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Spartan deficiency causes genomic instability and progeroid phenotypes.

Maskey RS, Kim MS, Baker DJ, Childs B, Malureanu LA, Jeganathan KB, Machida Y, van Deursen JM, Machida YJ - Nat Commun (2014)

Bottom Line: However, the physiological relevance of Spartan has not been established.Cre-mediated depletion of Spartan from conditional knockout mouse embryonic fibroblasts results in impaired lesion bypass, incomplete DNA replication, formation of micronuclei and chromatin bridges and eventually cell death.These data demonstrate that Spartan plays a key role in maintaining structural and numerical chromosome integrity and suggest a link between Spartan insufficiency and progeria.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, 200 First Street SW, Rochester, Minnesota 55905, USA.

ABSTRACT
Spartan (also known as DVC1 and C1orf124) is a PCNA-interacting protein implicated in translesion synthesis, a DNA damage tolerance process that allows the DNA replication machinery to replicate past nucleotide lesions. However, the physiological relevance of Spartan has not been established. Here we report that Spartan insufficiency in mice causes chromosomal instability, cellular senescence and early onset of age-related phenotypes. Whereas complete loss of Spartan causes early embryonic lethality, hypomorphic mice with low amounts of Spartan are viable. These mice are growth retarded and develop cataracts, lordokyphosis and cachexia at a young age. Cre-mediated depletion of Spartan from conditional knockout mouse embryonic fibroblasts results in impaired lesion bypass, incomplete DNA replication, formation of micronuclei and chromatin bridges and eventually cell death. These data demonstrate that Spartan plays a key role in maintaining structural and numerical chromosome integrity and suggest a link between Spartan insufficiency and progeria.

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Sprtn KO causes embryonic lethality.(a) Schematic of the mouse Sprtn gene and the targeted alleles. An inverted Neo cassette was inserted in the second intron with flanking FLP recognition target (FRT) sequences. LoxP sites were also inserted at the indicated positions. The floxed and KO alleles were created by crossing heterozygote mice with FLP and Cre-transgenic mice, respectively. Positions of genotyping primers are indicated by arrows. (b) PCR-based genotyping (at weaning) of wild-type and Sprtn heterozygote mice produced by intercrossing Sprtn+/−. (c) PCR-based genotyping of wild-type, heterozygote and KO blastocysts. (d) Blastocysts from Sprtn+/− intercrosses were cultured in vitro and observed by phase-contrast microscopy on 6 consecutive days. Representative images of Sprtn+/+, Sprtn+/− and Sprtn−/− blastocysts are shown. Scale bar, 100 μm.
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f1: Sprtn KO causes embryonic lethality.(a) Schematic of the mouse Sprtn gene and the targeted alleles. An inverted Neo cassette was inserted in the second intron with flanking FLP recognition target (FRT) sequences. LoxP sites were also inserted at the indicated positions. The floxed and KO alleles were created by crossing heterozygote mice with FLP and Cre-transgenic mice, respectively. Positions of genotyping primers are indicated by arrows. (b) PCR-based genotyping (at weaning) of wild-type and Sprtn heterozygote mice produced by intercrossing Sprtn+/−. (c) PCR-based genotyping of wild-type, heterozygote and KO blastocysts. (d) Blastocysts from Sprtn+/− intercrosses were cultured in vitro and observed by phase-contrast microscopy on 6 consecutive days. Representative images of Sprtn+/+, Sprtn+/− and Sprtn−/− blastocysts are shown. Scale bar, 100 μm.

Mentions: To explore the physiological relevance of Spartan, we generated a series of mice with graded reduction in Spartan expression using hypomorphic (H) and knockout (KO) Sprtn alleles that we generated by gene targeting (Fig. 1a,b). Intercrossing of Sprtn+/− mice failed to produce Sprtn−/−-live offspring and no Sprtn−/− embryos were found from embryonic day (E) 7.5 to E13.5 (Table 1), suggesting that Sprtn KO in mice causes early embryonic lethality. To examine the effect of Sprtn KO on early embryogenesis, we isolated blastocysts at E3.5 and cultured them in vitro for 6 days. Genotyping was successful for all the blastocysts collected, and Sprtn−/− blastocysts were obtained at normal Mendelian frequency (Fig. 1c; Table 2). However, Sprtn−/− blastocysts failed to hatch and enlarge the inner cell mass when cultured in vitro (Fig. 1d; Table 2), indicating that death occurred prior at the implantation stage.


Spartan deficiency causes genomic instability and progeroid phenotypes.

Maskey RS, Kim MS, Baker DJ, Childs B, Malureanu LA, Jeganathan KB, Machida Y, van Deursen JM, Machida YJ - Nat Commun (2014)

Sprtn KO causes embryonic lethality.(a) Schematic of the mouse Sprtn gene and the targeted alleles. An inverted Neo cassette was inserted in the second intron with flanking FLP recognition target (FRT) sequences. LoxP sites were also inserted at the indicated positions. The floxed and KO alleles were created by crossing heterozygote mice with FLP and Cre-transgenic mice, respectively. Positions of genotyping primers are indicated by arrows. (b) PCR-based genotyping (at weaning) of wild-type and Sprtn heterozygote mice produced by intercrossing Sprtn+/−. (c) PCR-based genotyping of wild-type, heterozygote and KO blastocysts. (d) Blastocysts from Sprtn+/− intercrosses were cultured in vitro and observed by phase-contrast microscopy on 6 consecutive days. Representative images of Sprtn+/+, Sprtn+/− and Sprtn−/− blastocysts are shown. Scale bar, 100 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4269170&req=5

f1: Sprtn KO causes embryonic lethality.(a) Schematic of the mouse Sprtn gene and the targeted alleles. An inverted Neo cassette was inserted in the second intron with flanking FLP recognition target (FRT) sequences. LoxP sites were also inserted at the indicated positions. The floxed and KO alleles were created by crossing heterozygote mice with FLP and Cre-transgenic mice, respectively. Positions of genotyping primers are indicated by arrows. (b) PCR-based genotyping (at weaning) of wild-type and Sprtn heterozygote mice produced by intercrossing Sprtn+/−. (c) PCR-based genotyping of wild-type, heterozygote and KO blastocysts. (d) Blastocysts from Sprtn+/− intercrosses were cultured in vitro and observed by phase-contrast microscopy on 6 consecutive days. Representative images of Sprtn+/+, Sprtn+/− and Sprtn−/− blastocysts are shown. Scale bar, 100 μm.
Mentions: To explore the physiological relevance of Spartan, we generated a series of mice with graded reduction in Spartan expression using hypomorphic (H) and knockout (KO) Sprtn alleles that we generated by gene targeting (Fig. 1a,b). Intercrossing of Sprtn+/− mice failed to produce Sprtn−/−-live offspring and no Sprtn−/− embryos were found from embryonic day (E) 7.5 to E13.5 (Table 1), suggesting that Sprtn KO in mice causes early embryonic lethality. To examine the effect of Sprtn KO on early embryogenesis, we isolated blastocysts at E3.5 and cultured them in vitro for 6 days. Genotyping was successful for all the blastocysts collected, and Sprtn−/− blastocysts were obtained at normal Mendelian frequency (Fig. 1c; Table 2). However, Sprtn−/− blastocysts failed to hatch and enlarge the inner cell mass when cultured in vitro (Fig. 1d; Table 2), indicating that death occurred prior at the implantation stage.

Bottom Line: However, the physiological relevance of Spartan has not been established.Cre-mediated depletion of Spartan from conditional knockout mouse embryonic fibroblasts results in impaired lesion bypass, incomplete DNA replication, formation of micronuclei and chromatin bridges and eventually cell death.These data demonstrate that Spartan plays a key role in maintaining structural and numerical chromosome integrity and suggest a link between Spartan insufficiency and progeria.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, 200 First Street SW, Rochester, Minnesota 55905, USA.

ABSTRACT
Spartan (also known as DVC1 and C1orf124) is a PCNA-interacting protein implicated in translesion synthesis, a DNA damage tolerance process that allows the DNA replication machinery to replicate past nucleotide lesions. However, the physiological relevance of Spartan has not been established. Here we report that Spartan insufficiency in mice causes chromosomal instability, cellular senescence and early onset of age-related phenotypes. Whereas complete loss of Spartan causes early embryonic lethality, hypomorphic mice with low amounts of Spartan are viable. These mice are growth retarded and develop cataracts, lordokyphosis and cachexia at a young age. Cre-mediated depletion of Spartan from conditional knockout mouse embryonic fibroblasts results in impaired lesion bypass, incomplete DNA replication, formation of micronuclei and chromatin bridges and eventually cell death. These data demonstrate that Spartan plays a key role in maintaining structural and numerical chromosome integrity and suggest a link between Spartan insufficiency and progeria.

Show MeSH
Related in: MedlinePlus