Evaluating the Dose Dependent Response to DTT Treatment Resulting in HSF Activation in Drosophila cells

Abstract

Proteins drive many of the biological processes in cells. To do this, they fold into complex 3D structures that are integral for their functionality. Notably, various environmental and chemical stressors can disrupt protein folding and thus disable the functions of proteins, threatening the livelihood of cells. To mitigate this stress, organisms initiate the highly‐conserved heat shock stress response. In eukaryotes, the master heat shock activator heat shock factor (HSF) is rapidly recruited to the Hsp70 heat shock protein genes and triggers the recruitment of additional co‐activator proteins that facilitate gene expression. This leads to the production of heat shock proteins that function as molecular chaperones to promote refolding of proteins, prevent aggregation and increase protein degradation pathways. Notably, activation of the heat shock response pathway, can be visualized by measuring GFP‐tagged HSF binding to the heat shock protein genes in living Drosophila salivary gland nuclei. Our lab is currently using this technique to identify novel compounds that induce the heat shock response pathway. Our pioneering experiments have shown that diamide and hydrogen peroxide, two chemicals known to cause protein misfolding and activation of the heat shock response pathway, trigger the recruitment of GFP‐HSF to the Hsp70 loci in living salivary gland cells to a similar level as HS stress. In addition, to our surprise, high levels of Dithiothreitol (DTT, 100 mM), a chemical known to cause protein misfolding and activation of the unfolded protein response pathway (UPR), results in the recruitment of GFP‐HSF to the Hsp70 gene loci. Here we describe experiments to further explore a possible dose dependent activation of HSF by DTT. Support or Funding Information Student Faculty Research Grant for Slippery Rock University