The balance of nuclear import and export determines the intracellular distribution and function of tomato heat stress transcription factor HsfA2

Tomato heat stress transcription factor HsfA2 is a shuttling protein with d ominant cytoplasmic localization as a result of a nuclear import combined w ith an efficient export. Besides the nuclear localization signal (NLS) adja cent to the oligomerization domain, a C-terminal leucine-rich motif functio ns as a nuclear export signal (NES). Mutant forms of HsfA2 with a defective or an absent NES are nuclear proteins. The same is true for the wild-type HsfA2 if coexpressed with HsfA1 or in the presence of export inhibitor lept omycin B (LMB). Fusion of the NES domain of HsfA2 to HsfB1, which is a nucl ear protein, caused export of the HsfB1-A2NES hybrid protein, and this effe ct was reversed by the addition of LMB. Due to the lack of background probl ems, Chinese hamster ovary (CHO) cells represent an excellent system for ex pression and functional analysis of tomato Hsfs. The results faithfully ref lect the situation found in plant cells (tobacco protoplasts). The intrigui ng role of NLS and NES accessibility for the intracellular distribution of HsfA2 is underlined by the results of heat stress treatments of CHO cells ( 41 degreesC). Despite the fact that nuclear import and export are not marke dly affected, HsfA2 remains completely cytoplasmic at 41 degreesC even in t he presence of LR;IB. The temperature-dependent conformational transition o f HsfA2 with shielding of the NLS evidently needs intramolecular interactio n between the internal HR-A/B and the C-terminal HR-C regions. It is not ob served with the HR oligomerization domain (HR-A/B region) deletion form of HsfA2 or in HsfA2-HsfA1 hetero-oligomers.