Transcription of heat shock (hsp) genes is governed by HSFs - constitutively synthesized transcription factors. HSF1 is the major stress responsive HSF in mammalian cells. Preliminary studies demonstrate that stress-induced activation of HSF 1 requires at least two associated factors: a translation elongation factor EF- 1A and novel untranslated RNA (HSR). The long-term objective of this project is to provide a complete structural and mechanistic description of the HSF1 activation mechanism and its control. It is further proposed to design small modulators of heat shock response based on HSR antisense oligonucleotides for applications in cancer therapy.
Specific aims are: I. Structure/functional characterization of the HSF - EF-1A - HSR ternary complex. Experiments are proposed to determine major structural determinants of HSR from higher and lower eukaryotic cells that are responsible for thermosensing and HSF activation. A combination of protein-protein and protein-RNA footprinting along with cross-linking approaches will be used to map sites within HSF and/or EF-1A that directly contact HSR as well as regions in HSR that participate in such interaction. II. Understanding cellular mechanisms underlying HSF activation by HSR and EF-1A. Biochemical and immunological approaches will be used to determine subcellular localization of HSR, EF- 1A, and HSF before and after heat shock. Experiments in Drosophila and yeast are proposed to test whether a similar complex mechanism of HSF activation exists in lower eukaryotes. It will be determined if other cellular factors associate with HSF-1-HSR-EF-1A complex before and after heat shock and if other members of HSF family require HSR-like RNA for their activity. III. Design novel pro-apoptotic anti-cancer drugs based on HSR antisense oligonucleotides. Preliminary studies demonstrate that transfection of mammalian cells with short oligos complementary to certain regions of HSR suppresses hsp expression and sensitizes cells to heat. Chemically modified stable HSR antisense oligos will be obtained for transfection of thermotolerant breast cancer cultural cells and nude mice tumors for making them more susceptible to apoptosis and existing therapies.
| Shen, Xihui; Banga, Simran; Liu, Yancheng et al. (2009) Targeting eEF1A by a Legionella pneumophila effector leads to inhibition of protein synthesis and induction of host stress response. Cell Microbiol 11:911-26 |
| Shamovsky, Ilya; Nudler, Evgeny (2009) Isolation and characterization of the heat shock RNA 1. Methods Mol Biol 540:265-79 |
| Shamovsky, I; Nudler, E (2008) New insights into the mechanism of heat shock response activation. Cell Mol Life Sci 65:855-61 |
