18 research outputs found
χ<sup>2</sup> and <i>p</i> value analyses of intergenerational GAA repeat transmissions.
<p>χ<sup>2</sup> and <i>p</i> value analyses of intergenerational GAA repeat transmissions.</p
Effect of Mlh1 on somatic GAA repeat dynamics.
<p>Representative image of the ethidium bromide-stained agarose gels used to determine GAA repeat expansion dynamics from different tissues of 3–5 month-old mice in absence of presence of Mlh1. M = 100 bp size marker, B = brain, C = cerebellum, H = heart, L = liver. WT (<i>Mlh1<sup>+/+</sup></i>) n = 2, KO (<i>Mlh</i><sup>−/−</sup>) n = 7.</p
Effect of MutLα-heterodimers on GAA repeat expansion dynamics and <i>FXN</i> transcription levels.
<p>Effect of MutLα-heterodimers on GAA repeat expansion dynamics and <i>FXN</i> transcription levels.</p
Proposed mechanism of MMR action on intergenetional GAA repeat expansions.
<p>Schematic images representing: (<b>A</b>) a small loop caused by triplex DNA structure; (<b>B</b>) recognition of the loop by the MutS complex, cleavage with endonuclease (ENDO), opening of the loop, recruitment of MutLα and synthesis of expanded DNA, and (<b>C</b>) end of repair by ligation of the further expanded strand and release of MMR proteins.</p
Effect of the MutLα-complex on <i>FXN</i> transcription <i>in vitro</i>.
<p>Relative RT-qPCR analyses of the mean <i>FXN</i> transcriptional level, isolated from MMR-proficient human cells, NCM-460, and MutLα heterodimer-deficient cells, HCT-116 (n = 3). Statistical analysis of the experiment was perform using the student's <i>t</i> test. Error bars = S.E.M, * = p<0.05, ** = p<0.01, *** = p<0.001.</p
Proposed mechanism of MMR action on post-mitotic somatic GAA repeat expansions.
<p>Images represent: (<b>A</b>) a small GAA loop is formed as part of the triplex DNA<b>•</b>RNA R-loop structure caused by transcription within GAA repeats; (<b>B</b>) recognition of the small GAA loop by MutS-heterodimers, cleavage of the CTT DNA strand with an endonuclease (ENDO) and recruitment of MutLα, and (<b>C</b>) release of the RNA, synthesis of expanded DNA and end of repair by ligation of the expanded strand and release of MMR proteins.</p
Effect of MutLα components on somatic <i>FXN</i> transcription <i>in vivo</i>.
<p>Relative RT-qPCR analyses of somatic <i>FXN</i> transcription level based on the MMR genotype (WT, <i>Pms2</i> KO or <i>Mlh1</i> KO) in (<b>A</b>) <i>FXN</i><sup>GAA</sup>/<i>MMR</i> brain tissues (n = 2–4), and (<b>B</b>) <i>FXN</i><sup>GAA</sup>/<i>MMR</i> cerebellum tissues (n = 2–4). Statistical analysis of the experiment was perform using the student's <i>t</i> test. Error bars = S.E.M, * = p<0.05, ** = p<0.01, *** = p<0.001.</p
Intergenerational GAA repeat frequencies.
<p>Analysis of intergenerational transmission of GAA repeat expansion frequency based on the parental genotype (WT = GAA<sup>+</sup>/<i>Mlh1</i><sup>+/+</sup>, Het = GAA<sup>+</sup>/<i>Mlh1</i><sup>+/−</sup>). Frequencies of ‘GAA expansions’, ‘no change’ and ‘GAA repeat contractions’ transmitted to offspring are represented as percentages of total GAA repeat transmissions. Chi squared (<i>x</i><sup>2</sup>) statistical testing was applied to this expriment. WT n = 30 GAA PCR products from 10 mice; Het n = 63 GAA PCR products from 21 mice; *** = p<0.001.</p
Proposed mechanism of MLH1 action on <i>FXN</i> transcription.
<p>Images illustrate: (<b>A</b>) inhibition of transcription by triplex DNA<b>•</b>RNA R-loop formation; (<b>B</b>) binding of MLH1 and MSH6 to the R-loop, together with other unknown factors (?), and (<b>C</b>) release of bound premature mRNA to allow continuation of transcription.</p
Intergenerational GAA repeats.
<p>Representative example of the ethidium bromide-stained agarose gels used to determine the GAA repeat sizes, showing GAA PCR products obtained from a YG22 GAA<sup>+</sup>/<i>Mlh1</i><sup>+/+</sup> parent and 10 GAA<sup>+</sup> offspring. M = 100 bp DNA size marker.</p