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Identification of Kinase-substrate Pairs Using High Throughput Screening
11:13

Identification of Kinase-substrate Pairs Using High Throughput Screening

Published on: August 29, 2015

A phospho-proteomic screen identifies novel S6K1 and mTORC1 substrates revealing additional complexity in the

Katarzyna Jastrzebski1, Katherine M Hannan, Colin M House

  • 1Research Division, Peter MacCallum Cancer Centre, Melbourne, Locked Bag 1, A'Beckett Street, Victoria 8006, Australia.

Cellular Signalling
|March 29, 2011
PubMed

Insights

The nuclear p85 ribosomal S6 kinase 1 (S6K1) isoform

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • Ribosomal protein S6 kinase 1 (S6K1) is a key mediator of mTORC1 signaling, regulating cell growth and protein synthesis.
  • While the cytoplasmic p70(S6K1) isoform's role in translation is well-established, the nuclear p85(S6K1) isoform's functions and substrates remain largely unknown.

Purpose of the Study:

  • To identify novel nuclear p85(S6K1) substrates using a phospho-proteomic screen.
  • To elucidate the functional consequences of p85(S6K1) phosphorylation on its substrates.

Main Methods:

  • Phospho-proteomic screening to identify p85(S6K1) targets.
  • Site-directed mutagenesis and in vitro assays to investigate phosphorylation site function.
  • Biochemical assays to assess chaperonin folding activity.

Main Results:

  • Identified four novel p85(S6K1) substrates (GRP75, CCTβ, PGK1, RACK1) and two mTORC1 substrates (ANXA4, PSMA6).
  • Determined that phosphorylation of the chaperonin subunit CCTβ at serine 260 modulates its folding activity.
  • The identified substrates are involved in diverse cellular processes including chaperone function, metabolism, and proteasome activity.

Conclusions:

  • The nuclear p85(S6K1) isoform phosphorylates diverse proteins involved in fundamental cellular processes.
  • Phosphorylation of CCTβ by p85(S6K1) directly impacts chaperonin folding activity, suggesting a role in regulating protein homeostasis.
  • This study expands the known functions of S6K1 into the nucleus, revealing new regulatory mechanisms in cell growth and protein synthesis.

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