CAG Repeat Lengths in X- and Y-bearing Sperm Indicate That Gender Bias during Transmission of Huntington's Disease Gene Is Determined in the Embryo

Abstract

The size of the CAG tract at the Huntington's disease (HD) locus upon transmission depends on the gender of the parent. However, the basis for the parent-of-origin effect is unknown. To test whether expansion and contraction in HD are "imprinted" in the germ cells, we isolated the X-and Y-bearing sperm of HD transgenic mice. Here we show that CAG repeat distributions in the X-and Y-bearing spermatozoa of founding fathers do not differ. These data show that gender-dependent changes in CAG repeat length arise in the embryo.The mechanism for expansion in HD 1 and other trinucleotide disorders is not known. However, one of the poorly understood features of expansion is the gender bias that is associated with transmission. Large changes in repeat number are known to be transmitted through the paternal line in HD (1, 2), SCA1 (3), dentatorubral-pallidoluysian atrophy (DRPLA) (4), MachadoJoseph disease (5), and spinal and bulbar muscular atrophy (SBMA) (6) and through the maternal line in Fragile X (7, 8). The gender bias transmission has also been documented in transgenic mice (9 -12). Similar to humans, HD transgenic mice transmit expansions predominantly through the male germ line (9,10,12). Moreover, we found that expansion and contraction of CAG repeat size in the hHD transgene depended on the gender of the progeny (9).Expansions are primarily seen in males and contractions dominate in females. The molecular basis for the parent-oforigin effect is unknown. The gender dependence of expansion can be due to differences in CAG repeat distribution in X-and Y-bearing sperm. Alternatively, the change in repeat number, contraction, or expansion can take place after fertilization in early embryogenesis. In order to distinguish between these two mechanisms, the isolation and purification of the X-and Ybearing sperm were required. This, however, was not possible using conventional technology.In present work, we have developed techniques and used advanced technology to separate pure populations of X-and Y-bearing germ cells from HD transgenic mice. We show here that CAG repeat distributions in X-and Y-bearing parental sperm are not different. Repeat expansion occurs equally well within both X and Y chromosomes. This indicates that the gender-dependent processing of CAG repeats must take place post-zygotically. Thus, these data provide evidence for a new kind of imprinting that may be important in the interpretation of genetic data in other systems. EXPERIMENTAL PROCEDURESAnimals-HD transgenic male mice (line B6CBA-TgN R6/1) were originally purchased from The Jackson Laboratory. The colony was maintained at the animal core facility, Mayo Clinic/Foundation. Animals were routinely screened for the presence of HD transgene by PCR (10, 18).Preparation of Single Cell Suspension and Flow Sorting-Mouse epididymis was dissected from the animal, placed in phosphate-buffered saline, and chopped with a razor. Resulting suspension was filtered through cheesecloth. The supernatant was centrifuged at 1500 rpm, and the pellet was resuspended...

Factors associated with HD CAG repeat instability in Huntington disease

“…Although in a previous study, offspring sex was not found to influence the intergenerational repeat instability of the HD CAG repeat,27 the discrepancy between that study and our data may lie in the fact that we analysed transmissions from male and female HD carriers separately. Interestingly, the direction bias of repeat-size changes we observed in male and female offspring parallels that seen in a transgenic HD mouse model 17 18…”

“…In a mouse model of HD, a postzygotic mechanism has been proposed to explain the finding that expansions predominate in male offspring whereas contractions predominate in female offspring from the same parent 17 18…”

Long-Term Effects of Mouse Intracytoplasmic Sperm Injection with DNA-Fragmented Sperm on Health and Behavior of Adult Offspring1

“…An association between ART and congenital malformation syndromes with imprinting defects and rare tumors, such as retinoblastoma, has been documented [72]. Also, epigenetic differences have been reported between male and female embryos that may explain why we have found obesity only in female mice [73][74][75][76].…”

Sex differences in behavior and striatal ascorbate release in the 140 CAG knock-in mouse model of Huntington's disease

“…Results showed that the CAG repeat was significantly expanded in the male offspring and contracted in the female offspring compared to the parent [11]. CAG repeats were the same length in both X and Y bearing sperm indicating that this change was associated with embryo development rather than changes occurring during spermatogenesis [12]. Thus, the CAG repeat is likely to be shorter in females and longer in males with successive generations.…”