Disruption of the HMGA2 Gene in Swine Reduces Growth

The high mobility group AT-hook 2 (HMGA2) has been shown to be a crucial gene for cell growth, proliferation, and apoptosis. HMGA2 is also a strong biological candidate for growth as mutations in this gene alter body size in mice and humans. Compared to wild-type controls, adult mice lacking HMGA2 are 60% smaller, and adult heterozygous mutants are 20% smaller. In humans, a microdeletion in HMGA2 locus resulted in short stature, with no dismorphologies, and normal puberty. In order to determine the effect of HMGA2 on fetal and adult growth in pigs, a transgenic pig line deficient in HMGA2 expression was generated by gene trap approach, using conventional gene targeting with homologous recombination or TALEN mediate homology direct repair (HDR), in fetal fibroblasts (FF).;Somatic cell nuclear transfer (SCNT) was used to generate 8 HMGA2-/+ boars, 11 HMGA2-/-Stra8 boars, and 6 HMGA2-/+ gilts. Weight data revealed that HMGA2-/+ SCNT boars were 17% and 16% lighter in weight over HMGA2+/+ SCNT littermate and naturally bred boars, respectively, since the time point of 14 weeks old. However, trending for 30 weeks was not significant (nonparametric trend test). A 1 HMGA2-/- Stra8 SCNT boar showed severe growth reduction during the early postnatal growth. At the age of 14 weeks, the weight of 1 HMGA2-/- Stra8 SCNT boar was 76.5% lighter than HMGA2+/+ naturally bred boar, the length was 34.7% shorter, the height was 38.9% shorter, and the body cavity was 38.9% smaller. Growth of HMGA2-/+ SCNT gilts showed similar pattern to HMGA2-/+ boars. Weights of HMGA2-/+ SCNT gilts were 20-35% lighter during mid postnatal stages and 25-30% in late stages compared to HMGA2+/+ naturally bred gilts. Nonparametric trend test confirmed that the growth reduction was significant (59 weeks).;Four of six HMGA2-/+ SCNT gilts did not show any estrus behavior. Analysis of follicular development visualization and histological analysis on ovaries showed normal folliculogenesis, despite abnormally low progesterone concentrations. Interestingly, HMGA2-/+ F1 gilts generated by breeding of HMGA2-/+ SCNT founders showed normal estrus at the expected pubertal range for normal gilts. In contrast, HMGA2-/+ SCNT boars had normal puberty and sexual behaviors up to the present (2 years and 4 months). Normally organized seminiferous tubules were identified by histological analysis on testis. While HMGA2-/+ SCNT boars were normal, all HMGA2 null boars (generated by SCNT or by breeding) had undescended testes (Cryptorchidism) with lack of gubernaculum bulbs, showing no evidence of spermatogenesis until 4 months of age.;Mating of HMGA2-/+ SCNT founders resulted in 3 pregnancies to term without HMGA2 null piglets. To determine the timing of fetal loss and to identify a cause for the loss, pregnancies at D40 and D78 were examined, resulting in confirmation of the presence of all four different genotypes, including HMGA2-/-. While no defects were detected in D40 and D78 fetuses per se, placental abnormalities were seen only in HMGA2-/- conceptuses from D78 litters at both macroscopic and microscopic level. To determine whether lack of HMGA2 or intrauterine resource depletion was responsible for lethality of HMGA2 null pigs after D78, SCNT was performed to generate a pregnancy carrying only the HMGA2 genotype. Successful generation of 10 live HMGA2 null clones indicates that HMGA2 deficiency does not cause fetal lethality.