A mutation in the myostatin gene increases muscle mass and enhances racing performance in heterozygote dogs

Double muscling is a trait previously described in several mammalian species including cattle and sheep and is caused by mutations in the myostatin (MSTN) gene (previously referred to as GDF8). Here we describe a new mutation in MSTN found in the whippet dog breed that results in a double-muscled phenotype known as the “bully” whippet. Individuals with this phenotype carry two copies of a two-base-pair deletion in the third exon of MSTN leading to a premature stop codon at amino acid 313. Individuals carrying only one copy of the mutation are, on average, more muscular than wild-type individuals (p = 7.43 × 10⁻⁶; Kruskal-Wallis Test) and are significantly faster than individuals carrying the wild-type genotype in competitive racing events (Kendall's nonparametric measure, τ = 0.3619; p ≈ 0.00028). These results highlight the utility of performance-enhancing polymorphisms, marking the first time a mutation in MSTN has been quantitatively linked to increased athletic performance.     

Induction of mdr1b mRNA and P-glycoprotein expression by tumor necrosis factor alpha in primary rat hepatocyte cultures

Mammalian liver exhibits expression of members of the family of multidrug resistance (mdr) transporters (P-glycoproteins). P-glycoprotein isoforms encoded by mdr1 genes participate in extrusion of an array of xenobiotics into the bile. Induction of mdr1b mRNA expression has been shown to occur in rat hepatocytes in response to hepatotrophic growth factors. As the cytokine tumor necrosis factor alpha (TNF-α) is known to exert a direct mitogenic effect on hepatocytes, its influence on mdr1b expression was investigated. In primary rat hepatocytes cultured in the absence of TNF-α, a time-dependent increase in basal expression of mdr1b mRNA and in immunodetectable P-glycoprotein was observed. In cells treated with TNF-α (4,000 U/ml) for 3 days, expression of mdr1b mRNA and of immunodetectable P-glycoprotein was induced approximately twofold. Moreover, intracellular steady-state levels of the mdr1 substrate rhodamine 123 were decreased in cells pretreated with TNF-α in comparison to controls, indicating an increase in functional transporter(s) mediating dye extrusion. Treatment of hepatocytes with antioxidants (1 mM ascorbic acid and 2% dimethyl sulfoxide) for 3 days markedly suppressed mdr1b mRNA and P-glycoprotein expression both in cells cultured in the presence of TNF-α and in the absence of the cytokine, but did not fully abolish mdr1b mRNA induction by TNF-α, supporting the notion that reactive oxygen species participate in regulation of basal mdr1b gene expression during hepatocyte culture. In conclusion, the present data indicate that by inducing mdr1b expression in hepatocytes, TNF-α may affect the capacity of the liver for extrusion or detoxification of endogenous or xenobiotic mdr1 substrates. J. Cell. Physiol. 176:506–515, 1998. © 1998 Wiley-Liss, Inc.     

Motor physiology: a brain of two halves

Brain changes after stroke suggest that undamaged areas may 'take over' the function of damaged regions. Recent studies using magnetic stimulation to disrupt the healthy human brain shed new light on the potential for dynamic compensation across the motor system.     

Translocation of an intracellular antigen to the surface of medullary breast cancer cells early in apoptosis allows for an antigen-driven antibody response elicited by tumor-infiltrating B cells

Tumor-infiltrating lymphoplasmacytic cells are a key feature of medullary carcinoma of the breast (MCB), a distinct subtype of human breast cancer that, despite cytologically anaplastic characteristics, has a more favorable prognosis than other types of breast cancer. Since it has been proposed that the improved clinical outcome is due at least in part to the presence of a prominent lymphoplasmacytic cell infiltrate in the tumor stroma, we recently examined the tumor-infiltrating B cell response in MCB and showed that it is oligoclonal and directed against an intracellular protein translocated to the cell surface upon MCB cell apoptosis. Human Abs cloned from MCB lymphoplasmacytic infiltrate-derived phage display libraries and reflecting the dominant part of the response were used to identify the target Ag as actin. Here, we have characterized in detail the cloned human IgG Abs and the translocation process of actin to the cell surface of apoptotic MCB cells. Our analysis shows that the cloned Abs bind specifically and with high affinity to actin, as determined by ELISA and surface plasmon resonance. Sequence analysis revealed that the Abs are highly somatically mutated, with high replacement to silent ratios, indicative of an Ag-driven, affinity-matured response. Interestingly, the tumor-infiltrating B cells in half the MCB patients mainly exhibited an IgG2 response, while IgG1 dominated in the others. To gain insight to the molecular events that may elicit such an Ab response, we examined the translocation of actin to the cell surface of apoptotic MCB cells using flow cytometry and laser scanning cytometry. Our results show that actin becomes exposed on the cell surface of a large proportion of apoptotic MCB cells as an early apoptotic event. We propose that the Ab response against actin produced by tumor-infiltrating B lymphoplasmacytic cells is Ag-driven, affinity-matured, and elicited due to the increased rate of apoptosis occurring within the MCB tumor that facilitates the translocation and proteolytic fragmentation of intracellular proteins.     

Large-scale purification,dissociation and functional reassembly of the maltose ATP-binding cassette transporter (MalFGK(2)) of Salmonella typhimurium

The maltose ATP-binding cassette (ABC) transporter of Salmonella typhimurium is composed of a membrane-associated complex (MalFGK(2)) and a periplasmic substrate binding protein. To further elucidate protein-protein interactions between the subunits, we have studied the dissociation and reassembly of the MalFGK(2) complex at the level of purified components in proteoliposomes. First, we optimized the yield in purified complex protein by taking advantage of a newly constructed expression plasmid that carries the malK, malF and malG genes in tandem orientation. Incorporated in proteoliposomes, the complex exhibited maltose binding protein/maltose-dependent ATPase activity with a V(max) of 1.25 micromol P(i)/min/mg and a K(m) of 0.1 mM. ATPase activity was sensitive to vanadate and enzyme IIA(Glc), a component of the enterobacterial glucose transport system. The proteoliposomes displayed maltose transport activity with an initial rate of 61 nmol/min/mg. Treatment of proteoliposomes with 6.6 M urea resulted in the release of medium-exposed MalK subunits concomitant with the complete loss of ATPase activity. By adding increasing amounts of purified MalK to urea-treated proteoliposomes, about 50% of vanadate-sensitive ATPase activity relative to the control could be recovered. Furthermore, the phenotype of MalKQ140K that exhibits ATPase activity in solution but not when associated with MalFG was confirmed by reassembly with MalK-depleted proteoliposomes.     

Uterine involution, day and variance of first postpartum ovulation in mares treated with progesterone and estradiol-17beta for 1 or 2 days postpartum

The effects of a single or double regimen of exogenous progesterone and estradiol-17beta (P/E, total dose 300 mg P/20 mg E) were investigated in 50 postparturient Quarter Horse mares. In Trial 1, at 1 and 24 h after foaling, mares were injected with progesterone (150 mg) and estradiol-17beta (10 mg) (n = 7) or 0.9% NaCl (control, n = 13). In Trial 2, within 12 h after foaling, mares were injected with progesterone (300 mg) and estradiol-17beta (20 mg) (n = 13) or 0.9% NaCl (control, n = 17). Mares were examined daily by palpation per rectum and transrectal ultrasonography to determine the day of ovulation. The largest cross sectional diameters of each uterine horn and uterine body were measured ultrasonographically on Day 15 postpartum. Mean uterine diameters did not differ between treatment groups (P > 0.05) in Trial 1, Trial 2 or for combined data for both Trials 1 and 2. For mares bred on the first postpartum estrus pregnancy rates did not differ (P > 0.05) between treatment groups (16/18, 89%) and controls (22/30, 81%) nor was there a difference in mean day to first postpartum ovulation (P > 0.05) between treated and control groups in Trial 1, Trial 2 or Trials 1 and 2 combined. However, fewer (P < 0.05) total P/E treated mares (0/20) ovulated prior to Day 10 postpartum than did control mares (6/30). Variance in days to ovulation was lower (P < 0.05) for P/E treated mares (var = 3.73 days) than for control mares (var = 7.64 days) for data combined from Trials 1 and 2.     

Influence of the estrous cycle on heterosexual aggression in two strains of rats (S3 and WEzob)

Two strains of rats (S₃ and WEzob), which show different levels of aggression in the laboratory, were tested in repeated heterosexual confrontations. Daily 15-min observations were made of the interactions between a female throughout a complete estrous cycle and the same male partner. In both strains the topography of aggression was similar in males and females, but the frequency of specific parameters varied. Males showed more offensive and females more defensive patterns. The overall level of aggression was very low on the day of estrous, when the female was sexually receptive. There were no differences in any elements of female or male behavior between the other 3 days of the cycle. The results support previous conclusions from single-sex encounters that in rats there is no sexual dimorphism in the ability to show aggression.