Related Experiment Videos
Cellular mechanisms of TGF-beta action
K D Rodland1, L L Muldoon, B E Magun
1Department of Cell Biology and Anatomy, Oregon Health Sciences University, Portland 97201-3098.
Abstract:
Transforming growth factor beta (TGF-beta), initially identified in platelet extracts by virtue of its ability to confer anchorage-independent growth and a neoplastic phenotype on mesenchymal cells, has subsequently been identified as a potent inhibitor of proliferation in most cells of epithelial origin. Our laboratory has investigated the role of specific second messengers in mediating the transcriptional responses of fibroblasts following addition of TGF-beta 1. Our studies indicate that TGF-beta 1, alone and in conjunction with epidermal growth factor (EGF), is capable of stimulating increases in both phosphoinositide metabolism and calcium influx, leading to significant increases in intracellular levels of Ca++ and inositol trisphosphate (IP3). Our data indicated that Ca++ influx and inositol phosphate release are coupled in Rat-1 cells, and suggested that influx of Ca++ from the extracellular medium is required for the change in IP3 accumulation observed in response to both EGF and TGF-beta 1. Using nuclear run-on analysis of the transcription of rat transin, a secreted metalloproteinase homologous to human stromelysin, we have also demonstrated a significant inhibition of transin transcription within 10 min of TGF-beta 1 treatment. The ability of TGF-beta 1 to inhibit transin gene transcription was not related to the TGF-beta 1-induced influx of Ca++ or to an increase in intracellular inositol phosphates, since inhibiting production of these second messengers failed to inhibit repression of the transin gene.
Insights
Transforming growth factor beta 1 (TGF-beta 1) inhibits transin gene transcription in fibroblasts. This inhibition is independent of TGF-beta 1-induced calcium influx or inositol phosphate production.
Area of Science:
- Cellular Biology
- Molecular Biology
- Biochemistry
Background:
- Transforming growth factor beta (TGF-beta) has diverse cellular effects, inhibiting epithelial cell proliferation while promoting mesenchymal cell growth.
- TGF-beta 1's role in mediating transcriptional responses in fibroblasts via second messengers is under investigation.
- Epidermal growth factor (EGF) is known to interact with TGF-beta 1 in cellular signaling pathways.
Purpose of the Study:
- To investigate the role of specific second messengers in TGF-beta 1-mediated transcriptional responses in fibroblasts.
- To determine the relationship between TGF-beta 1-induced phosphoinositide metabolism, calcium influx, and transin gene transcription.
- To elucidate the signaling mechanisms by which TGF-beta 1 regulates transin gene expression.
Main Methods:
- Measurement of phosphoinositide metabolism and intracellular calcium levels (Ca++) following TGF-beta 1 and EGF stimulation.
- Analysis of inositol trisphosphate (IP3) accumulation in response to growth factor treatment.
- Nuclear run-on assays to assess the rate of transin gene transcription.
Main Results:
- TGF-beta 1, alone and with EGF, stimulates phosphoinositide metabolism and Ca++ influx, increasing intracellular Ca++ and IP3 levels.
- Ca++ influx is required for the observed increase in IP3 accumulation in response to EGF and TGF-beta 1 in Rat-1 cells.
- TGF-beta 1 significantly inhibits transin gene transcription within 10 minutes, independently of Ca++ influx or IP3 increases.
Conclusions:
- TGF-beta 1 regulates transin gene transcription through a mechanism distinct from its effects on Ca++ influx and IP3 production.
- The inhibition of transin gene transcription by TGF-beta 1 is not mediated by the observed increases in intracellular Ca++ or IP3.
- These findings highlight a novel signaling pathway for TGF-beta 1 in regulating metalloproteinase gene expression in fibroblasts.