Molecular cloning of a mammalian nuclear phosphoprotein NUCKS, which serves as a substrate for Cdk1 in vivo

We have isolated and characterized a cDNA encoding a mammalian nuclear phos phoprotein NUCKS, previously designated P1. Molecular analyses of several o verlapping and full-length cDNAs from HeLa cells and rat brain revealed a p rotein with an apparent molecular mass of 27 kDa in both species. The deduc ed amino-acid sequences are highly conserved between human and rodents, but show no homology with primary structures in protein databases or with tran slated sequences of cDNAs in cDNA databanks. Although the protein has some features in common with the high mobility group proteins HMGI/Y, attempts t o find a putative protein family by database query using both sequence alig nment methods and amino-acid composition have failed. Northern blot analyse s revealed that human and rat tissues contain three NUCKS transcripts varyi ng in size from 1.5 to 6.5 kb. All human and rat tissues express the gene, but the level of transcripts varies among different tissues. Circular dichr oism analysis and secondary structure predictions based on the amino-acid s equence indicate a low level of or helical content and substantial amounts of beta turn structures. The protein is phosphorylated in all phases of the cell cycle and exhibits mitosis-specific phosphorylation of threonine resi dues. Phosphopeptide mapping and back-phosphorylation experiments employing NUCKS from HeLa interphase and metaphase cells show that the protein is ph osphorylated by Cdk1 during mitosis of the cell cycle.