STABLE TRANSFORMATION OF AN ARABIDOPSIS CELL-SUSPENSION CULTURE WITH FIREFLY LUCIFERASE PROVIDING A CELLULAR-SYSTEM FOR ANALYSIS OF CHAPERONE ACTIVITY IN-VIVO

Using Agrobacterium, we developed a method to transform an Arabidopsis cell suspension culture. A stably transformed cell line expressing hi gh levels of firefly luciferase (Luc) was used for in vivo studies of thermal denaturation and renaturation of the enzyme and the protective role of different chaperones. Luc activity was monitored under heat s tress and recovery conditions in control, thermotolerant cells and cel ls expressing plant chaperones after transient cotransformation with p lasmids encoding proteins of the heat shock protein Hsp90, Hsp70, or H sp20 family. The effects of the expressed proteins were specific. The Hsp17.6 class I protein maintained Luc activity on a level comparable with that observed in thermotolerant cells and improved Luc renaturati on. Although transient expression of Hsp90 did not protect Luc from th ermal denaturation, it accelerated Luc renaturation during recovery. I n contrast to the other chaperones tested, overexpression of Hsp70 alo ne had no effect on denaturation and renaturation of Luc but enhanced Luc renaturation if coexpressed with Hsp17.6.