Identification and characterization of cells factors involved in regeneration of the Ustilago maydis population after oxidative stress

Ustilago maydis je patogena gljiva kukuruza, otporna na ekstremne doze UV i jonizujućeg zračenja. U prirodi U. maydis prolazi kroz dva životna stadijuma, tokom kojih je izložen različitim vrstama stresa. Studije su pokazale da DNK reparacija ima veliki doprinos u oporavku populacije nakon snažnog stresa. Međutim, razumevanje molekularne osnove ekološkog uspeha U. maydis je ostalo nepotpuno. Cilj ovog istraživanja je bilo pružanje jasnijeg uvida u mehanizme koji omogućavaju opstanak populacije ustilaga u nepovoljnim uslovima. Kao metodološki pristup za istraživanje oporavka od snažnog oksidativnog stresa, primenjen je LH test (Liquid Holding)-praćenje vijabilnosti tretiranih ćelija tokom inkubacije u vodi. Rezultati su pokazali da U. maydis ima visok stepen opravka populacije i da se ova obnova ostvaruje multiplikacijom preživelih ćelija zahvaljujući njihovoj sposobnosti da preuzmu i recikliraju produkte degradacije umrlih ćelija. Identifikovano je i okarakterisano 10 ćelijskih faktora (Adr1, Did4, Kel1, Tbp1, Snf8, Slm1, Chk1, Gmt, Snf5, Hsf1) uključenih u proces, putem izolovanja mutanata koji su izgubili sposobnost obnove vijabilnosti populacije (RUS). Poznate su uloge identifikovanih ćelijskih faktora u različitim ćelijskim procesima, a ovo je istraživanje pokazalo da su neki od RUS-faktora uključeni u održavanje integriteta genoma. Takođe je pokazano i da je ustilago metabolički aktivan u vodi i da nakon pada brojnosti populacije usled autogenerisane hipoksije, koristeći RUS, uspeva da obnovi populaciju i u normoksičnim i u hipoksičnim uslovima. Sumirano, RUS bi mogao da predstavlja adaptaciju koja povećava fitnes U. maydis u nepovoljnim uslovima, putem repopulacije nakon delovanja različitih stresogenih faktora.Ustilago maydis is a pathogenic corn fungus, resistant to extreme doses of UV and ionizing radiation. In nature, U. maydis passes through two life stages, during which it is exposed to different types of stresses. Studies have shown that DNA reparation makes a major contribution to the recovery of the population after severe stress. However, understanding the molecular basis of environmental success of U. maydis remained incomplete. The aim of this research was to provide a clear insight into the mechanisms that enable survival of U. maydis population in unfavorable conditions. As methodoligical approach for investigation of recovery from strong oxidative stress, LH assay (Liquid Holding)- monitoring of viability of treated cells during incubation in water, was applied. The results showed that U. maydis had a high degree of population recovery, and that this renewal was accomplished by multiplication of the surviving cells at the expense of the released biomolecules from dead cells. Ten cellular factors (Adr1, Did4, Kel1, Tbp1, Snf8, Slm1, Chk1, Gmt, Snf5, Hsf1) involved in the process were identified and characterized, by isolating mutants that lost the ability to restore population viability (RUS). The roles in various cellular processes of the identified factors heave been already known, and this research showed that some of the RUS factors were involved in maintainance of genome integrity. It was also shown that U. maydis was metabolically active in water and that after population decline due to autogenerated hypoxia, it managed to reconstitute the population in both normoxic and hypoxic conditions, employing RUS. In summary, RUS could be an adaptation that increases fitness of U. maydis in adverse conditions, by repopulation after expsoure to various stressors