Role of heterogeneous ribosomes on translational activity from quiescent exist cells

FERNANDEZ, MARTÍN; ORTOLÁ MARTÍNEZ, MARÍA CLARA; ROSSI, SILVIA; GALELLO, FIORELLA; PORTELA, PAULA

Rosario

Congreso; SAIB 2023; 2023

SAIB

Cellular quiescence is the predominant state of all cells. S. cerevisiae is a useful model to study themolecular basis of quiescence and aging. Previously, we described that the ribosome composition ofmonosomes and polysomes changes during translational activation of quiescent cells. Now, weevaluated the role of individual ribosomal proteins (RPs), assessing cell growth and global translationalactivity of mutant cells (Δ) lacking one RP gene (RPG). First, we evaluated the consequences of a singleRPGΔ paralog or non-paralog on growth fitness during quiescence exit. All RPGΔ strains reach the samelevel of stationary phase growth and exhibit similar cellular viability. However, strains rpp2BΔ or rps7BΔshow delayed cellular growth, and a strain rpl38Δ partially reduces lag phase length during quiescenceexit. The results suggest that RP-specific effects control the efficiency of outgrowth from quiescence.Next, the role of RPs on global translational activation of nutrient-stimulated quiescent cells wasevaluated by puromycin incorporation and Renilla luciferase translation reporter assays. We verifiedthat the RPGΔs strain does not affect rRNA biogenesis. Compared to wild-type cells, rpl38Δ strains showa decrease in translational activity. Single deletions of the RPS7A/B, RPL16A/6B, and RPL37A/B paraloggenes affect translational activity during quiescence exit. From the Rps7A/B pair, the paralog Rps7B isthe most heavily expressed both in stationary phase and after nutrient stimulus, which differs fromexpression in exponential phase cells. Rps7B is associated with polysomes, whereas Rps7A remainsassociated with 80S fractions. The rps7AΔ strain reduces global translation activity in comparison torps7bΔ. Therefore, both Rps7 paralogs carry out different roles in translation. The Rpl16A/B paralogsexhibit different behaviors. Rpl16 paralogs show similar distribution in polysomes upon fresh mediumaddition to stationary phase cells. Pull-down assays of Rpl16A-GST suggest that a mixed population ofRpl16A and Rpl16B ribosomes exists on individual mRNAs. As the level of the Rpl16B paralog is generally higher than that of Rpl16A, polysomes containing only the Rpl16B paralog might also be expected. Therpl16BΔ strain shows lower translational activity than rpl16AΔ, suggesting a different role for theseparalogs in translation. Lastly, Rpl37A associates exclusively with the monosome fraction in quiescentcells, and its deletion decreases global translational activity, while RPL7B gene deletion has no effect. ForRP paralogs, it cannot be distinguished if the changes in global translation activity upon RPGΔ are aconsequence of gene dose or gene type requirements. Altogether, the differential expression of RPs andtheir association with ribosomes might give rise to the formation of heterogeneous ribosomes, allowinga diversity in translation functionality during quiescence exit.