HSP90 Controls SIR2 Mediated Gene Silencing

Abstract

In recent years, Hsp90 is found to interact with several telomeric proteins at various phases of cell cycle. The Hsp90 chaperone system controls assembly and disassembly of telomere structures and thus maintains the dynamic state of telomere. Here, for the first time we report that the activity of another telomeric protein Sir2p is modulated by Hsp82, the ortholog of Hsp90 from budding yeast (Saccharomyces cerevisiae). In a temperature sensitive Hsp90 deficient yeast strain (iG170Dhsp82), less abundant Sir2p is observed, resulting in de-repression of telomere silencing and a complete loss of mating type silencing. Intriguingly, over expression of Hsp90, either by exposing cells to heat shock or by introducing HSP82 overexpression plasmid also yields reduced level of Sir2p, with a consequential loss of telomere silencing. Thus, Hsp90 homeostasis maintains the cellular pool of Sir2p and thereby controls the reversible nature of telomere silencing. Interestingly, such regulation is independent of one of its major co-chaperones Sba1 (human ortholog of p23).

Both the Charged Linker Region and ATPase Domain of Hsp90 Are Essential for Rad51-Dependent DNA Repair

“…Its pivotal role in telomere capping and maintenance has been established (44). Recently, work on Sir2 protein dynamics as a function of Hsp82 activity has recognized a link between Hsp82 and telomere silencing (38). Two recent studies on Sba1 and Aha1, two important cochaperones of Hsp82, have unraveled an interesting connection with DNA repair proteins.…”

“…For the temperature-sensitive strains, the cells were divided into two groups; one part was incubated at 25°C for 4 h, and another part was incubated at 37°C overnight. After that, equal numbers of cells were taken and precipitated, and protein was isolated from them by the trichloroacetic acid (TCA) method, followed by Western blotting (38). A rabbit anti-Rad51 antibody (Promega), a mouse anti-Hsp82 antibody (Calbiochem), and a mouse anti-Act1 antibody (Abcam) were used at a 1:5,000 dilution.…”

“…Total RNA was isolated from the iG170Dhsp82 strain after growth at 25°C and at 37°C for 4 h. In another assay, wild-type strain HH1a-p2HG/Hsp82 and the charged linker deletion mutant, as well as P82a and the hsp82 T101I point mutant, were either left untreated or exposed to 0.03% MMS for 2 h, and by using the acid phenol method as described previously (38), we isolated RNA from these strains. We synthesized cDNA exactly as described in reference 38.…”

Heat Stress-Induced Cup9-Dependent Transcriptional Regulation of SIR2

“…The Hsp82 null condition not only leads to the reduced abundance of Sir2 but also results in the inactivation of Sir2 proteins. On the other hand, Hsp82 overexpression leads to the reduction of the steady-state level of Sir2 protein, though it does not affect the mating type silencing activity of Sir2 (5). In this report, we provide mechanistic insights into Hsp82 overexpression phenotype which occurs while cells are exposed to heat shock.…”

“…Sir2 deacetylates histone H3 (at K9, K14, and K56) and H4 (particularly K16) to regulate telomeric heterochromatin structure in yeast (3,4). Our previous studies have demonstrated that heat stress and concomitant overexpression of Hsp90 result in euchromatinization of silent subtelomeric chromatin by reducing the steady-state level of Sir2 (5).…”