Regulation of GATA-binding protein 2 levels via ubiquitin-dependent degradation by Fbw7: Involvement of cyclin B-cyclin-dependent kinase 1-mediated phosphorylation of Thr176 in GATA-binding protein 2

Tomomi Nakajima, Kyoko Kitagawa, Tatsuya Ohhata, Satoshi Sakai, Chiharu Uchida, Kiyoshi Shibata, Naoko Minegishi, Kanae Yumimoto, Keiichi I. Nakayama, Kazuma Masumoto, Fuminori Katou, Hiroyuki Niida, Masatoshi Kitagawa

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Abstract

AGATA family transcription factor, GATA-binding protein 2 (GATA2), participates in cell growth and differentiation of various cells, such as hematopoietic stem cells. Although its expression level is controlled by transcriptional induction and proteolytic degradation, the responsible E3 ligase has not been identified. Here, we demonstrate that F-box/WD repeat-containing protein 7 (Fbw7/Fbxw7), a component of Skp1, Cullin 1, F-box-containing complex (SCF)-type E3 ligase, is an E3 ligase for GATA2. GATA2 contains a cell division control protein 4 (Cdc4) phosphodegron (CPD), a consensus motif for ubiquitylation by Fbw7, which includes Thr176. Ectopic expression of Fbw7 destabilized GATA2 and promoted its proteasomal degradation. Substitution of threonine 176 to alanine in GATA2 inhibited binding with Fbw7, and the ubiquitylation and degradation of GATA2 by Fbw7 was suppressed. The CPD kinase, which mediates the phosphorylation of Thr176, was cyclin B-cyclin-dependent kinase 1 (CDK1). Moreover, depletion of endogenous Fbw7 stabilized endogenous GATA2 in K562 cells. Conditional Fbw7 depletion in mice increased GATA2 levels in hematopoietic stem cells and myeloid progenitors at the early stage. Increased GATA2 levels in Fbw7-conditional knock-out mice were correlated with a decrease in a c-Kit high expressing population of myeloid progenitor cells. Our results suggest that Fbw7 is a bona fide E3 ubiquitin ligase for GATA2 in vivo.

Original languageEnglish
Pages (from-to)10368-10381
Number of pages14
JournalJournal of Biological Chemistry
Volume290
Issue number16
DOIs
Publication statusPublished - Apr 17 2015

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GATA2 Transcription Factor
CDC2 Protein Kinase
Cyclin B
Phosphorylation
Ubiquitin
Degradation
Ubiquitin-Protein Ligases
Ubiquitination
Hematopoietic Stem Cells
Stem cells
Myeloid Progenitor Cells
Cells
K562 Cells
Transcription factors
Threonine
Cell growth
Knockout Mice
Alanine
Cell Division
Cell Differentiation

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Regulation of GATA-binding protein 2 levels via ubiquitin-dependent degradation by Fbw7 : Involvement of cyclin B-cyclin-dependent kinase 1-mediated phosphorylation of Thr176 in GATA-binding protein 2. / Nakajima, Tomomi; Kitagawa, Kyoko; Ohhata, Tatsuya; Sakai, Satoshi; Uchida, Chiharu; Shibata, Kiyoshi; Minegishi, Naoko; Yumimoto, Kanae; Nakayama, Keiichi I.; Masumoto, Kazuma; Katou, Fuminori; Niida, Hiroyuki; Kitagawa, Masatoshi.

In: Journal of Biological Chemistry, Vol. 290, No. 16, 17.04.2015, p. 10368-10381.

Research output: Contribution to journalArticle

Nakajima, T, Kitagawa, K, Ohhata, T, Sakai, S, Uchida, C, Shibata, K, Minegishi, N, Yumimoto, K, Nakayama, KI, Masumoto, K, Katou, F, Niida, H & Kitagawa, M 2015, 'Regulation of GATA-binding protein 2 levels via ubiquitin-dependent degradation by Fbw7: Involvement of cyclin B-cyclin-dependent kinase 1-mediated phosphorylation of Thr176 in GATA-binding protein 2', Journal of Biological Chemistry, vol. 290, no. 16, pp. 10368-10381. https://doi.org/10.1074/jbc.M114.613018
Nakajima, Tomomi ; Kitagawa, Kyoko ; Ohhata, Tatsuya ; Sakai, Satoshi ; Uchida, Chiharu ; Shibata, Kiyoshi ; Minegishi, Naoko ; Yumimoto, Kanae ; Nakayama, Keiichi I. ; Masumoto, Kazuma ; Katou, Fuminori ; Niida, Hiroyuki ; Kitagawa, Masatoshi. / Regulation of GATA-binding protein 2 levels via ubiquitin-dependent degradation by Fbw7 : Involvement of cyclin B-cyclin-dependent kinase 1-mediated phosphorylation of Thr176 in GATA-binding protein 2. In: Journal of Biological Chemistry. 2015 ; Vol. 290, No. 16. pp. 10368-10381.
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abstract = "AGATA family transcription factor, GATA-binding protein 2 (GATA2), participates in cell growth and differentiation of various cells, such as hematopoietic stem cells. Although its expression level is controlled by transcriptional induction and proteolytic degradation, the responsible E3 ligase has not been identified. Here, we demonstrate that F-box/WD repeat-containing protein 7 (Fbw7/Fbxw7), a component of Skp1, Cullin 1, F-box-containing complex (SCF)-type E3 ligase, is an E3 ligase for GATA2. GATA2 contains a cell division control protein 4 (Cdc4) phosphodegron (CPD), a consensus motif for ubiquitylation by Fbw7, which includes Thr176. Ectopic expression of Fbw7 destabilized GATA2 and promoted its proteasomal degradation. Substitution of threonine 176 to alanine in GATA2 inhibited binding with Fbw7, and the ubiquitylation and degradation of GATA2 by Fbw7 was suppressed. The CPD kinase, which mediates the phosphorylation of Thr176, was cyclin B-cyclin-dependent kinase 1 (CDK1). Moreover, depletion of endogenous Fbw7 stabilized endogenous GATA2 in K562 cells. Conditional Fbw7 depletion in mice increased GATA2 levels in hematopoietic stem cells and myeloid progenitors at the early stage. Increased GATA2 levels in Fbw7-conditional knock-out mice were correlated with a decrease in a c-Kit high expressing population of myeloid progenitor cells. Our results suggest that Fbw7 is a bona fide E3 ubiquitin ligase for GATA2 in vivo.",
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AU - Ohhata, Tatsuya

AU - Sakai, Satoshi

AU - Uchida, Chiharu

AU - Shibata, Kiyoshi

AU - Minegishi, Naoko

AU - Yumimoto, Kanae

AU - Nakayama, Keiichi I.

AU - Masumoto, Kazuma

AU - Katou, Fuminori

AU - Niida, Hiroyuki

AU - Kitagawa, Masatoshi

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