Genetically Determined Proteolytic Cleavage Modulates α7β1 Integrin Function

Abstract

The dystrophin-glycoprotein complex and the ␣7␤1 integrin are trans-sarcolemmal linkage systems that connect and transduce contractile forces between muscle fibers and the extracellular matrix. ␣7␤1 is the major laminin binding integrin in skeletal muscle. Different functional variants of this integrin are generated by alternative splicing and post-translational modifications such as glycosylation and ADP-ribosylation. Here we report a species-specific difference in ␣7 chains that results from an intra-peptide proteolytic cleavage, by a serine protease, at the 603 RRQ 605 site. Site-directed mutagenesis of RRQ to GRQ prevents this cleavage. This RRQ sequence in the ␣7 integrin chain is highly conserved among vertebrates but it is absent in mice. Protein structure modeling indicates this cleavage site is located in an open region between the ␤-propeller and thigh domains of the ␣7 chain. Compared with the non-cleavable ␣7 chain, the cleaved form enhances cell adhesion and spreading on laminin. Cleavage of the ␣7 chain is elevated upon myogenic differentiation, and this cleavage may be mediated by urokinase-type plasminogen activator. These results suggest proteolytic cleavage is a novel mechanism that regulates ␣7 integrin functions in skeletal muscle, and that the generation of such cleavage sites is another evolutionary mechanism for expanding and modifying protein functions.Integrins are ␣,␤-heterodimeric membrane receptors for extracellular matrix proteins (for reviews, see Refs. 1-3). They are used by cells to sense and modify their environments and they are involved in a wide range of cellular processes including cell adhesion, migration, differentiation, proliferation, apoptosis, and cancer metastasis (for reviews, see . Integrins are present in all metazoans and they are highly conserved in structure and function. In general, both ␣ and ␤ subunits have a short cytoplasmic domain, a large N-terminal extracellular domain, and a single hydrophobic transmembrane segment (9). The N-terminal of the ␣ subunit contains seven FG-GAP repeats forming a ␤-propeller domain that is important for ligand binding (10).The ␣7 chain is synthesized as a single 1135-amino acid polypeptide precursor, and like other integrin ␣ chains, it is cleaved within the cell to form a heavy (Ϸ100 kDa) and a light (Ϸ30 kDa) chain connected by a disulfide bond (11). ␣7 associates with ␤1 subunits and the ␣7␤1 integrin is expressed in skeletal and smooth muscle cells, neurons, Schwann cells, and cardiomyocytes where it functions as a receptor for laminin (12)(13)(14). Expression of ␣7 integrin in skeletal muscle is developmentally regulated at the transcriptional level and by alternative splicing, resulting in at least two extracellular (X1 and X2) and two cytoplasmic isoforms (A and B) (12,13,(15)(16)(17). The ␣7A cytoplasmic isoform is only found in skeletal muscle, and it is enriched at myotendinous and neuromuscular junctions (12,13,15,18). Unlike ␣7A, the ␣7B isoform is found throughout the sarcolemma and it is also expressed in other c...

Short form of α9 promotes α9β1 integrin-dependent cell adhesion by modulating the function of the full-length α9 subunit

“…In α6 and α7, the N-terminal cleaved product has β-propeller and partial thigh domains. Processing of α6 and α7 is important for inside-out signaling and enhanced cell adhesion, respectively [38, 39], similar to the results described here.…”

Levels of α7 integrin and laminin-α2 are increased following prednisone treatment in themdxmouse and GRMD dog models of Duchenne muscular dystrophy

“…The lack of negative feedback suppression of Itga7 transcription in the mdx mouse might allow for greater average α7 integrin protein levels at the sarcolemma than in other models. The mouse α7 integrin protein is more stable, lacking a secondary extracellular protease cleavage site that is conserved in rats, dogs and humans (Liu et al, 2008b). This non-cleavable mouse α7 integrin could enable the mdx mouse to stabilize their sarcolemma by reducing α7 integrin protein turnover, thus preventing the dystrophic progression.…”

Thiol switches in membrane proteins - Extracellular redox regulation in cell biology

“…The integrins, α4β1, αMβ2, and αIIbβ3, were among the first redox-regulated integrins to be discovered, with consequences on leukocyte integrin activation and platelet aggregation (Blouin et al 1999;Laragione et al 2003;Liu et al 2008;Murphy et al 2014;Yan and Smith 2000). Many of these studies revealed re-shuffling as well as reducing and reforming of disulfide bonds within the EGF domains of the cysteine-rich integrin β chain (Hu and Luo 2018;Mor-Cohen et al 2012).…”