Abstract
In eukaryotes, nutrient availability and metabolism are coordinated by sensing mechanisms and signaling pathways, which influence a broad set of cellular functions such as transcription and metabolic pathways to match environmental conditions. In yeast, PKA is activated in the presence of high glucose concentrations, favoring fast nutrient utilization, shutting down stress responses, and boosting growth. On the contrary, Snf1/AMPK is activated in the presence of low glucose or alternative carbon sources, thus promoting an energy saving program through transcriptional activation and phosphorylation of metabolic enzymes. The PKA and Snf1/AMPK pathways share common downstream targets. Moreover, PKA has been reported to negatively influence the activation of Snf1/AMPK. We report a new cross-talk mechanism with a Snf1-dependent regulation of the PKA pathway. We show that Snf1 and adenylate cyclase (Cyr1) interact in a nutrient-independent manner. Moreover, we identify Cyr1 as a Snf1 substrate and show that Snf1 activation state influences Cyr1 phosphorylation pattern, cAMP intracellular levels, and PKA-dependent transcription.
Highlights
The Snf1/AMPK and PKA pathways are crucial for nutrient sensing and utilization in yeast
Because the cellular metabolic behavior could be the result of the action of different and somehow opposite signaling pathways, we wondered whether Snf1 could directly regulate such pathways
Approach, immunoprecipitating myc-tagged Snf1 and detecting co-immunoprecipitated proteins with mass spectrometry after resolution with SDS-PAGE. We performed this experiment with protein extracts of exponentially growing cells in 2% glucose, because we already demonstrated that in this condition Snf1 is at least partially functional [22, 25]
Summary
The Snf1/AMPK and PKA pathways are crucial for nutrient sensing and utilization in yeast. Active Snf phosphorylates Cyr and negatively regulates cAMP content and PKA-dependent transcription. PKA phosphorylates and deactivates Adr, the transcriptional activator of several glucose repressed genes, whereas Snf indirectly causes its dephosphorylation and activation [33, 34]. Both PKA and Snf act as repressors of the transcription factor Msn, but, whereas PKA is the main regulator of this factor, Snf targets it only to shut off transcription of the stress response element regulon after prolonged stationary phase [9, 35, 36]. We demonstrate that constitutive activation of Snf reduces cAMP intracellular content and functionally impairs PKA activity as a regulator of glucose-repressed genes
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