Figure 1.

tRNAs are trimmed upon stress and repaired once normal growth conditions are restored. (A) Northern blot analyses of tRNAs on total RNA extracted from T. brucei. RNA was isolated from procyclic T. brucei cells either during the exponential growth phase (Exponential) or after heat shock, nutritional deprivation (Nutr. stress), oxidative stress, during the stationary phase or from exponentially growing bloodstream form (Exponential BSF). Full length tRNAs and 3′ trimmed tRNAs are indicated with filled and open arrowheads, respectively. (B) Total RNA extracted from cells growing exponentially (expo), subjected to nutritional stress (stress), or allowed to recover in normal media for 15 or 30 min after stress, was analyzed by northern blot for tRNA^Val and tRNA^Arg (NB, upper panel). Full length tRNAs and 3′ trimmed tRNAs are indicated with filled and open arrowheads, respectively. Ethidium bromide staining (EtBr, lower panel) shows the migration of the bulk of the tRNAs. (C) Decay of total endogenous RNA was investigated by a 2-day pulse using α³²P-UTP followed by washes and chase with cold UTP. Total RNA extracted from cells growing exponentially (expo), stressed for 2h in PBS (stress) or allowed to recover for 2 h in normal media (recov) was separated on 8% polyacrylamide/urea gels. (D) Cells were induced for expression of a tagged tRNA followed by removal of the inducer from the culture media. RNA was extracted from cells growing exponentially (expo), after nutritional stress (stress) or after 1 or 2 h of recovery in normal media and the tagged RNA investigated by northern blot. As control the same treatment was applied to uninduced cells (left part).