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Double-strand break repair deficiency in NONO knockout murine embryonic fibroblasts and compensation by spontaneous upregulation of the PSPC1 paralog.

Li S, Li Z, Shu FJ, Xiong H, Phillips AC, Dynan WS - Nucleic Acids Res. (2014)

Bottom Line: We tested whether NONO and PSPC1 might also affect repair indirectly by influencing mRNA levels for other DSB repair genes.Of 12 genes tested, none were downregulated, and several were upregulated.Thus, NONO or related proteins are critical for DSB repair, NONO and PSPC1 are functional homologs with partially interchangeable functions and a compensatory response involving PSPC1 blunts the effect of NONO deficiency.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiation Oncology, Emory University, Atlanta, GA 30322, USA Department of Biochemistry, Emory University, Atlanta, GA 30322, USA Institute of Molecular Medicine and Genetics, Georgia Regents University, Augusta, GA 30912, USA.

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Increased levels of PSPC1 and SFPQ–PSPC1 complex in NONO-deficient cells. (A) Immunostaining of WT-2 and gt-2 MEF isolates using anti-PSPC1 antibody. Scale bar, 10 μm. (B) Immunoblotting to determine levels of SFPQ, NONO and PSPC1 proteins in cells of indicated genotype. Two independent MEF populations, derived from different embryos, were analyzed for each type. Arrowheads denote proteins as indicated. (C) Quantification of data from panel (B). Values are normalized to β-actin. Error bars reflect standard deviation of values from independent MEF populations. (D) Immunoprecipitation (IP), followed by immunoblotting (IB), to protein–protein complexes in MEFs of indicated genotypes.
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Figure 3: Increased levels of PSPC1 and SFPQ–PSPC1 complex in NONO-deficient cells. (A) Immunostaining of WT-2 and gt-2 MEF isolates using anti-PSPC1 antibody. Scale bar, 10 μm. (B) Immunoblotting to determine levels of SFPQ, NONO and PSPC1 proteins in cells of indicated genotype. Two independent MEF populations, derived from different embryos, were analyzed for each type. Arrowheads denote proteins as indicated. (C) Quantification of data from panel (B). Values are normalized to β-actin. Error bars reflect standard deviation of values from independent MEF populations. (D) Immunoprecipitation (IP), followed by immunoblotting (IB), to protein–protein complexes in MEFs of indicated genotypes.

Mentions: We investigated PSPC1 expression by immunofluorescence and immunoblotting using anti-PSPC1 antibody (Figure 3A and B). PSPC1 was present at low levels, near the limit of detection, in wild-type MEFs (+/0). Expression was strongly increased in NONO-deficient MEFs (gt/0 and gt/gt). Thus, it appears there is a reciprocal relationship between loss of NONO and gain of PSPC1 expression.


Double-strand break repair deficiency in NONO knockout murine embryonic fibroblasts and compensation by spontaneous upregulation of the PSPC1 paralog.

Li S, Li Z, Shu FJ, Xiong H, Phillips AC, Dynan WS - Nucleic Acids Res. (2014)

Increased levels of PSPC1 and SFPQ–PSPC1 complex in NONO-deficient cells. (A) Immunostaining of WT-2 and gt-2 MEF isolates using anti-PSPC1 antibody. Scale bar, 10 μm. (B) Immunoblotting to determine levels of SFPQ, NONO and PSPC1 proteins in cells of indicated genotype. Two independent MEF populations, derived from different embryos, were analyzed for each type. Arrowheads denote proteins as indicated. (C) Quantification of data from panel (B). Values are normalized to β-actin. Error bars reflect standard deviation of values from independent MEF populations. (D) Immunoprecipitation (IP), followed by immunoblotting (IB), to protein–protein complexes in MEFs of indicated genotypes.
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Related In: Results  -  Collection

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Figure 3: Increased levels of PSPC1 and SFPQ–PSPC1 complex in NONO-deficient cells. (A) Immunostaining of WT-2 and gt-2 MEF isolates using anti-PSPC1 antibody. Scale bar, 10 μm. (B) Immunoblotting to determine levels of SFPQ, NONO and PSPC1 proteins in cells of indicated genotype. Two independent MEF populations, derived from different embryos, were analyzed for each type. Arrowheads denote proteins as indicated. (C) Quantification of data from panel (B). Values are normalized to β-actin. Error bars reflect standard deviation of values from independent MEF populations. (D) Immunoprecipitation (IP), followed by immunoblotting (IB), to protein–protein complexes in MEFs of indicated genotypes.
Mentions: We investigated PSPC1 expression by immunofluorescence and immunoblotting using anti-PSPC1 antibody (Figure 3A and B). PSPC1 was present at low levels, near the limit of detection, in wild-type MEFs (+/0). Expression was strongly increased in NONO-deficient MEFs (gt/0 and gt/gt). Thus, it appears there is a reciprocal relationship between loss of NONO and gain of PSPC1 expression.

Bottom Line: We tested whether NONO and PSPC1 might also affect repair indirectly by influencing mRNA levels for other DSB repair genes.Of 12 genes tested, none were downregulated, and several were upregulated.Thus, NONO or related proteins are critical for DSB repair, NONO and PSPC1 are functional homologs with partially interchangeable functions and a compensatory response involving PSPC1 blunts the effect of NONO deficiency.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiation Oncology, Emory University, Atlanta, GA 30322, USA Department of Biochemistry, Emory University, Atlanta, GA 30322, USA Institute of Molecular Medicine and Genetics, Georgia Regents University, Augusta, GA 30912, USA.

Show MeSH
Related in: MedlinePlus