8099

CDK2AP1

DOC1|DORC1|ST19|doc-1|p12DOC-1;cyclin-dependent kinase 2 associated protein 1;CDK2AP1;cyclin-dependent kinase 2 associated protein 1

CDK2-associated protein 1|Deleted in oral cancer-1|cyclin-dependent kinase 2-associated protein 1|deleted in oral cancer 1|putative oral cancer suppressor

12q24.31

protein-coding

We evaluated the effect of expressing the cell cycle regulator protein cdk2-associating protein1 (cdk2ap1) in inhibiting growth of squamous cell carcinoma (SCC). expression of cdk2ap1 correlated with reduction in several SCC malignant cell phenotypes, including reduced angiogenesis. We observed several alterations in gene expression consistent with classical functions of cdk2ap1, including upregulation of cell cycle inhibitory genes, and an upregulation in expression of genes belonging to both intrinsic and extrinsic apoptotic cascades. Interestingly, we also uncovered a profile of gene expression and activation of signaling pathways that may suggest new tumor-suppressive functions for cdk2ap1 through downregulation of invasion/metastasis and modulation of antiangiogenesis by upregulation of the TGFbeta signaling pathway. Blocking of the TGFbeta1 pathway resulted in inhibition of the cdk2ap1 antiangiogenesis phenotype. In combination, these data support the role of cdk2ap1 as a tumor suppressor gene that can regulate SCC tumor growth in a cell autonomous manner through decreases in invasiveness and a non-cell autonomous manner through decreases in angiogenesis phenotypes, and these are novel phenotypes induced by cdk2ap1.

The present study was undertaken to investigate the regulation of P12(CDK2AP1) by miRNAs. A conserved target site for miR-21 within the CDK2AP1-3-UTR at nt 349-370 was predicted by bioinformatics software and an inverse correlation of miR-21 and CDK2AP1 protein was observed. Highly specific amplification and quantification of miR-21 was achieved using real-time RT-PCR. Transfection of HaCaT cells with pre-miR-21 significantly suppressed a luciferase reporter including the CDK2AP1-3-UTR, whereas transfection of Tca8113 with anti-miR-21 increased activity of this reporter. This was abolished when a construct mutated at the miR-21/nt 349-370 target site was used instead. Anti-miR-21-transfected Tca8113 cells showed an increase of CDK2AP1 protein and reduced proliferation and invasion. Resected primary tumors and tumor-free surgical margins of 18 patients with head and neck squamous cell carcinomas demonstrated an inverse correlation between miR-21 and P12(CDK2AP1). This study shows that P12(CDK2AP1) is downregulated by miR-21 and that miR-21 promotes proliferation and invasion in cultured cells.

We have isolated a human cDNA encoding a 115-amino-acid polypeptide that revealed 97% identity to a candidate tumor suppressor gene for oral cancer in Mesocricetus auratus (deleted in oral cancer-1; doc-1). It also showed a high degree of homology to a gene induced by TNF-alpha in Mus musculus. To investigate its possible role in esophageal carcinogenesis, we examined genetic alterations and expression levels of the gene in 13 esophageal carcinoma cell lines and 10 primary esophageal carcinomas. No mutation nor reduction of expression was observed in any of the 23 cancer materials examined. These results imply that the human doc-1 homologue is unlikely to play a significant role in esophageal carcinogenesis, although its role in the TNF-alpha signaling pathway remains unclear. We mapped DOC1 to chromosome band 12q24.31 by fluorescence in situ hybridization.

doc-1 is a putative tumor suppressor gene isolated and identified from the hamster oral cancer model. Here, we report the molecular cloning and the functional characterization of the human ortholog of the hamster doc-1 gene. Human doc-1 cDNA is 1.6 kilobase pairs in length and encodes for a 115-amino acid polypeptide (12.4 kDa, pI 9. 53). Sequence analysis showed 98% identity between human and hamster doc-1 protein sequences. DOC-1 is expressed in all normal human tissues examined. In oral keratinocytes, expression of DOC-1 is restricted to normal oral keratinocytes. By immunostaining of normal human mucosa, DOC-1 is detected in both the cytoplasm and nuclei of basal oral keratinocytes; while in suprabasilar cells, it is primarily found in the nuclei. Human oral cancers in vivo did not exhibit immunostaining for DOC-1. Like murine DOC-1, human DOC-1 associates with DNA polymerase alpha/primase and mediates the phosphorylation of the large p180 catalytic subunit, suggesting it may be a potential regulator of DNA replication in the S phase of the cell cycle. Using a human doc-1 cosmid as a probe, human doc-1 is mapped to chromosome 12q24. We identified four exons in the entire human doc-1 gene and determined the intron-exon boundaries. By polymerase chain reaction and direct sequencing, we examined premalignant oral lesion and oral cancer cell lines and found no intragenic mutations.