Xin repeats define a novel actin-binding motif

Abstract

Xin is a protein that is expressed during early developmental stages of cardiac and skeletal muscles. Immunolocalization studies indicated a peripheral localization in embryonic mouse heart, where Xin localizes with β-catenin and N-cadherin. In adult tissues, Xin is found primarily in the intercalated discs of cardiomyocytes and the myotendinous junctions of skeletal muscle cells, both specialized attachment sites of the myofibrillar ends to the sarcolemma. A large part of the Xin protein consists of unique 16 amino acid repeats with unknown function. We have investigated the characteristics of the Xin repeats by transfection experiments and actin-binding assays and ascertained that, upon expression in cultured cells, these repeats bind to and stabilize the actin-based cytoskeleton. In vitro co-sedimentation assays with skeletal muscle actin indicated that they not only directly bind actin filaments, but also have the capability of arranging microfilaments into networks that sediment upon low-speed centrifugation. Very similar repeats were also found in `Xin-repeat protein 2' (XIRP2), a novel protein that seems to be expressed mainly in striated muscles. Human XIRP2 contains 28 Xin repeats with properties identical to those of Xin. We conclude that the Xin repeats define a novel, repetitive actin-binding motif present in at least two different muscle proteins. These Xin-repeat proteins therefore constitute the first two members of a novel family of actin-binding proteins.

UNC-87, a calponin-related protein in C. elegans, antagonizes ADF/cofilin-mediated actin filament dynamics

“…Although, at present, no calponin-like protein has been identified in vertebrate striated muscle, several actin-stabilizing proteins including nebulin (McElhinny et al, 2005), Xin (Pacholsky et al, 2004) and myotilin (Salmikangas et al, 2003) also have repetitive actin-binding motifs. By contrast, C. elegans does not have nebulin or Xin, but Ce-kettin has immunoglobulin-like repeats and shows some similarity to myotilin.…”

N-WASP deficiency impairs EGF internalization and actin assembly at clathrin-coated pits

“…The expression construct for EGFPtagged human β-actin was purchased from BD Biosciences; mRFPactin has been described previously (Pacholsky et al, 2004). Human ARPC5A cDNA was fused into pEGFP-C1 (BD Biosciences) using BglII and SalI.…”

SH4-domain-induced plasma membrane dynamization promotes bleb-associated cell motility

“…The expression plasmids for N18-Yes-wt-GFP and N18-Yes-⌬Myr-GFP as well as the full-length HASPB, HASPB-GFP and N18-HASPBmCherry were generated by subcloning the respective coding sequences into pREV-TRE2. Expression plasmids for mCherry (Shaner et al, 2004) and mRFP-actin (Pacholsky et al, 2004) were kindly provided by Roger Tsien (UCSD, San Diego, CA) and Ulrike Engel (Nikon Imaging Center, Germany), respectively. The expression plasmid for the GFP fusion of Lck was generated by subcloning the Lck ORF into pEGFP-N1.…”