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Assay for Adhesion and Agar Invasion in S. cerevisiae
Published on: November 8, 2006
Localization of Ras signaling complex in budding yeast
Fiorella Belotti1, Renata Tisi, Chiara Paiardi
1Department of Biotechnology and Biosciences, Umiversity of Milano-Bicocca, Milan, Italy.
Biochimica Et Biophysica Acta
|May 12, 2012
Summary
The Ras signaling complex in yeast localizes to different cellular membranes, including ER and mitochondria, not just the plasma membrane. Protein kinase A (PKA) activity influences this localization, impacting cellular processes.
Area of Science:
- Cellular Biology
- Molecular Biology
- Yeast Genetics
Background:
- The cAMP/protein kinase A (pKA) pathway is crucial for regulating metabolism, stress resistance, and proliferation in Saccharomyces cerevisiae.
- Ras proteins, regulated by guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs), are key activators of adenylate cyclase, which produces cAMP.
- Recent findings suggest Ras proteins and associated GEFs are found in internal membranes, challenging the traditional plasma membrane localization model.
Purpose of the Study:
- To investigate the subcellular localization of Ras signaling complex proteins within Saccharomyces cerevisiae.
- To determine if different components of the Ras signaling pathway reside in distinct cellular compartments.
- To explore the role of protein kinase A (PKA) in regulating the localization of Ras signaling complex proteins.
Main Methods:
- Fluorescent tagging of Ras signaling complex proteins for visualization.
- Biochemical cell membrane fractionation using sucrose gradients.
- Analysis of protein localization under conditions of altered PKA activity (Tpk1 overexpression).
Main Results:
- A minor fraction of Ras signaling complex components was detected at the plasma membrane during exponential growth on glucose.
- Cdc25 (RasGEF) predominantly localized to ER membranes.
- Ira2 (GAP) and Cyr1 (adenylate cyclase) were significantly present on mitochondria, with Ira2 relocating to ER membranes upon Tpk1 overexpression.
Conclusions:
- The Ras signaling pathway in yeast is compartmentalized, with different components localizing to distinct subcellular membranes (ER, mitochondria).
- Subcellular relocalization of Ras signaling complex proteins is dynamically regulated by PKA activity.
- These findings support a model where distinct branches of Ras signaling operate in different cellular compartments, influencing cellular responses.
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