Identification and characterization of the novel centrosome-associated protein CCCAP.
We have cloned and characterized the cDNA, expression pattern, and subcellular localization of the human and murine orthologs of the centrosomal colon cancer autoantigen protein (CCCAP). We identified both the transcriptional start site of murine CCCAP (mCCCAP) and its TATA-less promoter within BAC genomic clones of the mCCCAP 5' region. The mCCCAP transcript is ubiquitously present in mouse tissues, but at very low copy number. The 2151 bp open reading frame of mCCCAP encodes an 83 kDa protein that possesses a large C-terminal coiled-coil domain, which is able to homo-oligomerize in the yeast 2-hybrid system. Endogenous mCCCAP localizes to the centrosomes of murine BALB/c 3T3 fibroblasts during both interphase and mitosis. This centrosomal localization was not disrupted by nocodazole-induced depolymerization of the microtubule cytoskeleton, suggesting that mCCCAP is an integral component of the centrosome rather than simply a microtubule-associated protein. We also cloned human CCCAP (hCCCAP). The 2139 bp open reading frame of hCCCAP encodes an 82.5 kDa protein that is 71% identical to mCCCAP at the amino acid level and has the same predicted secondary structure. Ectopically expressed full-length hCCCAP in human U2-osteosarcoma cells also displayed centrosomal localization during interphase and mitosis. This pattern of localization was abolished by truncations of the N- and C-terminus of the protein. We further discovered that the C-terminal portion of hCCCAP is identical to the human colon cancer autoantigen NY-CO-8 (Human Gene Nomenclature symbol SDCCAG8).
Kenedy
Andrew A
AA
Division of Hematology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
Cohen
Kenneth J
KJ
Loveys
Deborah A
DA
Kato
Gregory J
GJ
Dang
Chi V
CV
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001324:Autoantigens / Q000235:genetics*; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D015152:Blotting, Northern; D018385:Centrosome / Q000378:metabolism*; D003001:Cloning, Molecular; D018387:Codon, Initiator / Q000235:genetics; D003599:Cytoskeleton / Q000378:metabolism; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008870:Microtubules / Q000378:metabolism; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D017422:Sequence Analysis, DNA; D024363:Transcription Initiation Site; D014407:Tumor Cells, Cultured
D001324:Autoantigens; D018387:Codon, Initiator; D018076:DNA, Complementary; D009363:Neoplasm Proteins; D012333:RNA, Messenger; C472380:SDCCAG8 protein, human; C472381:Sdccag8 protein, mouse