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Patched1 interacts with cyclin B1 to regulate cell cycle progression
E A Barnes1, M Kong, V Ollendorff
1Department of Chemistry and Biochemistry, Center for Molecular Genetics, University of California-San Diego, La Jolla, CA 92093-0367, USA.
Abstract:
The initiation of mitosis requires the activation of M-phase promoting factor (MPF). MPF activation and its subcellular localization are dependent on the phosphorylation state of its components, cdc2 and cyclin B1. In a two-hybrid screen using a bait protein to mimic phosphorylated cyclin B1, we identified a novel interaction between cyclin B1 and patched1 (ptc1), a tumor suppressor associated with basal cell carcinoma (BCC). Ptc1 interacted specifically with constitutively phosphorylated cyclin B1 derivatives and was able to alter their normal subcellular localization. Furthermore, addition of the ptc1 ligand, sonic hedgehog (shh), disrupts this interaction and allows cyclin B1 to localize to the nucleus. Expression of ptc1 in 293T cells was inhibitory to cell proliferation; this inhibition could be relieved by coexpression of a cyclin B1 derivative that constitutively localizes to the nucleus and that could not interact with ptc1 due to phosphorylation-site mutations to ALA: In addition, we demonstrate that endogenous ptc1 and endogenous cyclin B1 interact in vivo. The findings reported here demonstrate that ptc1 participates in determining the subcellular localization of cyclin B1 and suggest a link between the tumor suppressor activity of ptc1 and the regulation of cell division. Thus, we propose that ptc1 participates in a G(2)/M checkpoint by regulating the localization of MPF.
Insights
The tumor suppressor patched1 (ptc1) interacts with cyclin B1, regulating its cell localization and impacting cell division. Sonic hedgehog signaling disrupts this interaction, allowing cell cycle progression.
Area of Science:
- Cell Biology
- Molecular Biology
- Cancer Research
Background:
- Mitosis initiation depends on M-phase promoting factor (MPF) activation.
- MPF regulation involves cdc2 and cyclin B1 phosphorylation and localization.
- Patched1 (ptc1) is a tumor suppressor implicated in basal cell carcinoma (BCC).
Purpose of the Study:
- To identify novel interactions with phosphorylated cyclin B1.
- To investigate the role of ptc1 in cell division regulation.
- To explore the link between ptc1 tumor suppressor activity and cell cycle control.
Main Methods:
- Yeast two-hybrid screening with a phosphorylated cyclin B1 bait.
- Co-immunoprecipitation and subcellular localization studies in 293T cells.
- Analysis of cell proliferation upon ptc1 and cyclin B1 derivative expression.
Main Results:
- Identified a novel interaction between cyclin B1 and ptc1.
- Ptc1 specifically binds phosphorylated cyclin B1, altering its localization.
- Sonic hedgehog (shh) disrupts the ptc1-cyclin B1 interaction, promoting nuclear cyclin B1 localization.
- Ptc1 inhibits cell proliferation, an effect reversed by a non-interacting cyclin B1 mutant.
- Endogenous ptc1 and cyclin B1 interact in vivo.
Conclusions:
- Ptc1 regulates the subcellular localization of cyclin B1.
- Ptc1's tumor suppressor function is linked to cell division regulation.
- Ptc1 likely acts in a G(2)/M checkpoint by controlling MPF localization.