Functional characterization of C21ORF2 association with the NEK1 kinase mutated in human in diseases.

The NEK1 kinase controls ciliogenesis, mitosis, and DNA repair, and NEK1 mutations cause human diseases including axial spondylometaphyseal dysplasia and amyotrophic lateral sclerosis. C21ORF2 mutations cause a similar pattern of human diseases, suggesting close functional links with NEK1 Here, we report that endogenous NEK1 and C21ORF2 form a tight complex in human cells. A C21ORF2 interaction domain "CID" at the C-terminus of NEK1 is necessary for its association with C21ORF2 in cells, and pathogenic mutations in this region disrupt the complex. AlphaFold modelling predicts an extended binding interface between a leucine-rich repeat domain in C21ORF2 and the NEK1-CID, and our model may explain why pathogenic mutations perturb the complex. We show that NEK1 mutations that inhibit kinase activity or weaken its association with C21ORF2 severely compromise ciliogenesis, and that C21ORF2, like NEK1 is required for homologous recombination. These data enhance our understanding of how the NEK1 kinase is regulated, and they shed light on NEK1-C21ORF2-associated diseases.

A case of siblings with juvenile retinitis pigmentosa associated with NEK1 gene variants.

BACKGROUND: Axial spondylometaphyseal dysplasia(axial SMD) is associated with early-onset retinal dystrophy and various skeletal dysplasias of varying severity. NEK1 is the causative gene for short rib polydactyly syndrome and axial SMD. Here, we report a case of siblings with juvenile retinitis pigmentosa (RP) and NEK1 variants not associated with systemic disorders. MATERIALS AND METHODS: The patients were a 7-year-old-girl and a 9-year-old boy with RP, who were followed for 9 years. Whole exome sequencing (WES) was performed on the siblings and their parents, who were not consanguineous. RESULTS: The corrected visual acuity of the girl and the boy at first visit was binocular 20/63 and 20/100 OD and 20/63 OS, respectively. The siblings had narrowing of retinal blood vessels and retinal pigment epithelium atrophy in the fundus and showed an extinguished pattern in electroretinogram. On optical coherence tomography, there was a mottled ellipsoid band with progressive loss in the outer macular, the edges of which corresponded to the ring of hyperautofluorescence on fundus autofluorescence imaging. The siblings showed progressive visual field constriction. Radiological examination did not reveal any skeletal abnormalities. We identified two rare heterozygous NEK1 variants in the patients: c.240 G>A; p.(M80I) and c.634_639dup;p.(V212_L213dup). Heterozygous variants were recognized in the father and mother, respectively. According to the guidelines of the American College of Medical Genetics and Genomics, both variants were classified as likely pathogenic. CONCLUSION: This is the first report of RP patients with NEK1 variants not associated with skeletal abnormalities.

NEK1 and STMN2 short tandem repeat lengths are not associated with Australian amyotrophic lateral sclerosis risk.

Sporadic amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with a complex genetic architecture. The lengths of two short tandem repeats (STRs), at the NEK1 and STMN2 loci, were recently associated with ALS risk in cohorts of European descent. The STMN2 STR was proposed to be predictive of clinical features including the age of onset and disease duration in bulbar onset cases. We sought to investigate NEK1 and STMN2 STR lengths in a cohort of Australian sporadic ALS cases (n = 608) and neurologically healthy controls (n = 4689) of European ancestry. ExpansionHunter was used to determine NEK1 and STMN2 STR length genotypes from whole-genome sequencing data followed by PCR validation of predicted lengths. No significant association was identified between sporadic ALS risk and the length of either STR. Further, neither NEK1 nor STMN2 STR lengths were indicative of the age of onset or disease duration. We report that the NEK1 and STMN2 STRs were not associated with ALS risk or clinical features in this Australian sporadic ALS cohort.

Nek1-inhibitor and temozolomide-loaded microfibers as a co-therapy strategy for glioblastoma treatment.

Malignant glioblastoma (GB) is the predominant primary brain tumour in adults, but despite the efforts towards novel therapies, the median survival of GB patients has not significantly improved in the last decades. Therefore, localised approaches that treat GB straight into the tumour site provide an alternative to enhance chemotherapy bioavailability and efficacy, reducing systemic toxicity. Likewise, the discovery of protein targets, such as the NIMA-related kinase 1 (Nek1), which was previously shown to be associated with temozolomide (TMZ) resistance in GB, has stimulated the clinical development of target therapy approaches to treat GB patients. In this study, we report an electrospun polyvinyl alcohol (PVA) microfiber (MF) brain-implant prepared for the controlled release of Nek1 protein inhibitor (iNek1) and TMZ or TMZ-loaded nanoparticles. The formulations revealed adequate stability and drug loading, which prolonged the drugs' release allowing a sustained exposure of the GB cells to the treatment and enhancing the drugs' therapeutic effects. TMZ-loaded MF provided the highest concentration of TMZ within the brain of tumour-bearing rats, and it was statistically significant when compared to TMZ via intraperitoneal (IP). All animals treated with either co-therapy formulation (TMZ + iNek1 MF or TMZ nanoparticles + iNek1 MF) survived until the endpoint (60 days), whereas the Blank MF (drug-unloaded), TMZ MF and TMZ IP-treated rats' median survival was found to be 16, 31 and 25 days, respectively. The tumour/brain area ratio of the rats implanted with either MF co-therapy was found to be reduced by 5-fold when compared to Blank MF-implanted rats. Taken together, our results strongly suggest that Nek1 is an important GB oncotarget and the inhibition of Nek1's activity significantly decreases GB cells' viability and tumour size when combined with TMZ treatment.

Illuminating a Dark Kinase: Structure-Guided Design, Synthesis, and Evaluation of a Potent Nek1 Inhibitor and Its Effects on the Embryonic Zebrafish Pronephros.

NIMA-related kinase 1 (Nek1) has lately garnered attention for its widespread function in ciliogenesis, apoptosis, and the DNA-damage response. Despite its involvement in various diseases and its potential as a cancer drug target, no directed medicinal chemistry efforts toward inhibitors against this dark kinase are published. Here, we report the structure-guided design of a potent small-molecule Nek1 inhibitor, starting from a scaffold identified by kinase cross-screening analysis. Seven lead compounds were identified in silico and evaluated for their inhibitory activity. The top compound, 10f, was further profiled for efficacy, toxicity, and bioavailability in a zebrafish polycystic kidney disease model. Administration of 10f caused the expansion of fluorescence-labeled proximal convoluted tubules, supporting our hypothesis that Nek1-inhibition causes cystic kidneys in zebrafish embryos. Compound 10f displayed insignificant inhibition in 48 of 50 kinases in a selectivity test panel. The findings provide a powerful tool to further elucidate the function and pharmacology of this neglected kinase.

Design, synthesis and biological evaluation of novel aminopyrazole- and 7-azaindole-based Nek1 inhibitors and their effects on zebrafish kidney development.

NIMA-related protein kinase Nek1 is crucially involved in cell cycle regulation, DNA repair and microtubule regulation and dysfunctions of Nek1 play key roles in amyotrophic lateral sclerosis (ALS), polycystic kidney disease (PKD) and several types of radiotherapy resistant cancer. Targeting of Nek1 could reveal a new class of radiosensitizing substances and provide useful tools to better understand the aforementioned diseases. In this report we explore substituted aminopyrazoles and 7-azaindoles as potent inhibitors for the Nek1 kinase domain and examine their effect on kidney organogenesis in Danio rerio.

Mutation spectrum of amyotrophic lateral sclerosis in Central South China.

To analyze the mutational spectrum of known ALS causative genes in China ALS patients. We comprehensively analyzed 51 ALS causative genes by combining different sequencing technologies in 753 unrelated ALS patients from Central South China. The mean age at onset (AAO) was 53.7±11.4 years. The AAO was earlier in the autosomal dominant (AD) ALS patients than in the sporadic ALS (sALS) patients. Bulbar onset was more frequent in females than in males. SOD1 was the most frequently mutated gene in the AD-ALS and the sALS patients, followed by the ATXN2 and FUS genes in the AD-ALS patients and the NEK1 and CACNA1H genes in the sALS patients. Patients with RDVs in the SOD1 or FUS genes had an earlier AAO than the mean AAO of all the patients, while the patients with RDVs in the NEK1 gene showed later onset. SOD1 gene was the most commonly mutated gene in ALS patients in China, followed by ATXN2 and NEK1. The phenotype might be determined synergistically by sex and genetic variants.

Roles of NEK family in cell cycle regulation.

As a serine/threonine kinase, NIMA-related kinases (NEKs) play important roles in the regulation of cell cycle, and involve in several cellular activities such as centrosome separation, spindle assembly, chromatin condensation, nuclear envelope breakdown, spindle assembly checkpoint signaling, cytokinesis, cilia formation and DNA damage response. In this review, we summarize the component, structural characteristics and functions of NEK family in mitosis and meiosis based on the relevant researches in recent years, providing a reference for the further study on the roles of NEKs in the regulation of cell cycle and a theoretical basis for the clinical diagnosis and treatment of tumors.

Human Aurora kinase inhibitor Hesperadin reveals epistatic interaction between Plasmodium falciparum PfArk1 and PfNek1 kinases.

Mitosis has been validated by numerous anti-cancer drugs as being a druggable process, and selective inhibition of parasite proliferation provides an obvious opportunity for therapeutic intervention against malaria. Mitosis is controlled through the interplay between several protein kinases and phosphatases. We show here that inhibitors of human mitotic kinases belonging to the Aurora family inhibit P. falciparum proliferation in vitro with various potencies, and that a genetic selection for mutant parasites resistant to one of the drugs, Hesperadin, identifies a resistance mechanism mediated by a member of a different kinase family, PfNek1 (PF3D7_1228300). Intriguingly, loss of PfNek1 catalytic activity provides protection against drug action. This points to an undescribed functional interaction between Ark and Nek kinases and shows that existing inhibitors can be used to validate additional essential and druggable kinase functions in the parasite.

Protein-Bound Polysaccharides from Coriolus Versicolor Induce RIPK1/RIPK3/MLKL-Mediated Necroptosis in ER-Positive Breast Cancer and Amelanotic Melanoma Cells.

BACKGROUND/AIMS: The induction of necroptosis, a form of caspase-independent cell death, represents one of the most promising anticancer therapeutic modalities, as necroptosis serves as an alternative way to eliminate apoptosis-resistant tumor cells. Here, we investigated whether protein-bound polysaccharides (PBPs) derived from the fungus Coriolus versicolor (CV) induce the necroptotic death pathway in breast cancer and melanoma cells. METHODS: MCF-7 and SKMel-188 cells were exposed to PBPs either alone or in combination with necrostatin-1 (Nec-1), GSK'872 or necrosulfonamide (NSA), pharmacological inhibitors of the kinases receptor-interacting protein 1 kinase (RIPK1), receptor interacting protein kinase 3 (RIPK3) and mixed lineage kinase domain-like protein (MLKL), respectively, which are involved in necroptotic processes. The effects of cellular treatment with these inhibitors were quantified by measuring cell viability and reactive oxygen species (ROS) generation via 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) and 2',7'-dichlorofluorescein diacetate (DCF-DA) assays, respectively. The morphological changes induced in these cells were detected using holotomographic (HT) microscopy. Activation of the TNF-α/TNFR1 pathway in the PBP-stimulated cells was evaluated using TNF-α-neutralizing antibody, qRT-PCR and immunofluorescence-based assays. RESULTS: PBPs showed effective antitumor activity against MCF-7 and SKMel-188 cells. Cotreatment of the cells with Nec-1, GSK'872 or NSA abrogated PBP-induced cell death, and the cells were protected against membrane rupture. Moreover, breast cancer cell death caused by PBPs was mediated by induced activation of the TNF-α/TNFR1 pathway. Interestingly, the melanoma cells did not express TNF-α or TNFR1 after PBP stimulation; instead, PBPs triggered intracellular ROS generation, which was partially diminished by the inhibitors Nec-1, GSK'872 and NSA. CONCLUSION: These results suggest that PBPs from the fungus CV induce RIPK1/RIPK3/MLKL-mediated necroptosis in breast cancer and melanoma cells, providing novel insights into the molecular effects of PBPs on cancer cells.

Fractionation-Dependent Radiosensitization by Molecular Targeting of Nek1.

NIMA (never-in-mitosis gene A)-related kinase 1 (Nek1) is shown to impact on different cellular pathways such as DNA repair, checkpoint activation, and apoptosis. Its role as a molecular target for radiation sensitization of malignant cells, however, remains elusive. Stably transduced doxycycline (Dox)-inducible Nek1 shRNA HeLa cervix and siRNA-transfected HCT-15 colorectal carcinoma cells were irradiated in vitro and 3D clonogenic radiation survival, residual DNA damage, cell cycle distribution, and apoptosis were analyzed. Nek1 knockdown (KD) sensitized both cell lines to ionizing radiation following a single dose irradiation and more pronounced in combination with a 6 h fractionation (3 × 2 Gy) regime. For preclinical analyses we focused on cervical cancer. Nek1 shRNA HeLa cells were grafted into NOD/SCID/IL-2Rγc-/- (NSG) mice and Nek1 KD was induced by Dox-infused drinking water resulting in a significant cytostatic effect if combined with a 6 h fractionation (3 x 2 Gy) regime. In addition, we correlated Nek1 expression in biopsies of patients with cervical cancer with histopathological parameters and clinical follow-up. Our results indicate that elevated levels of Nek1 were associated with an increased rate of local or distant failure, as well as with impaired cancer-specific and overall survival in univariate analyses and for most endpoints in multivariable analyses. Finally, findings from The Cancer Genome Atlas (TCGA) validation cohort confirmed a significant association of high Nek1 expression with a reduced disease-free survival. In conclusion, we consider Nek1 to represent a novel biomarker and potential therapeutic target for drug development in the context of optimized fractionation intervals.

Expression of the NEK family in normal and cancer tissue: an immunohistochemical study.

BACKGROUND: The NEK serine/threonine protein kinases are involved in cell cycle checkpoints, DNA damage repair, and apoptosis. Alterations in these pathways are frequently associated with cell malignant cellular transformations. Thyroid cancer is the most common malignant tumour in the endocrine system. Despite good treatment methods, the number of cases has increased significantly in recent years. Here, we studied the expression of NEK1, NEK2, NEK3, and NEK5 in different types of normal and malignant tissues, using tissue microarray analysis, and identified NEKs as potential markers in thyroid malignancy. METHODS: The studied cases comprised multiple cancer tissue microarrays, including breast, colon, esophagus, kidney, lung, pancreas, prostate, stomach, thyroid and uterine cervix, as well as 281 patients who underwent thyroid resection for thyroid cancer or thyroid nodules. The expression of NEK1, NEK2, NEK3, and NEK5 was analyzed by immunohistochemistry. The expression pattern was evaluated in terms of intensity by two methods, semiquantitative and quantitative, and was compared between normal and cancer tissue. RESULTS: We analysed the expression of each member of the NEK family in a tissue-dependent manner. Compared to normal tissue, most of the evaluated proteins showed lower expression in lung tumour. However, in the thyroid, the expression was higher in malignant tissue, especially for NEK 1, NEK3 and NEK5. Concerning characteristics of the thyroid tumour, such as aggressiveness, NEK1 expression was higher in tumours with multifocality and in patients with lymph node metastasis. NEK3 expression was stronger in patients with stage II, that involved metastasis. NEK5, on the other hand, showed high expression in patients with invasion and metastasis and in patients with tumour size > 4 cm. Furthermore, this work, demonstrated for the first time a high specificity and sensitivity of over-expression of NEK1 in classical and follicular variants of papillary thyroid cancer and NEK3 in tall-cell papillary thyroid cancer. CONCLUSION: Taken together, the NEK protein kinases emerge as important proteins in thyroid cancer development and may help to identify malignancy and aggressiveness features during diagnosis. TRIAL REGISTRATION: This study was retrospectively registered.  www.accamargo.org.br/cientistas-pesquisadores/comite-de-etica-em-pequisa-cep.

The TLK1/Nek1 axis contributes to mitochondrial integrity and apoptosis prevention via phosphorylation of VDAC1.

The TLK1/Nek1 axis contributes to cell cycle arrest and implementation of the DDR to mediate survival upon DNA damage. However, when the damage is too severe, the cells typically are forced into apoptosis, and the contribution of TLKs in this process has not been investigated. In contrast, it is known that Nek1 may play a role by phosphorylating VDAC1 maintaining proper opening and closure of the channel and thus mitochondrial integrity. We now show that the activating phosphorylation of Nek1-T141 by TLK1 contributes to the phosphorylation and stability of VDAC1 and thereby to mitochondrial permeability and integrity. Treatment of three different cell lines model that overexpress Nek1-T141A mutant with doxorubicin showed exquisite sensitivity to the drug, with implementation of rapid accumulation of cells with subG1 DNA content (apoptotic) and other alterations in the cell cycle. In addition, these cells displayed reduced oxygen consumption under normal conditions and less reliance on mitochondria and more dependence on glycolysis for energy production. Consistent with greater apoptosis, upon treatment with low doses of doxorubicin, cells overexpressing Nek1-T141A displayed leakage of Cyt-C into the cytoplasmic fraction. This suggests that inhibiting the TLK1/Nek1/VDAC1 nexus could sensitize cancer cells to apoptotic killing in combination with an appropriate DNA damaging agent. We in fact have previously reported that Nek1 expression is elevated in advanced Prostate Cancer (PCa) and we now report that VDAC1 expression is elevated and correlated with disease stage, thereby making the TLK1/Nek1/VDAC1 nexus a very attractive target for PCa.

A new approach for rare variation collapsing on functional protein domains implicates specific genic regions in ALS.

Large-scale sequencing efforts in amyotrophic lateral sclerosis (ALS) have implicated novel genes using gene-based collapsing methods. However, pathogenic mutations may be concentrated in specific genic regions. To address this, we developed two collapsing strategies: One focuses rare variation collapsing on homology-based protein domains as the unit for collapsing, and the other is a gene-level approach that, unlike standard methods, leverages existing evidence of purifying selection against missense variation on said domains. The application of these two collapsing methods to 3093 ALS cases and 8186 controls of European ancestry, and also 3239 cases and 11,808 controls of diversified populations, pinpoints risk regions of ALS genes, including SOD1, NEK1, TARDBP, and FUS While not clearly implicating novel ALS genes, the new analyses not only pinpoint risk regions in known genes but also highlight candidate genes as well.

The TLK1-Nek1 axis promotes prostate cancer progression.

We recently uncovered the critical TLK1>NEK1>ATR > Chk1 axis in mediating the DDR and cell cycle checkpoint while transiting from Androgen Sensitive to Insensitive growth for LNCaP and TRAMP-C2 cells. However, we did not know the generality of this pathway in PCa progression since there are few cell lines where the transition has been studied. Furthermore, the identification of Nek1, and more importantly the TLK-mediated phosphorylation of T141, has never been studied in PCa biopsies. We now report the first study of a PCa TMA of p-Nek1-T141 and correlation to the Gleason score. In addition we found that TRAMP mice treated with the TLK inhibitor, thioridazine (THD), following castration did not recover cancerous growth of their prostates. Moreover, we recapitulated the process of translational increase in TLK1B expression in a naïve PDX model that was established from an AR + adenocarcinoma. Therefore, we believe that this TLK1-Nek1 mediated DDR axis is likely to be a common adaptive response during the transition of PCa cells toward androgen-insensitive growth, and hence CRPC progression, which has the potential to be targeted with THD and other TLK or Nek1 inhibitors.

TgCep250 is dynamically processed through the division cycle and is essential for structural integrity of the Toxoplasma centrosome.

The apicomplexan centrosome has a unique bipartite structure comprising an inner and outer core responsible for the nuclear cycle (mitosis) and budding cycles (cytokinesis), respectively. Although these two cores are always associated, they function independently to facilitate polyploid intermediates in the production of many progeny per replication round. Here, we describe the function of a large coiled-coil protein in Toxoplasma gondii, TgCep250, in connecting the two centrosomal cores and promoting their structural integrity. Throughout the cell cycle, TgCep250 localizes to the inner core but, associated with proteolytic processing, is also present on the outer core during the onset of cell division. In the absence of TgCep250, stray centrosome inner and outer core foci were observed. The detachment between centrosomal inner and outer cores was found in only one of the centrosomes during cell division, indicating distinct states of mother and daughter centrosomes. In mammals, Cep250 processing is required for centrosomal splitting and is mediated by Nek phopsphorylation. However, we show that neither the nonoverlapping spatiotemporal localization of TgNek1 and TgCep250 nor the distinct phenotypes upon their respective depletion support conservation of this mechanism in Toxoplasma. In conclusion, TgCep250 has a tethering function tailored to the unique bipartite centrosome in the Apicomplexa.

Targeting the TLK1/NEK1 DDR axis with Thioridazine suppresses outgrowth of androgen independent prostate tumors.

Standard therapy for advanced Prostate Cancer (PCa) consists of antiandrogens, which provide respite from disease progression, but ultimately fail resulting in the incurable phase of the disease: mCRPC. Targeting PCa cells before their progression to mCRPC would greatly improve the outcome. Combination therapy targeting the DNA Damage Response (DDR) has been limited by general toxicity, and a goal of clinical trials is how to target the DDR more specifically. We now show that androgen deprivation therapy (ADT) of LNCaP cells results in increased expression of TLK1B, a key kinase upstream of NEK1 and ATR and mediating the DDR that typically results in a temporary cell cycle arrest of androgen responsive PCa cells. Following DNA damage, addition of the TLK specific inhibitor, thioridazine (THD), impairs ATR and Chk1 activation, establishing the existence of a ADT > TLK1 > NEK1 > ATR > Chk1, DDR pathway, while its abrogation leads to apoptosis. Treatment with THD suppressed the outgrowth of androgen-independent (AI) colonies of LNCaP and TRAMP-C2 cells cultured with bicalutamide. Moreover, THD significantly inhibited the growth of several PCa cells in vitro (including AI lines). Administration of THD or bicalutamide was not effective at inhibiting long-term tumor growth of LNCaP xenografts. In contrast, combination therapy remarkably inhibited tumor growth via bypass of the DDR. Moreover, xenografts of LNCaP cells overexpressing a NEK1-T141A mutant were durably suppressed with bicalutamide. Collectively, these results suggest that targeting the TLK1/NEK1 axis might be a novel therapy for PCa in combination with standard of care (ADT).

VHL regulates NEK1 via both HIF-2α pathway and ubiquitin-proteasome pathway in renal cancer cell.

Both Von Hippel-Lindau tumor suppressor (VHL) and Never-in-mitosis A (NIMA)-related kinase 1 (NEK1) are involved in primary cilium formation, but whether VHL could regulate NEK1 is unknown. Here, we demonstrated that renal cancer cells Caki-1 and ACHN with wild-type VHL expressed lower level of NEK1 than that of VHL-defective cells including 786-O, 769-P and A498 cells. VHL-overexpression down-regulated NEK1 in 769-P cells, while VHL-knockdown up-regulated NEK1 in Caki-1 cells. In addition, we found the hypoxia response element (HRE) in the promoter sequence of NEK1 and hypoxia induced NEK1 expression both in vitro and in vivo. HIF-2α knockdown blocked hypoxia induced NEK1 upregulation instead of HIF-1α, which indicates that NEK1 may be a new target of HIF-2α. Moreover, we confirmed the association between VHL and NEK1 in Caki-1 cell, then showed VHL promoted NEK1 protein degradation and ubiquitination. In conclusion, our findings showed VHL regulates NEK1 via both HIF-2α pathway and ubiquitin-proteasome pathway in renal cancer cells.

NEK1 loss-of-function mutation induces DNA damage accumulation in ALS patient-derived motoneurons.

Mutations in genes coding for proteins involved in DNA damage response (DDR) and repair, such as C9orf72 and FUS (Fused in Sarcoma), are associated with neurodegenerative diseases and lead to amyotrophic lateral sclerosis (ALS). Heterozygous loss-of-function mutations in NEK1 (NIMA-related kinase 1) have also been recently found to cause ALS. NEK1 codes for a multifunctional protein, crucially involved in mitotic checkpoint control and DDR. To resolve pathological alterations associated with NEK1 mutation, we compared hiPSC-derived motoneurons carrying a NEK1 mutation with mutant C9orf72 and wild type neurons at basal level and after DNA damage induction. Motoneurons carrying a C9orf72 mutation exhibited cell specific signs of increased DNA damage. This phenotype was even more severe in NEK1c.2434A>T neurons that showed significantly increased DNA damage at basal level and impaired DDR after induction of DNA damage in an maturation-dependent manner. Our results provide first mechanistic insight in pathophysiological alterations induced by NEK1 mutations and point to a converging pathomechanism of different gene mutations causative for ALS. Therefore, our study contributes to the development of novel therapeutic strategies to reduce DNA damage accumulation in neurodegenerative diseases and ALS.

NIMA-related kinase 1 (NEK1) regulates meiosis I spindle assembly by altering the balance between α-Adducin and Myosin X.

NIMA-related kinase 1 (NEK1) is a serine/threonine and tyrosine kinase that is highly expressed in mammalian germ cells. Mutations in Nek1 induce anemia, polycystic kidney and infertility. In this study we evaluated the role of NEK1 in meiotic spindle formation in both male and female gametes. Our results show that the lack of NEK1 provokes an abnormal organization of the meiosis I spindle characterized by elongated and/or multipolar spindles, and abnormal chromosome congression. The aberrant spindle structure is concomitant with the disruption in localization and protein levels of myosin X (MYO10) and α-adducin (ADD1), both of which are implicated in the regulation of spindle formation during mitosis. Interaction of ADD1 with MYO10 is dependent on phosphorylation, whereby phosphorylation of ADD1 enables its binding to MYO10 on mitotic spindles. Reduction in ADD1 protein in NEK1 mutant mice is associated with hyperphosphorylation of ADD1, thereby preventing the interaction with MYO10 during meiotic spindle formation. Our results reveal a novel regulatory role for NEK1 in the regulation of spindle architecture and function during meiosis.