The ZASP-like Motif in Actinin-associated LIM Protein Is Required for Interaction with the α-Actinin Rod and for Targeting to the Muscle Z-line

Abstract

The Z-line is a specialized structure connecting adjacent sarcomeres in muscle cells. ␣-Actinin cross-links actin filaments in the Z-line. Several PDZ-LIM domain proteins localize to the Z-line and interact with ␣-actinin. Actinin-associated LIM protein (ALP), C-terminal LIM domain protein (CLP36), and Z band alternatively spliced PDZ-containing protein (ZASP) have a conserved region named the ZASP-like motif (ZM) between PDZ and LIM domains. To study the interactions and function of ALP we used purified recombinant proteins in surface plasmon resonance measurements. We show that ALP and ␣-actinin 2 have two interaction sites. The ZM motif was required for the interaction of ALP internal region with the ␣-actinin rod and for targeting of ALP to the Z-line. The PDZ domain of ALP bound to the C terminus of ␣-actinin. This is the first indication that the ZM motif would have a direct role in a proteinprotein interaction. These results suggest that the two interaction sites of ALP would stabilize certain conformations of ␣-actinin 2 that would strengthen the Z-line integrity.The muscle Z-line is a highly specialized structure between adjacent sarcomeres in muscle fibers that maintain the organization of the contractile machinery (for review, see Ref. 1). ␣-Actinin is one of the major components at the Z-line. The ␣-actinin polypeptide is composed of an N-terminal actin binding domain, four spectrin repeats that form the central rod region (2, 3), and two pairs of C-terminal EF-hands (4). ␣-Actinin forms an antiparallel homodimer. ␣-Actinin cross-links actin filaments from opposite sarcomeres to the Z-line and, therefore, has a major mechanical function in keeping the sarcomeres together.Several PDZ-LIM proteins have been detected in the muscle Z-line and shown to interact with ␣-actinin (5-12). PDZ domain is a widely expressed protein-protein interaction domain (for review, see Ref. 13). PDZ-LIM proteins form one subgroup of PDZ proteins (13) and are regarded as mediators between cytoskeletal structures and signaling cascades.PDZ-LIM proteins can be divided in two subclasses; they either contain one or three LIM domains. Actinin-associated LIM protein (ALP) 1 (5, 10), C-terminal LIM domain protein (CLP36) (14) (also called hCLIM1 (15) or Elfin (16)), Reversion-induced LIM protein (RIL) (17), and Mystique (Uniprot accession number Q7Z584) belong to the first class, which has one N-terminal PDZ domain and one C-terminal LIM domain. ALP is expressed in muscle (5, 10), whereas CLP36 and RIL are mainly expressed in nonmuscle tissues (11,18,19). CLP36 shows also high expression levels in muscle (7,15,16). Enigma, Enigma homology protein, and ZASP/Cypher/Oracle form the second class, with one N-terminal PDZ domain and three C-terminal LIM domains. They all are expressed mainly in muscle (6, 9, 20 -22). An interesting feature of these seven PDZ-LIM proteins is that ALP, CLP36, and ZASP contain a conserved region, named ZASP-like motif (ZM) (SMART prediction (23) accession number SM 00735, Interpro 006643), in the internal r...

A gene coexpression network for bovine skeletal muscle inferred from microarray data

“…These two proteins are both located in the z-disk and bind to each other (9), and they are located in the same cluster in the C2C12 differentiation data set (31). In addition, the expression of TCAP is correlated with the expression of PDLIM3, another component of the z-disk (11). The expression of both TCAP and PDLIM3 is also positively correlated with the expression of PFKM (a muscle cytoplasmic phosphofructokinase) and negatively correlated with the expression of SPP1 (osteopontin).…”

A Class III PDZ Binding Motif in the Myotilin and FATZ Families Binds Enigma Family Proteins: a Common Link for Z-Disc Myopathies

“…To date, there are six isoforms of ZASP/Cypher, all of which contain an N-terminal PDZ domain and none or three C-terminal LIM domains. ZASP/Cypher also contains a third domain known as the ZM motif which can also be found in ALP and CLP-36 (23,24). It interacts with two different regions of ␣-actinin-2; its PDZ domain binds to the C-terminal EH-hand region of ␣-actinin-2, whereas its ZM motif binds to the rod region of ␣-actinin-2 (2, 23).…”

Molecular mechanisms of mechanosensing in muscle development

“…Among several candidates, PDZ-LIM domain proteins comprised of ALP and Enigma subfamilies, which contain one or three LIM domains, respectively, have been identified as ␣-actinin binding partners (Xia et al, 1997;Pomies et al, 1999;Zhou et al, 1999;Pashmforoush et al, 2001;Klaavuniemi et al, 2004). Most members of this group colocalize with ␣-actinin in the Z-disks of striated and cardiac muscles and in intercalated disks of cardiac muscles.…”