Thermal stress and evolution: Molecular responses to temperature in natural populations of Gambusia

The western mosquitofish, Gambusia affinis, lives in highly fluctuating thermal environments. Such environments are a source of heat stress where one manifestation is the tertiary structure of proteins may take aberrant forms or the proteins may denature altogether. Such improperly folded proteins may also form large aggregates that interfere with other cellular activities. Molecular chaperones are proteins that facilitate the correct folding of other proteins so that they maintain a stable tertiary structure. In addition, they promote the proper refolding or degradation of denatured proteins after cellular stress. Heat shock proteins form one of the main classes of molecular chaperones. The genes encoding the 70 kd and 90 kd families were cloned and sequenced in Gambusia affinis. Phylogenetic analyses suggested these genes corresponded to heat shock cognate 70 (hsc70), heat shock protein 70-2 (hsp70-2), heat shock protein 90 a (hsp90a), and heat shock protein 90b (hsp90b). All or a substantial portion of the four hsps were characterized and found to be very similar to homologous fish sequences and other vertebrates.;Gambusia affinis from fifteen populations (spanning much of their native range in Oklahoma and Texas) were analyzed for nucleotide sequence variation in the ND2 mitochondrial gene. Molecular results indicate a lack of variation across populations suggesting that there is little population structure among the geographically widespread populations. This may be due to the founding of these populations by a monomorphic ancestor followed by dispersal. Additionally, sequences from two heat shock protein genes (hsp70 and hsp90) were analyzed across Gambusia spp. to determine if there is a link between successful colonization of thermally variable environments and fixed nucleotide sequence patterns. The extremely conserved sequences across species indicate the ability to withstand thermal stress is not a function of variation in primary protein structure.;Many organisms respond to fluctuations in temperature by altering the expression of a family of evolutionarily conserved genes known as heat shock proteins (hsps). The mediation of these hsp genes in the heat shock response of fish exposed to varying degrees of thermal stress over time confers an adaptive value. This study explores the differential regulation of four-heat shock protein genes in the mediation of thermal stress in three species of Gambusia where habitat conditions differ in thermal constancy and temperature extremes. Results suggest that the ability of these fish to live in thermally contrasting habitats is a function of the regulation of genes in the hsp70 and hsp90 families. Gambusia living in thermally variable habitats upregulate all hsp genes as heat shock temperatures increase; however, gene expression of hsp70-2, hsc70, and hsp90b decreases or remains steady in the obligate spring dweller, Gambusia geiseri...