FUNCTIONAL-ANALYSIS OF THE TEMPERATURE-DEPENDENT EXPRESSION OF THE BARLEY HVHSP17 GENE PROMOTER IN MONOCOT AND DICOT CELL SYSTEMS

The regulation of a barley heat shock gene (Hvhsp17) has been investig ated by means of transient expression assays with monocot and dicot pr otoplast systems. The complete 1700-bp 5' region of Hvhsp17, and the d eletions of 600 and 170 bp, respectively, were fused with the bacteria l beta-glucuronidase (beta-GUS) marker gene, to give the following exp ression vectors: pHSGUS1 (-1700 pHS/GUS fusion), pHSGD1 (-600 pHS/GUS fusion) and pHSGD2 (-170 pHS/GUS fusion). pHSGUS1 and pHSGD1 contain t wo heat shock elements (HSEs) located in position -238 and -168, respe ctively, while pHSGD2 contains only one HSE located near the TATA box. As control expression vector, plasmid pBI221 (Clontech) was used, in which the beta-GUS gene is under the control of the CaMV 35S constitut ive promoter. All expression vectors were transfected into protoplasts derived from leaf mesophyll cells of tobacco (dicot) and some cereal species (monocots). beta-GUS activity was determined after a heat shoc k treatment of 1 h at 40 degrees C. Transient expression assays showed that the promoter of Hvhsp17 was active and regulated by thermal indu ction only in monocot cell systems, while not expressed in tobacco cel ls. Analyses performed with the deletion constructs, indicated that th ermal activation of beta-GUS was obtained only when both the HSEs were present in the promoter region.