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A Mass Spectrometry-Based Approach to Identify Phosphoprotein Phosphatases and their Interactors
10:17

A Mass Spectrometry-Based Approach to Identify Phosphoprotein Phosphatases and their Interactors

Published on: April 29, 2022

PPP1CC2 can form a kinase/phosphatase complex with the testis-specific proteins TSSK1 and TSKS in the mouse testis

Graham MacLeod1, Peng Shang, Gregory T Booth

  • 1Department of Cell and Systems Biology, University of Toronto, 25 Harbord Street, Toronto, Ontario, Canada M5S 3G5.

Reproduction (Cambridge, England)
|October 4, 2013
PubMed

Insights

Protein phosphatase 1 regulatory subunit 15B (PPP1CC) is crucial for male fertility and spermatogenesis. A novel kinase/phosphatase complex involving PPP1CC2, TSSK1, and TSKS plays a key role in this process.

Area of Science:

  • Reproductive Biology
  • Molecular Cell Biology
  • Biochemistry

Background:

  • The mouse protein phosphatase gene Ppp1cc is essential for male fertility, with mutants exhibiting spermatogenesis failure.
  • Mutations lead to germ cell loss and mitochondrial sheath abnormalities, hypothesized to cause loss of the testis-specific isoform PPP1CC2.

Purpose of the Study:

  • To identify proteins interacting with PPP1CC2 that are involved in spermatogenesis.
  • To elucidate the molecular mechanisms underlying the role of PPP1CC2 in male fertility.

Main Methods:

  • GST pull-down assays using mouse testis lysates to identify interacting proteins.
  • Interaction experiments, phosphoproteomic analysis, and immunohistochemical analysis.
  • Electron microscopy to compare Ppp1cc and Tssk1/2 knockout phenotypes.

Main Results:

  • Identified testis-specific protein kinase TSSK1 as a PPP1CC2 interactor.
  • Confirmed a complex formation between PPP1CC2 and TSSK1, mediated by TSKS.
  • Discovered a novel phosphorylation site on TSKS that regulates PPP1CC2 binding.
  • Observed abnormalities in TSSK1 and TSKS distribution and mitochondrial sheath morphology in Ppp1cc mutants.

Conclusions:

  • Demonstrated a novel testis-specific kinase/phosphatase complex (PPP1CC2/TSSK1/TSKS).
  • This complex is critical for the completion of spermatogenesis.
  • The findings provide new insights into the molecular regulation of male fertility.

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