Molecular physiology of insect low temperature stress responses .

摘要

Insect responses to low temperature can be broadly-categorized into two groups: freeze-tolerant and freeze-intolerant. Freeze-tolerant insects respond to low-temperatures by allowing their body tissues to freeze, and damage is typically mitigated in this mechanism by limiting ice formation to the extracellular spaces. Freeze-tolerant species accomplish this by increasing the amount of intracellular solutes and promoting ice formation in the extracellullar spaces, which limits damage to membranes. Freeze-intolerant insects suffer damage from cold at temperatures above the point at which body tissues freeze. These organisms prevent damage from low-temperature by increasing the concentrations of polyols in the hemolymph and repair the damage from low-temperature by the production of heat shock proteins. None of what is previously known about insect low-temperature survival is complete, especially at the molecular level. Other than the production of heat shock proteins during recovery from low-temperature damage, no genes have definitively been identified as functionally upregulated to promote low-temperature survival. In addition, the body of knowledge concerning cytoskeletal, membranal, and metabolic changes in insects exposed to low-temperatures is sparse, but all three of these are hypothesized to be involved in low-temperature survival in other organisms. It is the purpose of this work to identify previously unknown molecular mechanisms by which insects deal with low temperatures. This exploration should encompass transcriptional, membranal, and metabolic methods in order to glean the maximal amount of information. The freeze tolerant midge, Belgica antarctica, and the freeze intolerant flesh fly, Sarcophaga crassipalpis, were chosen as model organisms for testing.; A gas chromatography-mass spectroscopic analysis of the membrane lipids from flesh flies revealed that diapause and rapid cold-hardening both dramatically altered the composition of fatty acids in cell membranes. Both physiological conditions increased oleic acid levels, which promotes cellular survival to low-temperatures by widening the window by which cell membranes maintain their liquid crystalline state. In addition, a thin-layer chromatographic analysis of phospholipid head groups revealed that phosphatidycholines were replaced by phosphatidylethanolamines, which also lowers the temperature window by which cell membranes maintain homeostasis. In short, membrane restructuring appears to contribute to low-temperature survival in the freeze-intolerant flesh fly.; A metabolomic analysis of whole-body metabolites isolated from flesh flies in diapause and rapid cold-hardening revealed wide-spread alterations in metabolism. Rapid cold-hardening produced the predictable increase in glycerol concentration, but this increase was also coupled with the increase of another polyol, sorbitol. Rapid cold-hardening also produced increases in many other metabolites that have been previously unknown to be increased, most notably alanine, glutamine and pyruvate. In an environment with these metabolic changes, flesh flies experiencing rapid cold-hardening are well-equipped to survive low-temperature stress because they have increased polyols to protect proteins and membranes, and glutamine is present should the need for heat shock proteins arise during recovery. Increases in glycerol, alanine, and pyruvate were also seen for diapause, but unlike rapid cold-hardening, diapause produced a metabolic profile consistent with a disruption of Krebs cycle activity. Not only does this cellular environment promote cold survival, but it also explains the previous observation that oxygen consumption is very low during flesh fly diapause.; A metabolomic analysis of the freeze tolerant midge, Belgica antarctica, revealed that freezing increased a number of different polyols in whole-body extracts, incuding glycerol, mannitol, and erythritol. Freezing also increased alanine, asparagine, and gl