Experiment
Muscle bulk in adult healthy humans is highly variable even after height, age, and sex are accounted for. Low muscle mass, due to fewer and/or smaller constituent muscle fibers, would exacerbate the impact of muscle loss occurring in aging or disease. Genetic variability substantially influences muscle mass differences, but causative genes remain largely unknown. In a genome-wide association study (GWAS) on appendicular lean mass (ALM) in a population of 85,750 middle-aged (aged 38-49 years) individuals from the UK Biobank (UKB), the authors found 182 loci associated with ALM (p < 5 x 10^-8). They replicated associations for 78% of these loci (p < 5 x 10^-8) with ALM in a population of 181,862 elderly (aged 6074 years) individuals from UKB. They also conducted a GWAS on hindlimb skeletal muscle mass of 1,867 mice from an advanced intercross line (AIL) between two inbred strains (LG/J and SM/J); this GWAS identified 39 QTL.
To maximize QTL detection power, the authors combined three cohorts of LGXSM AIL mice from their previous reports:
Cohort 1 included 490 mice (253 males and 237 females) from LGXSM filial generation 34 (F34).
Cohort 2 consisted of 506 male mice (84 days of age) from filial generations 50-54 (F50-54).
Cohort 3 included 887 mice (447 males and 440 females) from filial generations 50-56 (F50-56).
The authors collected muscle phenotypes after the animals were sacrificed and frozen. They dissected four muscles and one long bone (tibia or femur) from each mouse at the Pennsylvania State University (n = 584) and the University of Aberdeen (n = 1,283). Each of the four muscles exhibits a different proportion of muscle fiber types and often revealed muscle-specific QTL.
The authors weighed the muscles to 0.1 mg precision and measured long bone length of the hindlimb (mm). All LGSM AIL traits were quantile normalized before mapping QTL.
Cohort 1 was genotyped through the use of a custom SNP genotyping array. These SNPs (n = 2,965) were evenly distributed along the autosomes (mouse genome assembly MGSCv36 [mm8]). The median distance between adjacent SNPs was 446 Kb, and the maximum was 18 Mb.
Cohort 2 was genotyped at 75,746 SNPs (73,301 on the autosomes and 2,386 on X and Y) using the MEGA Mouse Universal Genotyping Array (MegaMUGA; Mus musculus) genome assembly MGSCv37 (mm9); after removing SNP markers that are not polymorphic between the LG/J and SM/J strains, they retained 7,168 autosomal SNPs for subsequent analyses. The median distance between adjacent SNPs was 126.9 Kb and the maximum distance was 15 Mb for all chromosomes except for chromosomes 8, 10, and 14, which had distances of 19, 16, and 16 Mb, respectively.
The authors used a conversion tool in Ensembl to convert SNP positions from mm8 and mm9 to Mus musculus genome assembly GRCm38 (mm10).
Cohort 3 genotypes were obtained from Gonzales and colleagues (Genome wide association analysis in a mouse advanced intercross line. Nat. Commun.
9, 5162 (2018)). These genotypes were generated through the use of genotyping by sequencing. This approach has recently been used and described in detail. Only autosomal SNPs known to be polymorphic in the LG/J and SM/J founder strains (n = 523,027; mm10, GRCm38) were retained for subsequent analyses.
The authors combined the genotype data from cohorts 13 by using PLINK v.1.9, and they imputed missing genotypes by using BEAGLE v.4.1. After applying stringency filters, they retained 434,249 SNPs.
The authors used the software GEMMA (genome-wide efficient mixed-model association) to analyze the mouse phenotypes, identifying QTL peaks associated with tibialis
anterior (TA), extensor digitorium longus (EDL), gastrocnemius (gastroc), and soleus muscle weights.
39 novel skeletal muscle weight QTL were identified:
Skmw48 (skeletal muscle weight 48, EDL) maps to Chr 1: 85550285 - 87395436 bp with a peak -log10 P value score of 6.32 at 87255973 bp.
Skmw49 (skeletal muscle weight 49, TA) maps to Chr 1: 83442398 - 88624908 bp with a peak -log10 P value score of 5.8 at 87325000 bp.
Skmw50 (skeletal muscle weight 50, soleus) maps to Chr 2: 97953543 - 102645782 bp with a peak -log10 P value score of 5.18 at 101662134 bp.
Skmw51 (skeletal muscle weight 51, gastroc) maps to Chr 3: 146300868 - 150982467 bp with a peak -log10 P value score of 5.55 at 148832808 bp.
Skmw52 (skeletal muscle weight 52, TA) maps to Chr 3: 148033317 - 151340029 bp with a peak -log10 P value score of 5.52 at 151047417 bp.
Skmw53 (skeletal muscle weight 53, TA) maps to Chr 4: 62220056 - 69919794 bp with a peak -log10 P value score of 5.7 at 62843781 bp.
Skmw54 (skeletal muscle weight 54, EDL) maps to Chr 4: 65246120 - 66536974 bp with a peak -log10 P value score of 13.57 at 65416188 bp.
Skmw55 (skeletal muscle weight 55, gastroc) maps to Chr 4: 131710493 - 133983265 bp with a peak -log10 P value score of 6.99 at 132914412 bp.
Skmw56 (skeletal muscle weight 56, EDL) maps to Chr 4: 129965551 - 133983265 bp with a peak -log10 P value score of 8.16 at 132943072 bp.
Skmw57 (skeletal muscle weight 57, gastroc) maps to Chr 7: 4577470 - 16953936 bp with a peak -log10 P value score of 6.34 at 5282714 bp.
Skmw58 (skeletal muscle weight 58, TA) maps to Chr 7: 3503912 - 6278482 bp with a peak -log10 P value score of 5.33 at 5282714 bp.
Skmw59 (skeletal muscle weight 59, TA) maps to Chr 7: 44575519 - 45025958 bp with a peak -log10 P value score of 6.78 at 44628444 bp.
Skmw60 (skeletal muscle weight 60, gastroc) maps to Chr 7: 44901740 - 45053023 bp with a peak -log10 P value score of 7.85 at 45025443 bp.
Skmw61 (skeletal muscle weight 61, EDL) maps to Chr 7: 44575519 - 45025958 bp with a peak -log10 P value score of 6.54 at 45025443 bp.
Skmw62 (skeletal muscle weight 62, gastroc) maps to Chr 7: 107294508 - 111013102 bp with a peak -log10 P value score of 8.72 at 109218379 bp.
Skmw63 (skeletal muscle weight 63, TA) maps to Chr 7: 109192251 - 111013102 bp with a peak -log10 P value score of 12.91 at 109218379 bp.
Skmw64 (skeletal muscle weight 64, soleus) maps to Chr 7: 109116573 - 114993501 bp with a peak -log10 P value score of 7.25 at 110686587 bp.
Skmw65 (skeletal muscle weight 65, EDL) maps to Chr 7: 109460130 - 111013102 bp with a peak -log10 P value score of 13.47 at 110986447 bp.
Skmw66 (skeletal muscle weight 66, TA) maps to Chr 8: 128601355 - 128683864 bp with a peak -log10 P value score of 6.95 at 128607127 bp.
Skmw67 (skeletal muscle weight 67, EDL) maps to Chr 10: 119565635 - 119969040 bp with a peak -log10 P value score of 7.5 at 119939128 bp.
Skmw68 (skeletal muscle weight 68, TA) maps to Chr 10: 121540206 - 122019225 bp with a peak -log10 P value score of 5.69 at 121639039 bp.
Skmw69 (skeletal muscle weight 69, TA) maps to Chr 11: 29749029 - 32379535 bp with a peak -log10 P value score of 6.86 at 31523332 bp.
Skmw70 (skeletal muscle weight 70, EDL) maps to Chr 11: 31372737 - 32644975 bp with a peak -log10 P value score of 11.18 at 31680504 bp.
Skmw71 (skeletal muscle weight 71, gastroc) maps to Chr 11: 31325432 - 32795567 bp with a peak -log10 P value score of 5.46 at 31753896 bp.
Skmw72 (skeletal muscle weight 72, soleus) maps to Chr 11: 31379430 - 32835551 bp with a peak -log10 P value score of 5.44 at 31753956 bp.
Skmw73 (skeletal muscle weight 73, TA) maps to Chr 11: 108184643 - 109792775 bp with a peak -log10 P value score of 6.71 at 109541001 bp.
Skmw74 (skeletal muscle weight 74, EDL) maps to Chr 11: 111144720 - 111811380 bp with a peak -log10 P value score of 13.37 at 111390245 bp.
Skmw75 (skeletal muscle weight 75, gastroc) maps to Chr 12: 8162576 - 10401853 bp with a peak -log10 P value score of 6.53 at 9417325 bp.
Skmw76 (skeletal muscle weight 76, TA) maps to Chr 12: 102626836 - 106975629 bp with a peak -log10 P value score of 5.96 at 105094850 bp.
Skmw77 (skeletal muscle weight 77, EDL) maps to Chr 13: 104284915 - 107527275 bp with a peak -log10 P value score of 20.53 at 104435003 bp.
Skmw78 (skeletal muscle weight 78, gastroc) maps to Chr 13: 104261222 - 107527275 bp with a peak -log10 P value score of 12.76 at 104617159 bp.
The SM/J allele confers increased muscle mass at Skmw78.
Skmw79 (skeletal muscle weight 79, soleus) maps to Chr 13: 103150117 - 107527275 bp with a peak -log10 P value score of 6.18 at 104617159 bp.
Skmw80 (skeletal muscle weight 80, TA) maps to Chr 13: 104261222 - 107527275 bp with a peak -log10 P value score of 12.81 at 105148844 bp.
Skmw81 (skeletal muscle weight 81, EDL) maps to Chr 15: 50795676 - 53285678 bp with a peak -log10 P value score of 5.96 at 52165605 bp.
Skmw82 (skeletal muscle weight 82, soleus) maps to Chr 15: 81550235 - 81786938 bp with a peak -log10 P value score of 5.9 at 81743512 bp.
Skmw83 (skeletal muscle weight 83, soleus) maps to Chr 16: 31485647 - 33369337 bp with a peak -log10 P value score of 6.3 at 32545004 bp.
Skmw84 (skeletal muscle weight 84, TA) maps to Chr 17: 32441452 - 35301060 bp with a peak -log10 P value score of 5.27 at 34968724 bp.
Skmw85 (skeletal muscle weight 85, gastroc) maps to Chr 17: 43396777 - 46521031 bp with a peak -log10 P value score of 7.23 at 44724282 bp.
Skmw86 (skeletal muscle weight 86, TA) maps to Chr 19: 5014789 - 8980461 bp with a peak -log10 P value score of 5.65 at 5922485 bp.
In vitro studies of positional candidates confirmed Cpne1 (Skmw50) and Stc2 (Skmw69 - Skmw72) as modifiers of myogenesis.
* Please note that QTL described in the manuscript were renamed to reflect current MGD nomenclature policy.
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