NEK8 Mutations Affect Ciliary and Centrosomal Localization and May Cause Nephronophthisis

Abstract

Nephronophthisis, an autosomal recessive kidney disease, is the most frequent genetic cause of chronic renal failure in the first 3 decades of life. Causative mutations in 8 genes (NPHP1-8) have been identified, and homologous mouse models for NPHP2/INVS and NPHP3 have been described. The jck mouse is another model of recessive cystic kidney disease, and this mouse harbors a missense mutation, G448V, in the highly conserved RCC1 domain of Nek8. We hypothesized that mutations in NEK8 might cause nephronophthisis in humans, so we performed mutational analysis in a worldwide cohort of 588 patients. We identified 3 different amino acid changes that were conserved through evolution (L330F, H425Y, and A497P) and that were absent from at least 80 ethnically matched controls. All 3 mutations were within RCC1 domains, and the mutation H425Y was positioned within the same RCC1 repeat as the mouse jck mutation. To test the functional significance of these mutations, we introduced them into full-length mouse Nek8 GFP-tagged cDNA constructs. We transiently overexpressed the constructs in inner medullary collecting duct cells (IMCD-3 cell line) and compared the subcellular localization of mutant Nek8 to wild-type Nek8. All mutant forms of Nek8 showed defects in ciliary localization to varying degrees; the H431Y mutant (human H425Y) was completely absent from cilia and the amount localized to centrosomes was decreased. Overexpression of these mutants did not affect overall ciliogenesis, mitosis, or centriole number. Our genetic and functional data support the assumption that mutations in NEK8 cause nephronophthisis (NPHP9), adding another link between proteins mutated in cystic kidney disease and their localization to cilia and centrosomes.

Loss of the Ciliary Kinase Nek8 Causes Left-Right Asymmetry Defects

“…Subsequent studies have reported missense mutations in the RCC domain-encoding regions of both rat Nek8 in the Lewis PKD model 16 and human NEK8, in which three independent mutations were identified from 700 NPHP patients, making it a candidate for NPHP type 9. 11,17 Together, these data demonstrate that the NEK8 C terminus is critical for its function and the protein plays an important role in the maintenance of renal tubule integrity in the postnatal kidney. Of note, all of the reported mutations of Nek8 are missense mutations.…”

Downregulating Hedgehog Signaling Reduces Renal Cystogenic Potential of Mouse Models

“…The jck mutant phenotypically resembles ADPKD 29 and carries a mutation in the Nek8 kinase. 36 Mutations in NEK8 have been identified in patients with nephronophthisis-9 37 and in the Lewis PKD rat. 38 At 7 weeks of age, jck/jck mice showed a mean (6SEM) %KW/BW of 4.960.33 (n=11) compared with 1.360.04 in wt mice (n=19).…”

Nek1 and TAZ Interact to Maintain Normal Levels of Polycystin 2

“…2 Nek kinases function in the formation of the primary cilium in many eukaryotes. [3][4][5][6][7][8] They also play roles in DNA repair 9 and apoptosis. 10 Spontaneous mutations in genes encoding two members of the Nek family give rise to autosomal recessive forms of polycystic kidney disease (PKD) in the mouse.…”

“…Nek8 nevertheless regulates the expression and localization of the polycystins 4,20 and has been linked to medullary cystic kidney disease (nephronopthisis). 8 Knockdown of Nek8 in zebrafish embryos results in cystic kidney. 16 Independent investigations have shown that knockouts of TAZ (Wwtr1) result in the development of PKD.…”