PROTEOME WIDE ANALYSIS OF GRANULAR PROTEIN COMPLEXES EVOKED BY DIFFERENTIAL HEAT STRESS STRIGENCY IN Saccharomyces cerevisiae

GALELLO, FIORELLA; ROSSI S.; SOLARI, CLARA A.; FERNANDEZ G.; PORTELA P.; BARRAZA, CARLA E.; VALACCO P.; MORENO S.

Buenos Aires

Congreso; Reunión Conjunta de Sociedades de Biociencias; 2017

Cellular responses to stress comprise a variety of different mechanisms, including translation arrest and the relocation of translationally repressed mRNAs to ribonucleic particles (mRNPs) like stress granules (SGs) and processing bodies (PBs). Given recent evidence on the role of liquid phase transition in signalling and cytosolic mRNPs formation, it is possible that SGs might represent regions where certain processes or the activity of enzymes are concentrated. Our results from biochemical approaches and microscopy analysis show that under mild heat stress the catalytic subunit of PKA isoform Tpk3 aggregates and promotes aggregation of eIF4G, Pab1 and eIF4E, whereas severe heat stress leads to the formation of PBs and SGs that contain the Tpk2 isoform and a larger 48S translation initiation complex. PKA affects the translational response to heat stress, where each Tpk catalytic isoform appears to have a different role, with Tpk2 and Tpk3 playing negative and positive roles in the translational response, respectively. We favour a model where depending on the severity of an external stimulus, such as heat stress, each catalytic isoform of PKA interacts with a complex network of distinct protein factors and potential substrates. Our ongoing studies are focused on a global characterization of protein complexes under different heat stress conditions. To this end, we performed a label-free quantitative proteomic analysis of granular enriched fraction from mild and severe heat stress using a QExactive. We identified proteins exclusively enriched in mild and severe heat stress, as well as proteins common to both groups. Gene Onthology analysis showed annotations associated with several GO biological processes, such as structural constituent of ribosome, RNA binding and translation factor activity. Networks built from these results will let us start defining how different degrees of severity of the same stress evoke a specific response on RNP assembly.