Identification of a novel cardiac lineage-associated Protein(cCLP-1): A candidate regulator of cardiogenesis

We describe the isolation and characterization of a cDNA clone, called cCLP-1, that is a candidate for the previously described early cardiac-specific transcription factor BBF-1. BBF-1 binds the MEF2 (or element B) binding site within the cardiac myosin light chain 2 (MLC2) gene promoter. We used the element B sequence as a probe to screen an expression library constructed from mRNA obtained from the presumptive heart-forming regions of stage 6 chicken embryos. This yielded the cCLP-1 cDNA clone. Gel-shift analysis of stage 6 embryonic chicken protein extracts suggests that a protein that is recognized by the anti-cCLP-1 antibody binds to the same element B binding site to which BBF-1 binds, cCLP-1 mRNA was detected early in chicken development, prior to cardiac fate assignment at stage 4. The expression pattern of cCLP-1, based on whole mount in situ hybridization, coincides remarkably well with the established morphogenetic field of early heart formation. The nuclear localization of cCLP-1 is phosphorylation-dependent, suggesting that cCLP-1 may be a member of that class of transcription factors whose activity is regulated by cytoplasm to nucleus transport. Taken together, these data suggest that cCLP-1 may encode a novel transcription factor whose expression pattern is in agreement with that of the cardiogenic precursor cells of the early chicken embryo. (C) 1999 Academic Press.