Kinase Subfamily NEK9

Kinase Classification: Group Other: Family NEK: Subfamily NEK9

NEK9 is a mitotic NEK protein found in some animals, which phosphorylates and activates NEK6 subfamily kinases.

Evolution

NEK9 is present as a single copy in animal and choanoflagellate genomes, but is repeatedly lost, including in sponges, arthropods (including Drosophila), and nematodes (including C. elegans).

Domain Structure

Like NEK8, NEK9 has a N-terminal kinase domain followed by 7 RCC1 domains (356-788 in human NEK9) that form a beta-propellor structure, followed by an extended C-terminal region that includes a coiled-coil region (disordered regions and some short conserved segments. A highly conserved region (human: 810-828, GWLRKELENAEFIPMPDSP) binds to and activates the NEK7 kinase domain by displacing an autoinhibitory region [1], while another conserved region, WCLL form 967-970 is an LC3B-binding motif.

Functions

NEK9 is activated during mitosis and in turn, activates the NEK6 subfamily members NEK6 and NEK7 in mammals, by phosphorylation of their activation loops. NEK9 also phosphorylates the NLS of the nuclear microtubule-associated protein TPX2, keeping it cytoplasmic before mitosis [2], enabling centrosome development. Nek9 phosphorylates NEDD1 on Ser377 driving recruitment of NEDD1 and γ-tubulin to the mitotic centrosome [3].

NEK9 is phosphorylated at the centrosomes during prophase, probably by Plk1 and CDK1 upstream kinases, regulating centrosome separation, leading to mitosis [4]. NEK9 is required during G1-S transition, and is known to bind the FACT complex and be activated by it during this phase [5], while loss of NEK9 prolongs G1-S.

A null human NEK9 mutation is homozgous lethal, with severe skeletal dysplasia, reduced proliferation in isolated fibroblasts, and a reduction in cilia [6], which may be the cause of the skeletal phenotype. Somatic gain-of-function mutations in NEK9 were found in several cases of nevus comedonicus, a rare inflammatory acne syndrome, suggesting that NEK9 may modulate follicle cell differentiation [7].

NEK9 knockdown selectively inhibited cell growth in p53-deficient cells [8]. In gastric cancer, NEK9 was downregulated by miR-520f-3p. IL6-STAT3 signaling surpressed this, and the elevated NEK9 was shown to phosphoryate ARHGEF2, impacting RhoA activation and cell motility

Peptide array profiling of NEK kinases shows that the in vitro substrate specificity of NEK9 was similar to that of NEK6/7, with a preference for acidic residues at the -2 position, and somewhat at -4 and -5, unlike other NEKs. This profile is similar to that of the NEK9-activating kinase, PLK1, suggesting that NEK9/6 subfamilies may amplify the PLK1 phosphorylation signal [9].

NEK9 phosphorylates LC3B on T50, and binds to it via a C-terminal motif (WCLL, 967-970 on human NEK9) [10]. This motif also binds the autophagy protein ATG8a (GABARAP), and may regulate ciliogenesis by binding and autophagic degradation of myosin IIA (MYH9) [11]

References

1. Richards MW, O'Regan L, Mas-Droux C, Blot JM, Cheung J, Hoelder S, Fry AM, and Bayliss R. An autoinhibitory tyrosine motif in the cell-cycle-regulated Nek7 kinase is released through binding of Nek9. Mol Cell. 2009 Nov 25;36(4):560-70. DOI:10.1016/j.molcel.2009.09.038 |
2. Eibes S, Gallisà-Suñé N, Rosas-Salvans M, Martínez-Delgado P, Vernos I, and Roig J. Nek9 Phosphorylation Defines a New Role for TPX2 in Eg5-Dependent Centrosome Separation before Nuclear Envelope Breakdown. Curr Biol. 2018 Jan 8;28(1):121-129.e4. DOI:10.1016/j.cub.2017.11.046 |
3. Sdelci S, Schütz M, Pinyol R, Bertran MT, Regué L, Caelles C, Vernos I, and Roig J. Nek9 phosphorylation of NEDD1/GCP-WD contributes to Plk1 control of γ-tubulin recruitment to the mitotic centrosome. Curr Biol. 2012 Aug 21;22(16):1516-23. DOI:10.1016/j.cub.2012.06.027 |
4. Bertran MT, Sdelci S, Regué L, Avruch J, Caelles C, and Roig J. Nek9 is a Plk1-activated kinase that controls early centrosome separation through Nek6/7 and Eg5. EMBO J. 2011 Jun 3;30(13):2634-47. DOI:10.1038/emboj.2011.179 |
5. Tan BC and Lee SC. Nek9, a novel FACT-associated protein, modulates interphase progression. J Biol Chem. 2004 Mar 5;279(10):9321-30. DOI:10.1074/jbc.M311477200 |
6. Casey JP, Brennan K, Scheidel N, McGettigan P, Lavin PT, Carter S, Ennis S, Dorkins H, Ghali N, Blacque OE, Mc Gee MM, Murphy H, and Lynch SA. Recessive NEK9 mutation causes a lethal skeletal dysplasia with evidence of cell cycle and ciliary defects. Hum Mol Genet. 2016 May 1;25(9):1824-35. DOI:10.1093/hmg/ddw054 |
7. Levinsohn JL, Sugarman JL, Yale Center for Mendelian Genomics, McNiff JM, Antaya RJ, and Choate KA. Somatic Mutations in NEK9 Cause Nevus Comedonicus. Am J Hum Genet. 2016 May 5;98(5):1030-1037. DOI:10.1016/j.ajhg.2016.03.019 |
8. Kurioka D, Takeshita F, Tsuta K, Sakamoto H, Watanabe S, Matsumoto K, Watanabe M, Nakagama H, Ochiya T, Yokota J, Kohno T, and Tsuchiya N. NEK9-dependent proliferation of cancer cells lacking functional p53. Sci Rep. 2014 Aug 18;4:6111. DOI:10.1038/srep06111 |
9. van de Kooij B, Creixell P, van Vlimmeren A, Joughin BA, Miller CJ, Haider N, Simpson CD, Linding R, Stambolic V, Turk BE, and Yaffe MB. Comprehensive substrate specificity profiling of the human Nek kinome reveals unexpected signaling outputs. Elife. 2019 May 24;8. DOI:10.7554/eLife.44635 |
10. Shrestha BK, Skytte Rasmussen M, Abudu YP, Bruun JA, Larsen KB, Alemu EA, Sjøttem E, Lamark T, and Johansen T. NIMA-related kinase 9-mediated phosphorylation of the microtubule-associated LC3B protein at Thr-50 suppresses selective autophagy of p62/sequestosome 1. J Biol Chem. 2020 Jan 31;295(5):1240-1260. DOI:10.1074/jbc.RA119.010068 |
11. Yamamoto Y, Chino H, Tsukamoto S, Ode KL, Ueda HR, and Mizushima N. NEK9 regulates primary cilia formation by acting as a selective autophagy adaptor for MYH9/myosin IIA. Nat Commun. 2021 Jun 2;12(1):3292. DOI:10.1038/s41467-021-23599-7 |