Regulation of protein homoeostasis in aging and age-related diseases

Aging can be explained as a chronic stress which is manifested by the progressive decline in protein homeostasis or proteostasis. In age-related diseases proteostasis is further compromised. As the average human life expectancy has increased, so too has the impact of aging and age-related diseases on our society. Thus research on proteostasis maintenance and repair is becoming increasingly important.

However, measuring proteostasis capacity in an elegant way was not possible so far because of unavailability of effective tools which can be used in variety of model systems. To measure and compare the status of the protein quality control system under different conditions, we generated a series of proteostasis sensors. These proteins are increasingly destabilized versions of firefly luciferase, a protein known to depend on molecular chaperones for de novo folding and refolding. GFP-tagged versions of these mutants resulted in aggregates in mammalian cells over-expressing huntingtin exon1 with extended polyQ. These sensors also aggregated with aging in C. elegans confirming gradual collapse of proteostasis. In summary, we have generated a set of widely applicable proteostasis sensors from a single reporter protein. These can be used to obtain snap-shots of dysregulation of proteostasis at different physiological states as well for screening of small molecule modulators of proteostasis for therapeutic intervention.

Exposure to proteotoxic environmental conditions leads to activation of the cytosolic stress response to restore protein homeostasis. This stress esponse system is mal-functional during aging. Thus inducing cytosolic stress response is a promising strategy to counteract age-related proteostasis collapse. The key feature of this response is the heat shock transcription factor 1 (HSF1) dependent transient expression of stress proteins including molecular chaperones. We have performed a genome-scale RNA interference screen in HeLa cells to identify HSF1 modulators. Our study reveals that signals from multiple key pathways are integrated to regulate the activation and attenuation of HSF1. These include chromatin remodeling, transcription, splicing, membrane signaling, protein folding and degradation. Interestingly, we found that in the absence of a chromatin modifier, namely EP300, HSF1 gets destabilized and inactive. In addition, SILAC based quantitative proteomics experiments revealed that chaperones and proteasome are enriched in nucleus during thermal stress. By quantitative proteomics we also discovered that specific proteasomal subunits interact with transcriptionally active HSF1 during heat stress. Further cell biology experiments highlighted the importance of the proteasomal system in rapid degradation of active HSF1. These results assign a novel and direct role to the proteasome in attenuating the cytosolic stress response.