Experiment
To characterize the genetic architecture of hybrid male sterility on a genome wide scale an F2 intercross between two wild derived lines of inbred mice were used.
M. m. domesticus mice, WSB/EiJ and M. m. musculus, PWD/PhJ mice were crossed in reciprocal directions to generate F1 hybrids that were then bred with siblings. Pups were weaned into same sex groups at 21 days and males were separated into individual cages at ~56 days. Males were then phenotype at 70 days.
Five diagnostic measures of subfertility and sterility were quantified: testis weight (absolute weight of right testis), sperm density (estimated using a Makler chamber), sperm head morphology ( measured in samples collected from the epididymides), proportion of abnormal sperm (4 primary abnormalities: proximal bent tail, distal bent tail, missing head or missing tail, and severely amorphous head) and cross sectional area of seminiferous tubules (measured only in stage VII tubules).
Genomic DNA was extracted from liver tissue. SNP markers were designed from the Perlegen phase 4 release of the mouse resequencing project. A total of 331 SNP markers were genotyped using the Sequenom iPLEX MassARRAY system. After conservative filtering steps for quality control, 198 SNPs were genotyped across the autosomes, X Chromosome, Y Chromosome and mitochondrion of 310 F2 male mice.
All phenotypes were analyzed using standard interval mapping except for abnormal sperm types, which were analyzed using the extended Haley-Knott method. Genome wide significance thresholds were calculated from 1000 permutations of the autosomes and separate permutations for the X chromosome. For phenotypes that did not follow normal distribution results from several alternative mapping procedures were compared, including nonparametric interval mapping, interval mapping for binary traits, and a two part model for phenotypes that exhibited a spike near zero in the distribution.
Standard, single QTL interval mapping detected significant QTL for each hybrid sterility phenotype in the F2 male mice [Table 3, Fig 4, Fig S2].
Each X-linked QTL associated with the M. m. musculus genotype (PWD/PhJ) was associated with a large reduction in fertility. In contrast, infertility was associated with both
M. m. domesticus (WSB/EiJ) and M. m. musculus (PWD/PhJ) alleles at autosomal QTL. For 6 of the 11 QTL the allele causing hybrid male sterility was recessive. One case of underdominance, where the least fertile genotype was associated with heterzygous males, was observed at locus Spdq2 on Chr 17. [Table 3.]
Two-QTL model analysis was performed and several pairs of QTL were identified affecting each other in an additive manner; however, little evidence of epistatic interactions were observed among the QTL.
Multiple models that jointly fit any number of QTL were also considered using an automated forward/backward step wise search algorithm. In addition to statistical improvements over the one-QTL model several additional QTL were found [Table 4].
In the single QTL interval mapping QTL Semtaq1 (seminiferous tubule area QTL 1) mapped to Chromosome 18 with a LOD score of 4.36 at 34.0 cM (69.6 Mb), the 1.5 LOD interval spanned from 60.3 to 80.0 Mb. In the multiple model analysis the QTL mapped to 33 cM, LOD=4.62, CI 58.0-78.1 Mb accounting for 6.0% of trait variance with PWD/PhJ alleles inhertied in a dominant pattern associated with sterility.
Additional seniniferous tubule QTL were mapped in the multiple model analysis:
QTL Semtaq2 (seminiferous tubule area QTL 2) mapped to 27.8 cM on Chr 8, LOD=4.76, CI 35.5-102.7 Mb, accounting for 6.09% of trait variance with WSB/EiJ alleles contributing to sterility when inherited in an additive pattern.
QTL Semtaq3 (seminiferous tubule area QTL 3) mapped to 16.1 cM on Chr 14, LOD=4.35, CI 25.6-63.2 Mb, accounting for 5.55% of trait variance with PWD/PhJ alleles contributing to sterility when inherited in an additive pattern.
QTL Semtaq4 (seminiferous tubule area QTL 4) mapped to 39.0 cM on Chr 16, LOD=4.02, Ci 66.8-97.2 Mb accounting for 5.11% of trait varaince with PWD/PhJ alleles contributing to sterility when inherited in a dominant pattern.
Focusing on interactions involving the X chromosome, conditioned on PWD/PhJ (M. m. musculus) alleles, 5 autosomal QTL were detected interacting with the X Chromosome. For each abnormal sperm QTL in the interaction there were higher proportions of abnormal sperm when the X-linked PWD/PhJ allele was combined with autosomal QTL homozygous for the M.m domesticusWSB/EiJ alleles. [Table 5.]
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