP at residues N169 and N170, generating 2 fragments: FAM49B (1-169) and FAM49B (171-324). Both fragments are upregulated in VSMCs with the development of atherosclerotic plaques. The overexpression of full-length FAM49B inhibits the migration of human aortic VSMCs, whereas the overexpression of FAM49B fragments promotes VSMC migration. FAM49B fragments bind to Rac1 (Ras-related C3 botulinum toxin substrate 1) and increase its activity, thereby inducing actin polymerization and promoting cell migration. The overexpression of FAM49B fragments in mouse aortic VSMCs results in a higher atherosclerotic plaque burden, whereas the deletion of AEP blocks FAM49B fragmentation and decreases plaque size in mouse models of atherosclerosis. Furthermore, the administration of compound 11 blocked FAM49B fragmentation and alleviated atherosclerotic lesions.

Conclusions: Our results indicate that AEP-derived FAM49B fragments facilitate Rac1-mediated VSMC migration and promote atherosclerosis progression. Inhibiting AEP-mediated FAM49B fragmentation may be a therapeutic strategy for atherosclerosis.