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Mutation of NIMA-related kinase 1 (NEK1) leads to chromosome instability.

Chen Y, Chen CF, Chiang HC, Pena M, Polci R, Wei RL, Edwards RA, Hansel DE, Chen PL, Riley DJ - Mol. Cancer (2011)

Bottom Line: These NEK1-deficient cells transform, acquire the ability to grow in anchorage-independent conditions, and form tumors when injected into syngeneic mice.Genomic instability is also manifest in NEK1 +/- mice, which late in life develop lymphomas with a much higher incidence than wild type littermates.NEK1 is required for the maintenance of genome stability by acting at multiple junctures, including control of chromosome stability.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Medicine, Division of Endocrinology, University of California at Irvine, 1130 Gross Hall, Irvine, CA 92697, USA. yumayc@uci.edu

ABSTRACT

Background: NEK1, the first mammalian ortholog of the fungal protein kinase never-in-mitosis A (NIMA), is involved early in the DNA damage sensing/repair pathway. A defect in DNA repair in NEK1-deficient cells is suggested by persistence of DNA double strand breaks after low dose ionizing radiation (IR). NEK1-deficient cells also fail to activate the checkpoint kinases CHK1 and CHK2, and fail to arrest properly at G1/S or G2/M-phase checkpoints after DNA damage.

Results: We show here that NEK1-deficient cells suffer major errors in mitotic chromosome segregation and cytokinesis, and become aneuploid. These NEK1-deficient cells transform, acquire the ability to grow in anchorage-independent conditions, and form tumors when injected into syngeneic mice. Genomic instability is also manifest in NEK1 +/- mice, which late in life develop lymphomas with a much higher incidence than wild type littermates.

Conclusion: NEK1 is required for the maintenance of genome stability by acting at multiple junctures, including control of chromosome stability.

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Related in: MedlinePlus

Cultured NEK1 -/- cells form colonies in soft agar and tumors in when injected into mice. A, B. Equal numbers of unclumped cells (high density (1 × 105/60 mm); low density, (2 × 104/60 mm) of the indicated passage were seeded in duplicate in 0.367% agar. Total colony numbers were scored after 21 days. Contiguous clusters of >50 cells were considered colonies. Representative colonies were photographed as shown in B. Bar = 100 μm. C. H&E stained sections of tumors, 21 days after subcutaneous injection of NEK1 -/- RTEs into kat2J mice. Cy = cyst, arrows = abnormal nuclei. Bars = 100 μm. D. p19ARF expression in two distinct NEK1 -/- cell lines at different passages in culture, and in wild type (NEK1 +/+) cells at passage 7. Alpha tubulin expression served to control for protein loading in each lane.
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Figure 5: Cultured NEK1 -/- cells form colonies in soft agar and tumors in when injected into mice. A, B. Equal numbers of unclumped cells (high density (1 × 105/60 mm); low density, (2 × 104/60 mm) of the indicated passage were seeded in duplicate in 0.367% agar. Total colony numbers were scored after 21 days. Contiguous clusters of >50 cells were considered colonies. Representative colonies were photographed as shown in B. Bar = 100 μm. C. H&E stained sections of tumors, 21 days after subcutaneous injection of NEK1 -/- RTEs into kat2J mice. Cy = cyst, arrows = abnormal nuclei. Bars = 100 μm. D. p19ARF expression in two distinct NEK1 -/- cell lines at different passages in culture, and in wild type (NEK1 +/+) cells at passage 7. Alpha tubulin expression served to control for protein loading in each lane.

Mentions: The phenotypes we observed in NEK1 -/- cells prompted us to look for neoplastic behavior. A good predictor of whether cancer cells can form tumors in vivo is their ability to grow in soft agar without attachment to plastic. We therefore employed the soft agar assay to look for anchorage-independent colony formation in NEK1 -/- and wild type RTEs derived from sex-matched, littermate mice and cultured in identical conditions. At different passages, well-separated cells were seeded into soft agar and examined for their ability to form colonies. As early as passage five, small numbers of NEK1 -/- cells were observed to form colonies. At higher passages, NEK1 -/- cells were even more efficient in forming colonies. In contrast, wild type cells grew poorly and failed to proliferate beyond the single cell stage in the soft agar, even after many passages in monolayer culture (Figure 5A). NEK1 inactivation therefore seems to lead to a transformed phenotype in clones of cultured cells.


Mutation of NIMA-related kinase 1 (NEK1) leads to chromosome instability.

Chen Y, Chen CF, Chiang HC, Pena M, Polci R, Wei RL, Edwards RA, Hansel DE, Chen PL, Riley DJ - Mol. Cancer (2011)

Cultured NEK1 -/- cells form colonies in soft agar and tumors in when injected into mice. A, B. Equal numbers of unclumped cells (high density (1 × 105/60 mm); low density, (2 × 104/60 mm) of the indicated passage were seeded in duplicate in 0.367% agar. Total colony numbers were scored after 21 days. Contiguous clusters of >50 cells were considered colonies. Representative colonies were photographed as shown in B. Bar = 100 μm. C. H&E stained sections of tumors, 21 days after subcutaneous injection of NEK1 -/- RTEs into kat2J mice. Cy = cyst, arrows = abnormal nuclei. Bars = 100 μm. D. p19ARF expression in two distinct NEK1 -/- cell lines at different passages in culture, and in wild type (NEK1 +/+) cells at passage 7. Alpha tubulin expression served to control for protein loading in each lane.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3025975&req=5

Figure 5: Cultured NEK1 -/- cells form colonies in soft agar and tumors in when injected into mice. A, B. Equal numbers of unclumped cells (high density (1 × 105/60 mm); low density, (2 × 104/60 mm) of the indicated passage were seeded in duplicate in 0.367% agar. Total colony numbers were scored after 21 days. Contiguous clusters of >50 cells were considered colonies. Representative colonies were photographed as shown in B. Bar = 100 μm. C. H&E stained sections of tumors, 21 days after subcutaneous injection of NEK1 -/- RTEs into kat2J mice. Cy = cyst, arrows = abnormal nuclei. Bars = 100 μm. D. p19ARF expression in two distinct NEK1 -/- cell lines at different passages in culture, and in wild type (NEK1 +/+) cells at passage 7. Alpha tubulin expression served to control for protein loading in each lane.
Mentions: The phenotypes we observed in NEK1 -/- cells prompted us to look for neoplastic behavior. A good predictor of whether cancer cells can form tumors in vivo is their ability to grow in soft agar without attachment to plastic. We therefore employed the soft agar assay to look for anchorage-independent colony formation in NEK1 -/- and wild type RTEs derived from sex-matched, littermate mice and cultured in identical conditions. At different passages, well-separated cells were seeded into soft agar and examined for their ability to form colonies. As early as passage five, small numbers of NEK1 -/- cells were observed to form colonies. At higher passages, NEK1 -/- cells were even more efficient in forming colonies. In contrast, wild type cells grew poorly and failed to proliferate beyond the single cell stage in the soft agar, even after many passages in monolayer culture (Figure 5A). NEK1 inactivation therefore seems to lead to a transformed phenotype in clones of cultured cells.

Bottom Line: These NEK1-deficient cells transform, acquire the ability to grow in anchorage-independent conditions, and form tumors when injected into syngeneic mice.Genomic instability is also manifest in NEK1 +/- mice, which late in life develop lymphomas with a much higher incidence than wild type littermates.NEK1 is required for the maintenance of genome stability by acting at multiple junctures, including control of chromosome stability.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Medicine, Division of Endocrinology, University of California at Irvine, 1130 Gross Hall, Irvine, CA 92697, USA. yumayc@uci.edu

ABSTRACT

Background: NEK1, the first mammalian ortholog of the fungal protein kinase never-in-mitosis A (NIMA), is involved early in the DNA damage sensing/repair pathway. A defect in DNA repair in NEK1-deficient cells is suggested by persistence of DNA double strand breaks after low dose ionizing radiation (IR). NEK1-deficient cells also fail to activate the checkpoint kinases CHK1 and CHK2, and fail to arrest properly at G1/S or G2/M-phase checkpoints after DNA damage.

Results: We show here that NEK1-deficient cells suffer major errors in mitotic chromosome segregation and cytokinesis, and become aneuploid. These NEK1-deficient cells transform, acquire the ability to grow in anchorage-independent conditions, and form tumors when injected into syngeneic mice. Genomic instability is also manifest in NEK1 +/- mice, which late in life develop lymphomas with a much higher incidence than wild type littermates.

Conclusion: NEK1 is required for the maintenance of genome stability by acting at multiple junctures, including control of chromosome stability.

Show MeSH
Related in: MedlinePlus