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Transformation by Rous sarcoma virus induces clathrin heavy chain phosphorylation
J Martin-Perez1, D Bar-Zvi, D Branton
1Department of Cellular and Developmental Biology, Harvard University, Cambridge, Massachusetts 02138.
The Journal of Cell Biology
|August 1, 1989
Summary
Chicken embryo fibroblast cells transformed by Rous sarcoma virus show phosphorylated clathrin heavy chain, unlike normal cells. This phosphorylation, potentially by pp60v-src, may alter cellular processes like endocytosis.
Area of Science:
- Cell Biology
- Virology
- Biochemistry
Background:
- Clathrin heavy chain phosphorylation is observed in cells transformed by Rous sarcoma virus.
- This phosphorylation differs significantly from that in normal cells.
Purpose of the Study:
- To investigate the phosphorylation of clathrin heavy chain in Rous sarcoma virus-transformed cells.
- To determine the role of pp60v-src in clathrin phosphorylation.
- To explore the consequences of clathrin phosphorylation on cellular morphology and function.
Main Methods:
- Phosphorylation assays on clathrin heavy chain.
- In vitro kinase assays using pp60v-src.
- Proteolytic digestion (V8) and peptide analysis (Cleveland analysis).
- Immunofluorescent staining for clathrin localization.
Main Results:
- Clathrin heavy chain is phosphorylated in transformed cells, with phosphate on serine and tyrosine residues.
- pp60v-src phosphorylates clathrin heavy chain in vitro, and peptide analysis suggests it's responsible in vivo.
- Phosphorylation occurs in both assembled and unassembled clathrin pools.
- Immunofluorescence reveals altered clathrin distribution in transformed cells (loss of perinuclear staining).
Conclusions:
- pp60v-src likely directly phosphorylates clathrin heavy chain in Rous sarcoma virus-transformed cells.
- Clathrin heavy chain phosphorylation may contribute to transformation-dependent changes in receptor-mediated endocytosis.