| This thesis describes an investigation of the expression of an alfalfa (Medicago sativa L.) class A4 heat shock transcription factor (HSF), elements of the heat shock response and the potential utility of engineering HSFs to protect against low temperature stress. To study HSFs in alfalfa, a new, cold-inducible HSF homologue, designated MsHSFA4, was isolated from a field-acclimated cDNA library. Using northern and western macroarrays, transcript levels for HSF, HSP 18, HSP 86 and protein levels for low molecular weight HSPs (ImwHSPs) were monitored under controlled chamber and field conditions. To test the potential to improve low temperature tolerance through the overexpression of a HSF, an Arabidopsis HSF (AtHSFA4a) was engineered into alfalfa, and the plants were evaluated for winter survival and biomass accumulation.; Under both heat and cold stress, HSF, HSP 18, HSP 86 mRNAs and ImwHSP protein accumulations exhibited a variety of expression patterns. Some of these patterns exhibited distinct responses that were directly related to the dormancy classification among 10 alfalfa cultivars. Under both heat and cold stress, MsHSFA4 expression levels were greater in magnitude and induction rate in dormant cultivars as compared to non-dormant cultivars. Under field stress, HSF, HSP 18, HSP 86 mRNAs and ImwHSP protein accumulations also exhibited a variety of expression patterns. Dormancy cultivar effects were observed under HSF, HSP 86 mRNAs and ImwHSP protein accumulations. The most prominent of these effects occurred among HSP 86 mRNA expression in bud tissue, which exhibited notably higher levels of expression in non-dormant cultivars as compared to dormant cultivars. The results herein confirm that MsHSFA4 is a functional transcriptional activator, active under heat and cold stress, and is most likely involved in activation of elements of the heat shock response under low temperature stress.; Engineering of AtHSFA4a in alfalfa exhibited effects on winter survival and fall biomass. Plants containing sense constructs exhibited increased winter survival and greater fall biomass. Plants containing antisense constructs exhibited decreased winter survival and a reduction in fall biomass. This trait of increased and decreased biomass was further observed in the F 1 progeny, indicating that the trait is heritable and a result of engineered HSF activity. (Abstract shortened by UMI.)... |
