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Epidermal growth factor signaling and mitogenesis in Plcg1 null mouse embryonic fibroblasts
1Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-0146, USA.
Abstract:
Gene targeting techniques and early mouse embryos have been used to produce immortalized fibroblasts genetically deficient in phospholipase C (PLC)-gamma1, a ubiquitous tyrosine kinase substrate. Plcg1(-/-) embryos die at embryonic day 9; however, cells derived from these embryos proliferate as well as cells from Plcg1(+/+) embryos. The null cells do grow to a higher saturation density in serum-containing media, as their capacity to spread out is decreased compared with that of wild-type cells. In terms of epidermal growth factor receptor activation and internalization, or growth factor induction of mitogen-activated protein kinase, c-fos, or DNA synthesis in quiescent cells, PLcg1(-/-) cells respond equivalently to PLcg1(+/+) cells. Also, null cells are able to migrate effectively in a wounded monolayer. Therefore, immortalized fibroblasts do not require PLC-gamma1 for many responses to growth factors.
Insights
Researchers created immortalized fibroblasts lacking phospholipase C (PLC)-gamma1. These cells, despite embryo lethality, proliferate normally and respond to growth factors, indicating PLC-gamma1 is not essential for many fibroblast growth factor responses.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Phospholipase C (PLC)-gamma1 is a key substrate in tyrosine kinase signaling pathways.
- PLC-gamma1 plays a role in cellular responses to growth factors, but its necessity in fibroblasts is not fully understood.
Purpose of the Study:
- To investigate the role of PLC-gamma1 in immortalized fibroblast function.
- To determine if PLC-gamma1 is required for various growth factor-mediated responses in fibroblasts.
Main Methods:
- Gene targeting in early mouse embryos to create Plcg1(-/-) fibroblasts.
- Culturing and characterizing Plcg1(-/-) and wild-type (Plcg1(+/+)) fibroblasts.
- Assessing cell proliferation, saturation density, epidermal growth factor receptor (EGFR) signaling, and cell migration.
Main Results:
- Plcg1(-/-) embryos are embryonic lethal at day 9, but derived fibroblasts proliferate similarly to wild-type cells.
- Null cells exhibit increased saturation density and reduced spreading capacity.
- PLC-gamma1 deficient cells show equivalent responses in EGFR activation, internalization, MAPK and c-fos induction, and DNA synthesis.
- Null cells maintain effective migration in wounded monolayers.
Conclusions:
- Immortalized fibroblasts do not require PLC-gamma1 for many critical responses to growth factors.
- PLC-gamma1's role in fibroblast proliferation and migration appears dispensable, though it influences cell spreading and saturation density.