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—To determine how myocardial terminal differentiation is regulated by cell cycle control genes, we studied cdc2 expression in rat cardiac muscle and found that cdc2 mRNA and protein levels were reduced in neonatal compared with fetal ventricles and became undetectable in juvenile and adult ventricles. To further determine whether cdc2 downregulation is attributed to a decrease in transcription, transient expression assay was performed using the progressively truncated 6.2-, 1.8-, 1.1-, 0.7-, and 0.1-kb human cdc2 5′ flanking regions. All five fragments activated reporter expression in fetal myocytes and were significantly less active in neonatal myocytes. The 0.1-kb fragment showed 65% of the activity of the 6.2-kb fragment. A protein binding site that contains an inverted CCAAT box was identified within the 0.1-kb fragment by DNase I footprint assay and named the cdc2 promoter binding factor (CPBF) site. Point mutations within the CPBF site that abolish CPBF binding significantly decreased both 0.1- and 6.2-kb promoter activities. Competition and antibody supershift assays suggested that CPBF was identical or related to the transcription factor, nuclear factor Y (NF-Y). The 0.1-kb promoter activity was suppressed by a dominant-negative NF-Y mutant in fetal myocytes. Taken together, our results demonstrate that cardiac cdc2 expression is downregulated after birth and turned off when the juvenile stage is attained. A 0.1-kb promoter fragment of cdc2 contains major information for both cdc2 transcriptional activation and suppression in fetal and neonatal myocytes, respectively. NF-Y or its related factor plays a critical role in activating the 0.1-kb cdc2 promoter.

Transcriptional Activation of the p34 cdc2 Gene by cdc2 Promoter Binding Factor/Nuclear Factor-Y in Fetal Rat Ventricular Myocytes

Transcriptional Activation of the p34 ^cdc2 Gene by cdc2 Promoter Binding Factor/Nuclear Factor-Y in Fetal Rat Ventricular Myocytes