Characterization of cDNA clones encoding a human fibroblast caldesmon isoform and analysis of caldesmon expression in normal and transformed cells.
Overlapping cDNA clones encoding a low M gamma human nonmuscle caldesmon isoform (HUM 1-CaD) span the entire coding region (538 amino acids) as well as 111 base pairs (bp) of 5'-noncoding and 1249 bp of 3'-noncoding region. Northern blot probes derived from either the coding or 3'-noncoding region hybridized to a 4.3-kilobase mRNA in nonmuscle cells and a 5.2-kilobase mRNA in stomach tissue. Primer extension results indicated that the 5'-noncoding region of the HUM 1-CaD mRNA is approximately 700 bp in length and also suggested that 1-CaD mRNAs with common 5'-noncoding regions are expressed in both liver and fibroblast cells. Comparisons of the human, rat, and chicken 1-CaD amino acids sequences demonstrated that although each isoform has unique characteristics, extensive regions of conservation exist. Amino acids 27-53 and 97-127 are 100% identical in these isoforms while amino acids 297-531 of HUM 1-CaD are 94 and 85% identical to the rat and chicken 1-CaDs, respectively. In addition, the levels of HUM 1-CaD mRNA and protein appeared to be decreased by 2-4 fold in the transformed derivatives of KD and WI38 cell lines as judged by Northern and Western blot analysis. The results suggest that the decrease of 1-CaD protein in these transformed cells is a direct result of decreased 1-CaD mRNA synthesis and/or increased mRNA turnover.
Novy
R E
RE
Department of Biology, University of Iowa, Iowa City 52242.
Lin
J L
JL
Lin
J J
JJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D002148:Calmodulin-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D002461:Cell Line, Transformed; D003001:Cloning, Molecular; D016384:Consensus Sequence; D004247:DNA; D005347:Fibroblasts / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009130:Muscle, Smooth / Q000378:metabolism; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D014407:Tumor Cells, Cultured
D002148:Calmodulin-Binding Proteins; D004247:DNA