Stress proteins have been found to play important roles in several areas of medical investigation, including cancer research. While most of these proteins have been intensively studied, a very few have been almost entirely ignored. Two examples of long recognized but unstudied stress proteins are the 110 kDa heat shock protein (HSP110) and the anoxia inducible 170 kDa glucose regulated protein (GRP170). We have cloned HSP110, determined its sequence and defined its position as a member of a new family of stress proteins, the only characterized member which is the sea urchin egg receptor for sperm. We have also cloned and sequenced GRP170 and shown that it is the HSP110 of the endoplasmic reticulum. Sequence analysis shows that this HSP110 family of proteins exhibits limited similarity with the HSP70s, being the most highly diverged known 'relatives' of this family. It is our working hypothesis that HSP110 plays a major role in the stress response of eukaryotes and that is and its family members have cellular functions not overlapped by the functions of other glucose regulated or heat induced stress proteins. An understanding of the mammalian stress response and its physiological and clinical manifestations requires an analysis of the functions, distributions and modes of regulation of the various members of this newly discovered HSP110 family. In the present application we propose to perform the following aims. 1. We will examine ATP binding, protein binding, and protein reactivation activities using recombinant HSP110. 2. We will also perform a mutational analysis of selected regions and reexamine these properties to identify functional domains. Loss of function mutants will be examined for their ability to confer heat resistance. 3. We will use peptide antibodies against highly conserved family sequences and corresponding oligonucleotide probes to identify additional mammalian HSP110 family members, e.g., a homolog of sea urchin egg receptor, and examine the properties of these proteins. 4. We will define the modes of regulation of each mammalian member of this family to identify the environmental conditions which lead to their overexpression, e.g., anoxia or hyperthermia. The results of these studies can be expected to create new directions of investigation in the field of stress protein biology and yield benefits in many areas of biomedical research, including cancer biology and treatment.
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