Quantitative Proteomics Illuminates a Functional Interaction between mDia2 and the Proteasome

Tadamoto Isogai, Rob Van Der Kammen, Onno B. Bleijerveld, Soenita S. Goerdayal, Elisabetta Argenzio, A. F.Maarten Altelaar, Metello Innocenti

Research output: Contribution to journalArticle

8 Scopus citations

Abstract

Formin mDia2 is a cytoskeleton-regulatory protein that switches reversibly between a closed, autoinhibited and an open, active conformation. Although the open conformation of mDia2 induces actin assembly thereby controlling many cellular processes, mDia2 possesses also actin-independent and conformation-insensitive scaffolding roles related to microtubules and p53, respectively. Thus, we hypothesize that mDia2 may have other unappreciated functions and regulatory modes. Here we identify and validate proteasome and Ubiquitin as mDia2-interacting partners using stable isotope labeling with amino acids in cell culture-based quantitative proteomics and biochemistry, respectively. Although mDia2 is ubiquitinated, binds ubiquitinated proteins and free Ubiquitin, it is not a proteasome substrate. Surprisingly, knockdown of mDia2 increases the activity of the proteasome in vitro, whereas mDia2 overexpression has opposite effects only when it adopts the open conformation and cannot induce actin assembly. Consistently, a combination of candidate and unbiased proteome-wide analyses indicates that mDia2 regulates the cellular levels of proteasome substrate β-catenin and a number of ubiquitinated actin-regulatory proteins. Hence, these findings add more complexity to the mDia2 activity cycle by showing that the open conformation may control actin dynamics also through actin-independent regulation of the proteasome.

Original languageEnglish (US)
Pages (from-to)4624-4637
Number of pages14
JournalJournal of Proteome Research
Volume15
Issue number12
DOIs
StatePublished - Dec 2 2016
Externally publishedYes

Keywords

  • affinity purification
  • biochemistry
  • Formin
  • mass spectrometry
  • quantitative proteomics
  • SILAC

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)

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  • Cite this

    Isogai, T., Van Der Kammen, R., Bleijerveld, O. B., Goerdayal, S. S., Argenzio, E., Altelaar, A. F. M., & Innocenti, M. (2016). Quantitative Proteomics Illuminates a Functional Interaction between mDia2 and the Proteasome. Journal of Proteome Research, 15(12), 4624-4637. https://doi.org/10.1021/acs.jproteome.6b00718