Incidental Mutation 'P0005:Nek6'

• ID: 7632
• Institutional Source: Beutler Lab
• Gene Symbol: Nek6
• Ensembl Gene: ENSMUSG00000026749
• Gene Name: NIMA (never in mitosis gene a)-related expressed kinase 6
• Synonyms: 1300007C09Rik
• MMRRC Submission: 038262-MU
• Essential gene?: Probably non essential (E-score: 0.118)
• Stock #: P0005 (G1)
• Status: Validated
• Chromosome: 2
• Chromosomal Location: 38401655-38484618 bp(+) (GRCm39)
• Type of Mutation: critical splice donor site (2 bp from exon)
• DNA Base Change (assembly): T to C at 38459749 bp (GRCm39)
• Zygosity: Heterozygous
• Ref Sequence: ENSEMBL: ENSMUSP00000108523
• Gene Model: predicted gene model for transcript(s): [ENSMUST00000054234] [ENSMUST00000112895] [ENSMUST00000112902]
• AlphaFold: Q9ES70
• Predicted Effect: probably null; Transcript: ENSMUST00000054234
• SMART Domains: Protein: ENSMUSP00000049723; Gene: ENSMUSG00000026749

Domain	Start	End	E-Value	Type
S_TKc	45	310	3.01e-91	SMART

• Predicted Effect: probably null; Transcript: ENSMUST00000112895
• SMART Domains: Protein: ENSMUSP00000108516; Gene: ENSMUSG00000026749

Domain	Start	End	E-Value	Type
S_TKc	45	310	3.01e-91	SMART

• Predicted Effect: probably null; Transcript: ENSMUST00000112902
• SMART Domains: Protein: ENSMUSP00000108523; Gene: ENSMUSG00000026749

Domain	Start	End	E-Value	Type
S_TKc	34	299	3.01e-91	SMART

• Predicted Effect: noncoding transcript; Transcript: ENSMUST00000203964
• Meta Mutation Damage Score: 0.9500
• Coding Region Coverage:
  • 1x: 85.5%
  • 3x: 80.5%
  • 10x: 66.1%
  • 20x: 49.6%
• Validation Efficiency: 95% (104/109)
• MGI Phenotype: FUNCTION: [Summary is not available for the mouse gene. This summary is for the human ortholog.] The protein encoded by this gene is a kinase required for progression through the metaphase portion of mitosis. Inhibition of the encoded protein can lead to apoptosis. This protein also can enhance tumorigenesis by suppressing tumor cell senescence. Several transcript variants encoding a few different isoforms have been found for this gene. [provided by RefSeq, Oct 2011] ; PHENOTYPE: No significant homozygous or heterozygous mutant phenotype was detected in a high throughput screen of a targeted mutation, however these homozygotes exhibit an increased response of the heart to induced stress, with aggravated cardiac hypertrophy. [provided by MGI curators]
• Allele List at MGI:

  All alleles(128) : Targeted(3) Gene trapped(125)

• Posted On: 2012-10-05
• Science Writer: Anne Murray

Protein Function and Prediction

The Aspergillus nidulans 'never in mitosis A' (NIMA) gene encodes a serine/threonine kinase that controls initiation of mitosis. NIMA-related kinases (NEKs) are a group of protein kinases that are homologous to NIMA. Evidence suggests that NEKs perform functions similar to those of NIMA (OMIM *604884). Nek6 is proposed to regulate p70 ribosomal S6 kinase activity, indicating that Nek6 is involved in growth signal transduction pathways (1).  siRNA of Nek6 results in cell arrest in M phase and triggers apoptosis (2).  Furthermore, examination of cell cycle progress after treatment with a kinase-inactive mutant of Nek6 showed mitotic arrest at metaphase before the cells progressed to apoptosis, indicating that Nek6 is required for the metaphase-anaphase transition (2).  Later studies showed that Nek6-induced cell cycle arrest upon DNA damage is a result of DNA damage-induced phosphorylation (3).  Nek6 is upregulated in various human cancers and Nek6 expression is decreased in both replicative senescence of normal fibroblasts as well as p53-induced premature senescence, indicating that Nek6 may have a role in tumorigenesis (4).

Expression/Localization

During early embryogenesis, Nek6 is expressed in primary giant trophoblast cells (5).  In later embryogenesis, expression of Nek6 is restricted to the nervous system (5). In the adult brain, Nek6 is expressed primarily in the piriform cortex, the archicortex. Lower levels were observed in the amygdala, olfactory bulb, and the hindbrain (5).  In addition, Nek6 was found to be expressed in the intestine, ovary, and the granulosa cells of large growing and antral follicles (5).

Genotyping

P0005:Nek6 (NEK6 ICSI) genotyping is performed by amplifying the region containing the mutation using PCR, followed by sequencing of the amplified region to detect the single nucleotide transition.
 

PCR Primers

NEK6 ICSI(F): 5’- GTTGCTTTCACCTGTGTGAGACCC-3’

NEK6 ICSI(R): 5’- AAGCCAGGCAGATGCAGTCCATTC-3’

Sequencing Primer

NEK6 ICSI_seq(F): 5’- TGTTACAGATGAATAAATGAGCC-3’

NEK6 ICSI_seq(R): 5’- GCAGTCCATTCTATATGACAGGGTC-3’
 

PCR program

1) 94°C             2:00

2) 94°C             0:30

3) 55°C             0:30

4) 72°C             1:00

5) repeat steps (2-4) 40X

6) 72°C             10:00

7) 4°C               ∞

The following sequence of 667 nucleotides (from GenBank genomic region NC_000068 encoding Nek6) is amplified:

57659                                                                gt

57661 tgctttcacc tgtgtgagac cctgccctag tcactgcagt gcaaagctta ctgactgtca

57721 gacagtgtct cccgtgcggt gacgctcgtc tggcacaggg gaggcagctg tacaaaagga

57781 gacagtagta tagaagtctc tgctccccac tcacagccaa cctgtgacat gggggagggg

57841 gatagtcatg ttacagatga agtaaatgag cctcctccca tcccttctaa gcctgctctc

57901 cccacaccaa ccctgtgtcc ttgccctgca gacatcaagc ccgccaacgt gttcatcaca

57961 gctacgggca ttgtgaagct tggtgacctc ggcctgggcc gcttcttcag ctcggagacc

58021 actgcggccc actcactagg taagaggcct atgtcccacc tacccgcagc ccctgtaggc

58081 ctgggtgacc cagccggaag ggcatccatt ctcctggaac tatttgtagt gtgtgggaag

58141 atcttttttg agtcttaaga gttcaccctt gctttgggat agatccaaag tagaggaaac

58201 tgaggcccag aggggccact gcagggggga agggggatct ccccatatcc ccaagtcatc

58261 tctctagtac ttcctggaag gccaacagac cctgtcatat agaatggact gcatctgcct

58321 ggctt

Primer binding sites are underlined and the sequencing primer is highlighted; the mutated nucleotide is shown in red text.

References

  1. Belham, C., Comb, M. J., and Avruch, J. (2001) Identification of the NIMA Family Kinases NEK6/7 as Regulators of the p70 Ribosomal S6 Kinase. Curr Biol. 11, 1155-1167.

  2. Yin, M. J., Shao, L., Voehringer, D., Smeal, T., and Jallal, B. (2003) The serine/threonine Kinase Nek6 is Required for Cell Cycle Progression through Mitosis. J Biol Chem. 278, 52454-52460.

  3. Lee, M. Y., Kim, H. J., Kim, M. A., Jee, H. J., Kim, A. J., Bae, Y. S., Park, J. I., Chung, J. H., and Yun, J. (2008) Nek6 is Involved in G2/M Phase Cell Cycle Arrest through DNA Damage-Induced Phosphorylation. Cell Cycle. 7, 2705-2709.

  4. Jee, H. J., Kim, A. J., Song, N., Kim, H. J., Kim, M., Koh, H., and Yun, J. (2010) Nek6 Overexpression Antagonizes p53-Induced Senescence in Human Cancer Cells. Cell Cycle. 9, 4703-4710.

  5. Feige, E., and Motro, B. (2002) The Related Murine Kinases, Nek6 and Nek7, Display Distinct Patterns of Expression. Mech Dev. 110, 219-223.