Hyperthermia is an important modality in cancer therapy and it is also a powerful sensitizer when used in combination with ionizing radiation. However, its effectiveness is limited by the ubiquitous homeostatic phenomenon of thermotolerance. To identify a strategy which antagonizes thermotolerance, and thereby enhance the effectiveness of hyperthermia as a cancer treatment, it will be necessary to understand the molecular basis of the response. Although the expression of heat shock proteins (HSP), the most highly conserved genetic system known, has long been associated with thermotolerance, their role in this phenomenon is unknown. Recently, it has been shown that the yeast 104 kDa hsp is required for the majority of thermotolerance in that organism. It is the objective of this application to extend these studies to the mammalian homologue of this protein, hsp 110. We have previously shown that this is a nucleolar protein (as is yeast hsp 104) and moreover, that nucleolar function (protected recovery of rRNA synthesis) is required for thermotolerance. To further examine hsp 110, we will clone the mammalian cDNA and use it to construct cells which either over- or under-express hsp 110. These will be used to examine the effect of altered expression of the protein on protection of rRNA synthesis and on heat resistance. We will also purify hsp 110 and analyze its structure and function using biochemical and microinjection techniques. The effect of hsp 110 on the efficiency of rRNA synthesis will be examined in an in vitro transcription system, by addition or removal of active hsp 110. The common nucleolar localization of the approximately 110 kDa hsp in yeast and mammalian cells argues that the function of these proteins is conserved. This study will determine if this is the case and will define the role of hsp 110 in mammalian thermotolerance.
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