Artificial combination of two cis-regulatory elements generates a unique pattern of expression in transgenic plants

We show that a 36-base-pair-long upstream fragment from the soybean hsp17.3-B gene comprising two partly overlapping heat-shock element (HSE)-like sequences can confer heat inducibility to a reporter gene in transgenic tobacco. The heat-shock response does not display organ specificity and is not affected by light. Insertion of these HSE-like elements into the pea rbcS-3A 5' flanking fragment (position -410 to +15) either at position -410 (5' to the enhancer) or at position -49 (between the enhancer and the "TATA" box) renders the transcript level of the reporter gene light-inducible and organ-specific under heat-shock conditions. These results demonstrate the possibility of generating a unique pattern of expression (e.g., light-dependent and organ-specific heat-shock response) by artificial combination of appropriate cis-acting regulatory elements. Moreover, by using the HSE-like sequences as a weak heat-inducible enhancer in the chimeric regulatory regions we uncover the function of negative elements within the pea rbcS-3A upstream region. These negative elements are responsible for a repressed transcript level in roots as well as in dark-adapted leaves. Therefore, the upstream fragment containing two HSE-like elements can be considered a useful tool to test the function of other cis-acting elements.