Experiment
Genetic control of sperm head morphological abnormalities in the B10.MOL-H2wm1/MsRbrc (TEN1) strain was the subject of this study. The TEN1 strain was a congenic line established at the National Institute of Genetics, Japan to define H2 alleles on Chr 17 in the wild mouse population. The original donor was a wild Mus musculus molossinus; the wild-derived H2 allele was transmitted to B10 over 12 generations and has been maintained via sibling breeding.
Teratozoospermia is a condition characterized by the presence of morphologically abnormal sperm and is considered a symptom of infertility. B10.MOL-H2wm1/MsRbrc mice show inherited teratozoospermia at high frequencies. Mean frequencies of abnormal sperm are 50.2% for B10.MOL-H2wm1/MsRbrc mice vs 2.4% for C3H/HeNCrlCrlj (C3H) mice.
An initial cross of B10.MOL-H2wm1/MsRbrc x (C3H/HeNCrlCrlj x B10.MOL-H2wm1/MsRbrc)F1 produced 176 backcross males. Sperm samples were collected from 3 to 5 month old male mice; animals were sacrificed and the epididymides were dissected to obtain sperm for assessing morphology. Morphology was observed by differential interference contrast microscopy (x400 magnification). Genomic DNA was prepared from mouse tails. A genome wide scan was done using 103 SNP markers spaced at 10 cM. QTL analysis was performed to identify loci controlling frequencies of sperm head abnormalities using R software (ver.3.1.3 at http://www.rqtl.org).
Independent effect QTL were considered significant if they exceeded the 95% genome wide adjusted threshold based on 1000 permutations for autosomes and on 22,849 permutations for X Chromosome. Significant and suggestive thresholds were 2.71 and 1.74 for autosomes and 2.81 and 1.86 for X Chromosome, respectively. Epistatic effects were investigated using the genome wide all pairs scan, significance was determined from 1000 permutations.
Two significant pairs of QTL, 24.3 cM on Chr 1 and 29.0 cM on Chr X and 61.8 cM on Chr 6 and 29.0 cM on Chr X were suggested. 592 F2 males were then used for further analysis. Microsatellite markers that covered the 95% confidence interval of the QTLs on Chr 1, 6 and X were added and the F2 samples were genotyped.
Three statistically significant, main effect QTL were detected, Table 1:
QTL Shm3, sperm head morphology 3, mapped the Chromosome 1 at 24.3 cM near rs3022803 with a LOD score of 29.25 and a 95% confidence interval spanning between 21.7 and 27.2 cM. Genetic mapping placed the QTL between Ercc5 and D1Mit303 (Fig 4). The B10.MOL-H2wm1/MsRbrc allele at this locus was considered to be the major locus for the high frequency of sperm head abnormalities when inherited recessively.
QTL Shm4, sperm head morphology 5, mapped to Chromosome X at 32.0 cM near rs3695066 with a LOD score of 6.80 and a 95% confidence interval spanning between 16.0 and 44.0 cM. Genetic mapping placed the QTL between DXMit1.1 and DXMit170 (Fig 4).
QTL Shm5, sperm head morphology 4, mapped to Chromosome 6 at 63.8 cM near rs3023094 with a LOD score of 3.58 and a 95% confidence interval spanning between 45.8-84.9 cM. Genetic mapping placed the QTL between D6Mit219a and D6Mit14 (Fig 4).
Pairwise genome scans estimated two candidates for significant interactions: Chr 1 at 24.1 cM with Chr X at 31.5 cM and Chr 6 at 61.8 cM with Chr X at 31.5 cM.
Both Shm4 and Shm5 were considered conditional loci because both loci acted in specific genotypes. The frequency of sperm head abnormalities was plotted for each genotype concerning the responsible loci Shm3 on Chr 1 (genotyped at D1Mit236), Shm4 on Chr X
(genotyped at DXMit114) and Shm5 on Chr 6 (genotyped at D6Mit59) see Table 2.
At least one additional interacting locus involved in sperm head abnormalities was predicted to exist in the genetic system analyzed in this study.
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