Journal
BIOCHEMICAL JOURNAL
Volume 457, Issue -, Pages 43-56Publisher
PORTLAND PRESS LTD
DOI: 10.1042/BJ20130461
Keywords
AZ191; cell cycle; cyclin D1; dual-specificity tyrosine phosphorylation-regulated kinase 1B (DYRK1B); glycogen synthase kinase 3 beta (GSK3 beta)
Categories
Funding
- Biotechnology and Biological Sciences Research Council via a CASE PhD studentship [3068901]
- Babraham Institute
- Institute Strategic Programme Grant [BBS/E/B/000C0417]
- Association for International Cancer Research [AICR09-0257]
- BBSRC [BBS/E/B/000C0417, BB/L008793/1] Funding Source: UKRI
- Biotechnology and Biological Sciences Research Council [BBS/B/01111, BBS/E/B/000C0417, BB/L008793/1, BBS/E/B/0000C199] Funding Source: researchfish
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DYRK1B (dual-specificity tyrosine phosphorylation-regulated kinase 1B) is amplified in certain cancers and may be an oncogene; however, our knowledge of DYRK1B has been limited by the lack of selective inhibitors. In the present study we describe AZ191, a potent small molecule inhibitor that selectively inhibits DYRK1B in vitro and in cells. CCND1 (cyclin D1), a key regulator of the mammalian G(1)-S-phase transition, is phosphorylated on Thr(286) by GSK3 beta (glycogen synthase kinase 3 beta) to promote its degradation. DYRK1B has also been proposed to promote CCND1 turnover, but was reported to phosphorylate Thr(288) rather than Thr(286). Using in vitro kinase assays, phospho-specific immunoblot analysis and MS in conjunction with AZ191 we now show that DYRK1B phosphorylates CCND1 at Thr(286), not Thr(288), in vitro and in cells. In HEK (human embryonic kidney)-293 and PANC-1 cells (which exhibit DYRK1B amplification) DYRK1B drives Thr(286) phosphorylation and proteasome-dependent turnover of CCND1 and this is abolished by AZ191 or DYRK1B RNAi, but not by GSK3 beta inhibitors or GSK3 beta RNAi. DYRK1B expression causes a G(1)-phase cell-cycle arrest, but overexpression of CCND1 (wild-type or T286A) fails to overcome this; indeed, DYRK1B also promotes the expression of p21(CIPI) (21 kDa CDK-interacting protein 1) and p27(KIPI) (CDK-inhibitory protein 1). The results of the present study demonstrate for the first time that DYRK1B is a novel Thr(286)-CCND1 kinase that acts independently of GSK3 beta to promote CCND1 degradation. Furthermore, we anticipate that AZ191 may prove useful in defining further substrates and biological functions of DYRK1B.
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