On Broken Ne(c)ks and Broken DNA: The Role of Human NEKs in the DNA Damage Response

Abstract

NIMA-related kinases, or NEKs, are a family of Ser/Thr protein kinases involved in cell cycle and mitosis, centrosome disjunction, primary cilia functions, and DNA damage responses among other biological functional contexts in vertebrate cells. In human cells, there are 11 members, termed NEK1 to 11, and the research has mainly focused on exploring the more predominant roles of NEKs in mitosis regulation and cell cycle. A possible important role of NEKs in DNA damage response (DDR) first emerged for NEK1, but recent studies for most NEKs showed participation in DDR. A detailed analysis of the protein interactions, phosphorylation events, and studies of functional aspects of NEKs from the literature led us to propose a more general role of NEKs in DDR. In this review, we express that NEK1 is an activator of ataxia telangiectasia and Rad3-related (ATR), and its activation results in cell cycle arrest, guaranteeing DNA repair while activating specific repair pathways such as homology repair (HR) and DNA double-strand break (DSB) repair. For NEK2, 6, 8, 9, and 11, we found a role downstream of ATR and ataxia telangiectasia mutated (ATM) that results in cell cycle arrest, but details of possible activated repair pathways are still being investigated. NEK4 shows a connection to the regulation of the nonhomologous end-joining (NHEJ) repair of DNA DSBs, through recruitment of DNA-PK to DNA damage foci. NEK5 interacts with topoisomerase IIβ, and its knockdown results in the accumulation of damaged DNA. NEK7 has a regulatory role in the detection of oxidative damage to telomeric DNA. Finally, NEK10 has recently been shown to phosphorylate p53 at Y327, promoting cell cycle arrest after exposure to DNA damaging agents. In summary, this review highlights important discoveries of the ever-growing involvement of NEK kinases in the DDR pathways. A better understanding of these roles may open new diagnostic possibilities or pharmaceutical interventions regarding the chemo-sensitizing inhibition of NEKs in various forms of cancer and other diseases.

Keywords: DNA damage response; cell cycle; kinase; protein kinase.

Figure 1

NEK1 and NEK2 play key roles in the DDR pathway. NEK1 is required for replication fork stability by phosphorylating Rad54 at S572. Moreover, the TLK1 > NEK1 > ATR > CHK1 axis plays a major role in DDR, involving cell cycle arrest through CDC25A phosphorylation and CDK1 inhibition. TLK1 phosphorylates NEK1 at T141 and NEK1 interacts with ATR, promoting the stabilization of ATR-ATRIP complex and autophosphorylation of ATR at T1989. NEK2 regulates centrosome disjunction since ATM activates the PP1 phosphatase, which in turn dephosphorylates NEK2, leading to the inhibition of centrosome splitting. The PP1a phosphatase activation also counteracts the NEK2-dependent phosphorylation of NPM/B23, which stabilizes the ARF pathway. NEK2 also upregulates the Wnt1/β-catenin pathway.

Figure 2

NEK3, NEK4, and NEK5 roles in DDR. The interaction of NEK3 and PCNA is a possible link between NEK3 and DDR. NEK4 interacts with the DNA-PK complex and is important for efficient DNA-PKcs recruitment to DNA damage foci, as well as the activation of pathways essential to induce cell cycle arrest after DSBs with IR or etoposide. We propose that NEK4 acts very early in DNA break recognition, phosphorylating Ku70 (possibly at S155 residue) and stabilizing the DNA-PK complex. Also, NEK4 possibly interacts with other NHEJ-related proteins, such as H2AX, and KAP-1, and PCNA, and FEN1. The interaction between NEK5 and topoisomerase IIβ might be related to cell cycle halting due to DNA damage, as this interaction increases during the first stage of DDR. NEK5-depleted cells overcome the G2/M checkpoint upon DNA damage, indicating that NEK5 is required for an appropriate DDR.

Figure 3

The possible involvement of NEK6 and NEK7 in DDR. NEK6 kinase activity is inhibited by ATM/ATR-Chk1/Chk2 caffeine-sensitive signaling pathway after DSBs induced by IR and UV treatment. NEK6 kinase activity inhibition by DDR is necessary for G2/M arrest. NEK6 is repressed by miR-23a, which is induced by berberine, a genotoxin that increases the accumulation of DSBs, in a p53-dependent manner. NEK7 is related to a protective function of telomeres in response to oxidative DNA damage. ATM activation mediates the function of NEK7 in telomeres. NEK7 is recruited to the telomeres and binds and phosphorylates TRF1 in S114, preventing its degradation. The integrity of telomeres is linked to the stability of TRF1.

Figure 4

Signaling pathways involving NEK8 and NEK9 in response to replication stress. NEK8 prevents DSB accumulation by suppressing cyclin A-associated CDK activity. NEK8 interacts with ATR, ATRIP (ATR-interacting protein), and CHK1 proteins after DNA damage and is important for the progression of replication forks. NEK8 also regulates RAD51 foci formation upon DNA damage. NEK9 is a replication stress response (RSR) protein since agents involved in replication stress increase its expression. NEK9 interacts with CHK1 and its knockdown decreases CHK1 autophosphorylation (S296) and its kinase activity in response to replicative stress.

Figure 5

The signaling pathway of NEK10 and NEK11 upon DNA damage. NEK10 assembles in a ternary complex with RAF-1 and MEK1 and mediates ERK1/2 activation, playing a role in the maintenance of the G2/M checkpoint after UV light irradiation. Upon Cisplatin and IR treatments, NEK10 phosphorylates p53 at Y327, increasing p21 levels. p21 is a cyclin-dependent kinase inhibitor that suppresses the cell cycle G1/S phase. NEK10 participates in the control of cell cycle progression through p53 phosphorylation. Upon DNA damage, NEK11 is involved with ATM/ATR checkpoint pathway. Degradation of CDC25A is regulated by NEK11 through the phosphorylation of S82 and S88 of CDC25A, in vivo. CHK1 is responsible for phosphorylating NEK11 at the S273 site in vitro.