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Published on: March 9, 2012
Decreased retinoylation in NIH 3T3 cells transformed with activated Ha-ras
N Takahashi1, L M De Luca, T R Breitman
1Department of Health Chemistry, Hoshi University, Tokyo, Japan.
Abstract:
Retinoylation (retinoic acid acylation) is a post-translation modification of proteins occurring in a variety of mammalian cell lines and in vivo. To gain further knowledge of the role of retinoylation we studied it in NIH 3T3 cells and NIH 3T3 cells transformed by an activated Ha-ras oncogene (NIH Ha-ras-3T3 cells). In serum-free medium retinoic acid (RA) inhibited growth of NIH 3T3 cells but did not inhibit growth of NIH Ha-ras-3T3 cells. After incubation with [3H]RA, the level of retinoylated protein in NIH 3T3 cells was about 1.5-fold greater than in NIH Ha-ras-3T3 cells. On one-dimensional polyacrylamide gel electrophoresis, both the rate and the extent of retinoylation were greater in NIH 3T3 cells. We detected about 40 retinoylated proteins in NIH 3T3 cells by two-dimensional polyacrylamide gel electrophoresis. Only about 15 proteins were retinoylated, but at reduced levels, in NIH Ha-ras-3T3 cells. These results suggest that the activated ras oncogene inhibits retinoylation. This inhibition may in turn be related to the loss of other RA responses of NIH 3T3 cells, including growth inhibition, retinoic acid catabolism, down-regulation of fibronectin biosynthesis, and induction of tissue-type transglutaminase, which are not seen to the same extent in NIH Ha-ras-3T3 cells.
Insights
The activated ras oncogene inhibits retinoylation, a protein modification by retinoic acid (RA). This inhibition may explain why ras-transformed cells lose sensitivity to RA's growth-inhibiting effects.
Area of Science:
- Biochemistry
- Cell Biology
- Molecular Biology
Background:
- Retinoylation, the acylation of proteins by retinoic acid (RA), is a significant post-translational modification observed in mammalian cells.
- Understanding retinoylation's role is crucial for deciphering cellular responses to RA.
Purpose of the Study:
- To investigate the impact of an activated Ha-ras oncogene on retinoylation processes.
- To compare retinoylation levels and patterns in normal NIH 3T3 cells versus NIH Ha-ras-3T3 transformed cells.
Main Methods:
- Utilized NIH 3T3 and NIH Ha-ras-3T3 cells for comparative analysis.
- Administered [3H]RA to cells and quantified retinoylated protein levels.
- Employed one-dimensional and two-dimensional polyacrylamide gel electrophoresis to analyze retinoylation extent and identify modified proteins.
Main Results:
- NIH 3T3 cells exhibited significantly higher rates and extents of retinoylation compared to NIH Ha-ras-3T3 cells.
- Approximately 40 retinoylated proteins were identified in NIH 3T3 cells, versus only 15 in NIH Ha-ras-3T3 cells, with reduced modification levels.
- Activated ras oncogene was found to inhibit retinoylation.
Conclusions:
- The activated ras oncogene suppresses retinoylation in NIH 3T3 cells.
- Ras-mediated inhibition of retinoylation may underlie the diminished cellular responses to RA, including growth inhibition and altered biosynthesis pathways.
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