Identification of two RNA-binding proteins (RBPs), FUS and TDP-43, as causative factors of Amyotrophic
Lateral Sclerosis (ALS) resulted in a paradigm shift centered on RNA dysfunction as a disease-driving
mechanism. We previously established a yeast model of FUS cytotoxicity. Using this model, we carried out
genome-wide overexpression screen and identified five yeast genes that rescue FUS toxicity. Three out of
five suppressor genes have human homologs and all three encode RBPs. We found that hUPF1, the
human homolog of one of the three RBPs, rescues the toxicity of FUS in mouse primary neurons and a rat
model of ALS. These findings support the value of our study of FUS toxicity in yeast and uncovered a novel
pathway, currently under development as new therapeutics for ALS. Motivated by this success, we
constructed a new genome-scale library, containing 13,570 full-length sequence verified human gene
clones individually cloned in an inducible yeast-expression vector. Using this library and a newly developed
efficient screening method, we identified 37 human genes, when overexpressed, robustly rescuing FUS
induced toxicity in yeast. Genes encoding RBPs are highly enriched among suppressors (12 out of 37).
Strikingly, all 12 RBP suppressors have known connections to stress granules (SG). The objective of this
application is to define mechanisms of FUS toxicity by studying how the 12 RBPs work to rescue cellular
defects induced by FUS. We hypothesize that toxicity of FUS involves direct impairment of SG function,
and human RBP suppressors rescue FUS toxicity by alleviating detrimental effect of FUS on SG. Two
specific Aims are proposed: i) examine the effect of suppressor RBPs on FUS aggregation and localization;;
and ii) examine the effects of suppressor RBPs on SG. Our hypothesis is consistent with findings that FUS
protein aggregates mis-localize to SG in yeast and in mammalian cells. Based on our preliminary data on
TAF15, one of the human RBP suppressors, we reason that the suppressor mechanisms likely involve
interactions between FUS and the suppressors as well as alterations in SG structure and function.
Regulation and mis-regulation of SG is a pathway that is of immense interest to ALS research field.
Completion of this project will provide deeper understanding of FUS mediated cytotoxicity, its connection to
RNA metabolism, particularly with respect to aberrant SG function. Projects proposed here will not only
expose more undergraduate students to meritorious research in the PI’s laboratory but also help promote
discussion on scientific discoveries in an upper lever undergraduate course, regularly offered by the PI.
Both serve to strengthen the research environment at the PI’s home institution.