Heat shock proteins (HSPs) are essential in the maintenance of cellular proteostasis. Their expression and regulation have been shown to vary even among isogenic C. elegans cohorts, with individuals expressing higher levels of HSPs experiencing higher survival rates, which might reflect an underlying ability of each individual to survive upon exposure to heat challenge[1]. Interestingly, though, only worms that express highly at a specific time after heat shock are significantly longer-lived. This suggests that, rather than the level of expression itself, it is the dynamics of expression which correlate with lifespan. Indeed, the expression of HSPs at the whole organism level follows a dynamical behavior in time. We hypothesize that HSPs expression dynamics correlate with worm survival capability, a relationship that we aim to characterize and study. Towards this end, we use the transgenic C. elegans strain TJ375 [
hsp-16.2p::GFP(gpIs1)] and screen the expression of HSP after heat shock. We have built a high-throughput continuous-flow microfluidic platform that enables C. elegans in vivo population screening and sorting based on the quantification of GFP at the whole organism level. Each worm is read in a serial manner while in motion through a straight channel. The quantity of fluorescent proteins within each worm is screened, by means of a laser light sheet which excites the proteins of interest and light is collected by a PMT. Information signals can be processed in real-time for sorting downstream into three possible bins. The sorting mechanism is achieved by hydraulic on-chip valves, which deliver a maximum sorting throughput of 600 worms/min. Worms are then collected in individual vials with no significant loss in viability. To study how HSPs are dynamically regulated in certain subpopulations and to understand the significance of these dynamics in both, stochastic distribution and longevity, we obtain quantitative assessments of the expression of native HSP-16.2 produced by isogenic cohorts at different ages, as well as at different times after exposure to heat shock. Using this data, we use the LASSO technique to infer the existence of singular subpopulations (with dynamics-specific gene expression profiles), which we will then validate with the appropriate sorting of subpopulations of interest and the measuring of their HSPs expression dynamics separately (through subsequent rounds of screening).We will also corroborate the existence of these specific expression dynamics by imaging single individuals immobilized in a microfluidic trap along 48 h after heat shock. Finally, the characterization of the effect of HSPs expression dynamics (i.e. level of transcriptional drift and time profile of expression) on the lifespan of worms will be assessed. 1. Rea, S.L., et al., Nature Genetics, 2005. 37(8): p. 894-898.