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Coamplification of the CDK4 gene with MDM2 and GLI in human sarcomas
Z A Khatib1, H Matsushime, M Valentine
1Department of Experimental Oncology and Hematology/Oncology, St. Jude Children's Research Hospital, Memphis, Tennessee 38105.
Abstract:
The 34-kilodalton cyclin-dependent kinase, p34cdk4, is a major catalytic subunit of mammalian D-type cyclins, which act during the G1 phase of the cell cycle to enforce the decision of cells to enter S phase. A murine complementary DNA clone was used to clone the cognate human CDK4 gene, which was localized to human chromosome 12, band q13, by fluorescence in situ hybridization. Because this chromosomal band contains the GLI and MDM2 genes, which are frequently amplified in human sarcomas, we analyzed CDK4 copy number and expression in a panel of sarcoma cell lines. An osteosarcoma cell line, OsACL, manifested a 25-fold increased copy number of CDK4, amplified concordantly with both GLI and MDM2, whereas a rhabdomyosarcoma cell line, SJRH30, was found to have an amplicon that included CDK4 and GLI but not MDM2. CDK4 mRNA and protein were overexpressed in both cell lines, and nucleotide sequencing analysis indicated that the gene had not sustained mutations. These observations provide the first evidence for amplification of a gene encoding a cell division cycle protein kinase, complement recent data indicating that genes encoding D-type cyclins are targets of chromosomal rearrangement and gene amplification in tumor cells, and suggest that CDK4 amplification might contribute to oncogenesis.
Insights
Gene amplification of CDK4, a cell cycle protein kinase, was observed in sarcoma cell lines. This suggests CDK4 gene amplification may play a role in cancer development.
Area of Science:
- Molecular Biology
- Cancer Genetics
- Cell Cycle Regulation
Background:
- Cyclin-dependent kinase 4 (CDK4) is a key regulator of the G1 phase in the cell cycle.
- CDK4 is a catalytic subunit of D-type cyclins, crucial for cell cycle progression.
- Genes located on chromosome 12, band q13, including GLI and MDM2, are frequently amplified in sarcomas.
Purpose of the Study:
- To clone the human CDK4 gene and determine its chromosomal localization.
- To investigate the copy number and expression of CDK4 in sarcoma cell lines.
- To explore the potential role of CDK4 amplification in oncogenesis.
Main Methods:
- Human CDK4 gene cloning using a murine cDNA.
- Fluorescence in situ hybridization (FISH) for gene localization.
- Analysis of CDK4 copy number and mRNA/protein expression in sarcoma cell lines.
- Nucleotide sequencing to detect gene mutations.
Main Results:
- CDK4 gene localized to human chromosome 12, band q13.
- Significant CDK4 gene amplification (25-fold) observed in an osteosarcoma cell line (OsACL), concordant with GLI and MDM2.
- CDK4 amplification and overexpression found in both osteosarcoma and rhabdomyosarcoma cell lines.
- No mutations detected in the CDK4 gene in the analyzed cell lines.
Conclusions:
- First evidence of amplification for a cell division cycle protein kinase gene (CDK4).
- Supports findings of D-type cyclin gene alterations in tumor cells.
- Suggests that CDK4 gene amplification may contribute to the development of sarcomas.