BIOCHEMICAL AND MOLECULAR CHARACTERIZATION OF THE CHICKEN CYSTEINE-RICH PROTEIN, A DEVELOPMENTALLY-REGULATED LIM-DOMAIN PROTEIN THAT IS ASSOCIATED WITH THE ACTIN CYTOSKELETON

LIM domains are present in a number of proteins including transcriptio n factors, a protooncogene product, and the adhesion plaque protein zy xin. The LIM domain exhibits a characteristic arrangement of cysteine and histidine residues and represents a novel zinc binding sequence (M ichelsen et al., 1993). Previously, we reported the identification of a 23-kD protein that interacts with zyxin in vitro (Sadler et al., 199 2). In this report, we describe the purification and characterization of this 23-kD zyxin-binding protein from avian smooth muscle. Isolatio n of a cDNA encoding the 23-kD protein has revealed that it consists o f 192 amino acids and exhibits two copies of the LIM motif. The 23-kD protein is 91% identical to the human cysteine-rich protein (hCRP); th erefore we refer to it as the chicken cysteine-rich protein (cCRP). Ex amination of a number of chick embryonic tissues by Western immunoblot analysis reveals that cCRP exhibits tissue-specific expression. cCRP is most prominent in tissues that are enriched in smooth muscle cells, such as gizzard, stomach, and intestine. In primary cell cultures der ived from embryonic gizzard, differentiated smooth muscle cells exhibi t the most striking staining with anti-cCRP antibodies. We have perfor med quantitative Western immunoblot analysis of cCRP, zyxin, and alpha -actinin levels during embryogenesis. By this approach, we have demons trated that the expression of cCRP is developmentally regulated.