Dataset Information

CDK and MAPK collaborate to regulate signaling dynamics via multi-site phosphorylation of the scaffold protein Ste5

ABSTRACT: CDKs and MAPKs phosphorylate similar sites yet generally have distinct functions/substrates. We report here an unanticipated system of cooperative regulation by CDK and MAPK, involving collaborative multi-site phosphorylation of a single substrate. The budding yeast protein Ste5 is a signaling scaffold for the pheromone-activated G1 arrest pathway. Upon cell cycle entry, CDK activity inhibits Ste5 via phosphorylation at numerous sites flanking its membrane-binding domain. Ste5 is also regulated by negative feedback from the pathway MAPK, though the mechanism was unknown. Using quantitative time-lapse microscopy, we examined Ste5 membrane recruitment dynamics at different cell cycle stages. Surprisingly, at stages where we expected Ste5 recruitment would be blocked, we observed transient recruitment followed by a steep decline, depending on both CDK and MAPK activities. The collective results of mutagenesis, mass spectrometry, and electrophoretic analyses suggest that the CDK and MAPK target shared sites, and that the substrate is most extensively phosphorylated when both kinases are active and able to bind their respective docking sites on Ste5. This collaborative phosphorylation can diversify regulatory options, ranging from mild tuning to strong blocking, and can yield distinct patterns of regulatory dynamics at different cell cycle stages.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Saccharomyces Cerevisiae (baker's Yeast)

TISSUE(S): Haploid Cell

SUBMITTER: Wolfgang Reiter

LAB HEAD: Wolfgang Reiter

PROVIDER: PXD006154 | Pride | 2018-03-12

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	20140818_QEx6_RSLC4_Reiter_Ammerer_MFPL_onbead_ste5_cdc15ts_10min_sample4_run1_5per.raw	Raw
	20140818_QEx6_RSLC4_Reiter_Ammerer_MFPL_onbead_ste5_cdc15ts_10min_sample4_run1_5per.tsv	Tabular
	20140818_QEx6_RSLC4_Reiter_Ammerer_MFPL_onbead_ste5_cdc15ts_10min_sample4_run2_5per.raw	Raw
	20140818_QEx6_RSLC4_Reiter_Ammerer_MFPL_onbead_ste5_cdc15ts_10min_sample4_run2_5per.tsv	Tabular
	20140818_QEx6_RSLC4_Reiter_Ammerer_MFPL_onbead_ste5_cdc15ts_70min_sample4_run1_5per.raw	Raw

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Publications

CDK and MAPK Synergistically Regulate Signaling Dynamics via a Shared Multi-site Phosphorylation Region on the Scaffold Protein Ste5.

Repetto María Victoria MV Winters Matthew J MJ Bush Alan A Reiter Wolfgang W Hollenstein David Maria DM Ammerer Gustav G Pryciak Peter M PM Colman-Lerner Alejandro A

Molecular cell 20180301 6

We report an unanticipated system of joint regulation by cyclin-dependent kinase (CDK) and mitogen-activated protein kinase (MAPK), involving collaborative multi-site phosphorylation of a single substrate. In budding yeast, the protein Ste5 controls signaling through a G1 arrest pathway. Upon cell-cycle entry, CDK inhibits Ste5 via multiple phosphorylation sites, disrupting its membrane association. Using quantitative time-lapse microscopy, we examined Ste5 membrane recruitment dynamics at diffe ...[more]

PMID: 29547722

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