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 phosphoprotein NUCKS, previously designated P1. Molecular analyses of several overlapping and full-length cDNAs from HeLa cells and rat brain revealed a protein with an apparent molecular mass of 27 kDa in both species. The deduced amino-acid sequences are highly conserved between human and rodents, but show no homology with primary structures in protein databases or with translated sequences of cDNAs in cDNA databanks. Although the protein has some features in common with the high mobility group proteins HMGI/Y, attempts to find a putative protein family by database query using both sequence alignment methods and amino-acid composition have failed. Northern blot analyses revealed that human and rat tissues contain three NUCKS transcripts varying 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 dichroism analysis and secondary structure predictions based on the amino-acid sequence indicate a low level of alpha 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 residues. Phosphopeptide mapping and back-phosphorylation experiments employing NUCKS from HeLa interphase and metaphase cells show that the protein is phosphorylated by Cdk1 during mitosis of the cell cycle.
Ostvold
A C
AC
Department of Medical Biochemistry, University of Oslo, Norway. annecari@basalmed.uio.no
Norum
J H
JH
Mathiesen
S
S
Wanvik
B
B
Sefland
I
I
Grundt
K
K
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism; D016203:CDC2 Protein Kinase / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism; D002453:Cell Cycle; D002467:Cell Nucleus / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D016208:Databases, Factual; D015723:Gene Library; D024243:HMGB1 Protein; D006367:HeLa Cells; D006609:High Mobility Group Proteins / Q000378:metabolism; D006801:Humans; D051379:Mice; D008938:Mitosis; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism*; D010449:Peptide Mapping; D010750:Phosphoproteins / Q000235:genetics* / Q000378:metabolism*; D010766:Phosphorylation; D011233:Precipitin Tests; D017433:Protein Structure, Secondary; D051381:Rats; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014018:Tissue Distribution
D002352:Carrier Proteins; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D024243:HMGB1 Protein; D006609:High Mobility Group Proteins; C426654:NUCKS1 protein, human; D009687:Nuclear Proteins; D010750:Phosphoproteins; D016203:CDC2 Protein Kinase