Related Experiment Videos
DOC-2, a candidate tumor suppressor gene in human epithelial ovarian cancer
1Department of Obstetrics, Gynecology and Reproductive Biology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
Abstract:
Using RNA fingerprinting (RAP) strategy and Northern blot analysis, we identified a differentially expressed sequence DOC-2 which is detectable in all normal human ovarian surface epithelial (HOSE) cell cultures but not in ovarian cancer cell lines and tissues. Subsequent cloning of DOC-2 from a cDNA library generated from the HOSE cells was carried out using the 3' and 5' RACE approach. A 3268 base pair full length cDNA of DOC-2 was isolated and sequenced. The predicted protein has a length of 770 amino acids. Homology search of all NCBI sequences indicated that the amino acid sequence of DOC-2 shares 93% homology with the mouse p96/mDab2 phosphoprotein and has a phosphotyrosine interacting domain (PID) and multiple SH3 binding motifs. Chromosomal localization by FISH showed that the DOC-2 gene is located on 5p13. Western blot analysis showed that the 105 kDa DOC-2 protein was down-regulated in all the carcinoma cell lines. In-situ immunohistochemistry performed on normal ovaries, and benign, borderline and invasive ovarian tumor tissues showed down regulation of DOC-2 protein particularly in serous ovarian tumor tissues. When DOC-2 was transfected into the ovarian carcinoma cell line SKOV3, the stable transfectants showed significantly reduced growth rate and ability to form tumors in nude mice. These data suggest that down-regulation of DOC-2 may play an important role in ovarian carcinogenesis.
Insights
Down-regulation of the DOC-2 gene, identified in normal ovarian cells but absent in cancer, correlates with ovarian cancer development. Restoring DOC-2 expression in cancer cells reduced tumor growth.
Area of Science:
- Molecular Biology
- Genetics
- Oncology
Background:
- The DOC-2 gene is expressed in normal human ovarian surface epithelial (HOSE) cells.
- DOC-2 is notably absent in ovarian cancer cell lines and tissues.
Purpose of the Study:
- To clone and characterize the DOC-2 gene.
- To investigate the role of DOC-2 in ovarian carcinogenesis.
Main Methods:
- RNA fingerprinting (RAP), Northern blot, 3' and 5' RACE, cDNA sequencing.
- Western blot, in-situ immunohistochemistry, FISH chromosomal localization.
- Transfection of DOC-2 into ovarian cancer cells (SKOV3) and tumor formation assays in nude mice.
Main Results:
- A full-length 3268 bp cDNA of DOC-2 was isolated, encoding a 770 amino acid protein with homology to mouse p96/mDab2.
- DOC-2 protein (105 kDa) was down-regulated in all tested ovarian carcinoma cell lines and particularly in serous ovarian tumors.
- Overexpression of DOC-2 in SKOV3 cells significantly reduced proliferation and tumor formation in vivo.
Conclusions:
- Down-regulation of DOC-2 is implicated in ovarian carcinogenesis.
- DOC-2 may function as a tumor suppressor in ovarian cancer.
- Further research into DOC-2's mechanism is warranted for therapeutic strategies.