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Protein expression and functional analysis of the FHIT gene in human tumor cells
G A Otterson1, G H Xiao, J Geradts
1Medicine Branch, Division of Clinical Sciences, National Cancer Institute, Bethesda, MD, USA.
Background:
The fragile histidine triad (FHIT) gene at chromosome 3p14.2 has been proposed to be a candidate tumor suppressor gene in human cancers. To test whether FHIT exhibits the functional properties of a tumor suppressor gene, we studied the expression of its protein (pFHIT) in human carcinoma cells and examined the ability of FHIT to inhibit the neoplastic phenotype of cancer cells.
Methods:
Subcellular localization and patterns of protein expression in tumor cells were determined by immunohistochemical analysis and immunoblotting with the use of polyclonal anti-pFHIT antisera. In tumor cells with undetectable pFHIT, we examined the effect of recombinant pFHIT expression on morphology, growth rate, colony formation, and in vivo tumor formation.
Results:
We demonstrated that pFHIT is a cytoplasmic 17-kd polypeptide whose expression could not be detected in 30 of 52 human carcinoma cell lines tested. We observed, however, that the stable overexpression of pFHIT did not alter cell morphology, inhibit colony formation, or inhibit cell proliferation in vitro. Furthermore, overexpression of pFHIT did not lead to altered cell cycle kinetics in dividing cells. The in vivo tumorigenicity of a tumor cell line that expressed high levels of recombinant pFHIT was equivalent to that of control transfectants and of parental cells.
Conclusions:
These results suggest that the replacement of pFHIT in human carcinoma cells does not suppress tumor cell growth and that this protein may be involved in tumorigenesis in ways that are distinct from the "classic" tumor suppressor paradigm.
Insights
The fragile histidine triad (FHIT) protein, proposed as a tumor suppressor, did not inhibit cancer cell growth or tumor formation when overexpressed. These findings suggest FHIT
Area of Science:
- Oncology
- Molecular Biology
- Cancer Genetics
Background:
- The fragile histidine triad (FHIT) gene at 3p14.2 is investigated as a potential tumor suppressor in human cancers.
- FHIT's role in tumorigenesis is explored through its protein product, pFHIT.
Purpose of the Study:
- To determine the functional properties of FHIT as a tumor suppressor gene.
- To investigate the expression of pFHIT in human carcinoma cells.
- To assess the impact of FHIT reintroduction on cancer cell phenotypes.
Main Methods:
- Immunohistochemistry and immunoblotting were used to analyze pFHIT expression and localization.
- Recombinant pFHIT was expressed in tumor cells lacking endogenous pFHIT.
- Effects on cell morphology, growth, colony formation, and in vivo tumorigenicity were evaluated.
Main Results:
- pFHIT is a 17-kd cytoplasmic polypeptide.
- pFHIT expression was undetectable in 30 of 52 tested human carcinoma cell lines.
- Overexpression of pFHIT did not alter cell morphology, inhibit proliferation, or affect in vivo tumor formation.
Conclusions:
- Reintroducing pFHIT into human carcinoma cells did not suppress tumor cell growth.
- FHIT may be involved in tumorigenesis through mechanisms distinct from classical tumor suppressor pathways.