Journal: bioRxiv

Article Title: Genetic modifiers of somatic expansion and clinical phenotypes in Huntington’s disease reveal shared and tissue-specific effects

doi: 10.1101/2024.06.10.597797

Figure Lengend Snippet: A. The somatic expansion ratio of the HTT exon 1 CAG repeat in blood DNA is allele length- and age-dependent. The scatterplot shows the somatic expansion ratio (SER = in blood DNA plotted against age at sampling in 3,999 Enroll-HD participants. Points are color-coded based on the length of the inherited uninterrupted CAG length determined by MiSeq. Lines on the scatterplot are the SER values predicted by the multiple linear regression ln(SER) ∼ β 0 + β 1 .CAG + β 2 .age + β 3 .(CAG x age) + β 4 .CAG 2 + β 5 .age 2 + β 6 .(CAG 2 x age) + β 7 .(CAG x age 2 ) + ε 0 . Boxplot (25th and 75th percentiles (box), median (line), and range (whiskers, capped at 1.5x the interquartile range)) of the relationship between canonical and non-canonical HTT repeat sequences with the somatic expansion ratio in blood DNA adjusted for CAG length, age, CCG length and PCR batch ( i.e., the residuals of the multiple linear regression ln(SER) ∼ β 0 + β 1 .CAG + β 2 .age + β 3 .(CAG x age) + β 4 .CAG 2 + β 5 .age 2 + β 6 .(CAG 2 x age) + β 7 .(CAG x age 2 ) + β 8 .CCG + β 9 .PCRbatch). B. Boxplot of average number of CAG repeats gained over four weeks for HTT exon 1 variants knocked into the AAVS1 locus in RPE1 cells. Clonal knock-in RPE1 lines with canonical (n = 7), CAACAG-dup (n = 8), or CAA/CCA-loss (n = 11) HTT exon 1 variants are each represented by a single dot, positioned based on the average repeat gain of triplicate cultures. The average repeat gain represents the difference between mean CAG repeat (weighted on the fragment analysis peak height) at four-weeks and day-zero, which was adjusted for the effect size of day zero mean repeat length that ranged between 112-120 CAGs. Both CAACAG-dup and CAA/CCA-loss alleles showed significantly reduced CAG repeat expansion compared to canonical (p < 0.001 and = 0.015, respectively). C. Somatic expansion ratio length in post-mortem frontal cortex from 488 HD individuals carrying canonical expanded repeat sequences (open circles) and 5 carrying CAA/CCA-loss HD alleles (filled red circles) is plotted versus modal HTT CAG repeat D. Somatic expansion ratio in post-mortem frontal cortex from 488 HD individuals carrying canonical repeat sequences (open circles) and 5 carrying CAA/CCA-loss HD chromosomes (filled red circles) is plotted versus their ages at death. E. Boxplot of the relationship between canonical and non-canonical HTT repeat sequences with the somatic expansion ratio in HD frontal cortex DNA, adjusted for CAG length, age, CCG length and PCR batch as described in B. The CAA/CCA-loss allele carriers were not significantly different from those with canonical alleles (p = 0.96)

Article Snippet: The resulting plasmids with matching uninterrupted CAG repeat sequences were nanopore sequenced (Plasmidsaurus, SNPsaurus LLC) to confirm sequence identity and the repeat size was quantified by fragment analysis as described in the HTT CAG repeat genotyping section.

Techniques: Sampling, Knock-In

Journal: bioRxiv

Article Title: Genetic modifiers of somatic expansion and clinical phenotypes in Huntington’s disease reveal shared and tissue-specific effects

doi: 10.1101/2024.06.10.597797

Figure Lengend Snippet: A . GWAS of somatic CAG expansion measured by the PPS method applied to ABI blood CAG sizing traces is shown for SNVs with MAF > 1%. The dashed black line represents the threshold for genome-wide significance (p = 5.0E-08). B . An example of ABI traces from a CAACAG-dup allele (top) and a canonical allele (bottom), both carrying uninterrupted repeat lengths of 44 CAGs. The top panel reveals a mispriming artefact that scores as CAG expansion in the PPS method, leading to spurious signal at HTT in the GWAS of panel A. C . Association analysis of the PPS CAG expansion phenotype conditioned on rs183415333, which tags the non-canonical CAACAG-dup allele on HD chromosomes, is shown for SNVs with MAF > 1% in the HTT region. Conditioning removes the spurious signal and leaves rs146151652 as the top SNV, the same HTT 5’-UTR SNV detected in the MiSeq somatic CAG expansion GWAS ( , ).

Article Snippet: The resulting plasmids with matching uninterrupted CAG repeat sequences were nanopore sequenced (Plasmidsaurus, SNPsaurus LLC) to confirm sequence identity and the repeat size was quantified by fragment analysis as described in the HTT CAG repeat genotyping section.

Techniques: Genome Wide