Identification of MHC class IIbeta resistance/susceptibility alleles to Aeromonas salmonicida in brook charr (Salvelinus fontinalis).

Pathogen-driven selection is believed to be important in the evolution and maintenance of the polymorphism of the major histocompatibility complex (MHC) genes but have been tested for very few vertebrates. In this study, we first investigate by SSCP (single strand conformational polymorphism) the diversity found at the MHC class IIbeta gene in a population of brook charr (Salvelinus fontinalis) from the Rupert River (Québec, Canada). Secondly, to explore the survival performances conferred by specific alleles and genotypes, individuals from 23 half- and full-sibling families were infected with Aeromonas salmonicida, the causative agent of furonculosis. From the initial brook charr population, a total of six MHC class IIbeta alleles were identified; four complete and two partial coding sequences that include the complete polymorphic beta1 domain. One allele, Safo-DAB*0101, was significantly associated with resistance against A. salmonicida. In addition to homozygotes for this allele, its resistance effect was also detected in heterozygotes for two specific genotypes. Other allelic combinations, namely heterozygous genotypes Safo-DAB*0201/*0301 and Safo-DAB*0301/*0401 were significantly associated with increased susceptibility to furonculosis. Given that its frequency was relatively low (0.0873), the negative frequency-dependent selection hypothesis could explain the advantage associated with the allele Safo-DAB*0101 over the other alleles and highlight the importance of this mechanism to sustain variation at the MHC in brook charr.

Temperature and length-dependent modulation of the MH class II beta gene expression in brook charr (Salvelinus fontinalis) by a cis-acting minisatellite.

It is widely recognized that the variation in gene regulation is an important factor from which evolutionary changes in diverse aspects of phenotype can be observed in all organisms. Distinctive elements with functional roles on gene regulation have been identified within the non-coding part of the genome, including repeated elements. Major histocompatibility complex (MHC) genes have been the subject of an abundant literature which made them unique candidates for studies of adaptation in natural populations. Yet, the vast majority of studies on MHC genes have dealt with patterns of polymorphism in sequence variation while very few paid attention to the possible implication of differential expression in adaptive responses. In this paper, we report the identification of a polymorphic minisatellite formed of a 32 nucleotides motif (38% G+C) involved in regulation of the major histocompatibility class II beta gene (MHII beta) of brook charr (Salvelinus fontinalis). Our main objectives were: to analyze the variability of this minisatellite found in the second intron of the MHII beta gene and to document its effect to the variation of expression level of this gene under different environmental conditions. Distinctive number of the minisatellite repeats were associated with each different MHII beta alleles identified from exon 2 sequences. Relative expression levels of specific alleles in heterozygous individuals were determined from fish lymphocytes in different genotypes. We found that alleles carrying the longest minisatellite showed a significant 1.67-2.56-fold reduction in the transcript expression relatively to the shortest one. Results obtained in three different genotypes also indicated that the repressive activity associated to the longest minisatellite was more effective at 18 degrees C compared to 6 degrees C. In contrast, no significant difference was observed in transcript levels between alleles with comparable minisatellite length at both temperatures. We also depicted a significant up-regulation of the total MHII beta transcript at 6 degrees C relative to 18 degrees C. These results reveal for the first time that a temperature-sensitive minisatellite could potentially play an important role in the gene regulation of the adaptive immune response in fishes.

Tamoxifen and raloxifene differ in their functional interactions with aspartate 351 of estrogen receptor alpha.

The bulky side chains of antiestrogens hinder folding of the ligand binding domain (LBD) of estrogen receptors (ERs) into a transcriptionally active conformation. The presence of a tertiary amine in the side chain of raloxifene, which interacts with a negatively charged residue in helix H3 of the ER LBD [Asp351 in human (h)ERalpha], is important for antiestrogenicity in animal and cellular models. To better understand the molecular basis of the differential activity of tamoxifen and raloxifene, we have examined the influence of tertiary amine substituents and of mutations at position 351 in hERalpha on the activity profiles of tamoxifen derivatives. Results obtained in several cellular model systems suggest that the degree of antagonist activity of tamoxifen derivatives does not strictly correlate with the basicity of the side chain but depends on an optimal spatial relationship between the tertiary amine of these antiestrogens and the negative charge at position 351. Although altering the position of the negative charge at residue 351 (mutation D351E) had little effect on transcriptional activity in the presence of tamoxifen, it drastically increased the partial agonist activity of a tamoxifen derivative with improved antagonist activity as well as that of raloxifene. Our results suggest that contrary to raloxifene, tamoxifen and most of its derivatives do not interact with Asp351 in an optimal manner, although this can be improved by modifying tertiary amine substituents.