Active segregation of yeast mitochondria by Myo2 is essential and mediated by Mmr1 and Ypt11.

Active segregation of essential organelles is required for successful cell division. The essential budding yeast myosin V Myo2 actively segregates most organelles along polarized actin cables. The mechanism of mitochondrial segregation remains controversial, with movement driven by actin polymerization, movement driven by association with transported cortical endoplasmic reticulum (ER), and direct transport by Myo2 proposed as models.

The yeast rapid tRNA decay pathway competes with elongation factor 1A for substrate tRNAs and acts on tRNAs lacking one or more of several modifications.

The structural and functional integrity of tRNA is crucial for translation. In the yeast Saccharomyces cerevisiae, certain aberrant pre-tRNA species are subject to nuclear surveillance, leading to 3' exonucleolytic degradation, and certain mature tRNA species are subject to rapid tRNA decay (RTD) if they are appropriately hypomodified or bear specific destabilizing mutations, leading to 5'-3' exonucleolytic degradation by Rat1 and Xrn1. Thus, trm8-Δ trm4-Δ strains are temperature sensitive due to lack of m(7)G(46) and m(5)C and the consequent RTD of tRNA(Val(AAC)), and tan1-Δ trm44-Δ strains are temperature sensitive due to lack of ac(4)C(12) and Um(44) and the consequent RTD of tRNA(Ser(CGA)) and tRNA(Ser(UGA)).

The yeast rapid tRNA decay pathway primarily monitors the structural integrity of the acceptor and T-stems of mature tRNA.

tRNAs, like other RNAs, are subject to quality control steps during and after biosynthesis. We previously described a rapid tRNA degradation (RTD) pathway in which the 5'-3' exonucleases Rat1 and Xrn1 degrade mature tRNA(Val(AAC)) in yeast mutants lacking m(7)G and m(5)C, and mature tRNA(Ser(CGA)) in mutants lacking Um and ac(4)C. To understand how the RTD pathway selects substrate tRNAs among different tRNAs lacking the same modifications, we used a genetic screen to examine tRNA(Ser(CGA)) variants.

Chapter 11. Identification and analysis of tRNAs that are degraded in Saccharomyces cerevisiae due to lack of modifications.

Mounting evidence shows that tRNA modifications play crucial roles in the maintenance of wild-type levels of several tRNA species. This chapter describes a generalized framework in which to study tRNA turnover in the yeast Saccharomyces cerevisiae as a consequence of a defect in tRNA modification status. It describes several approaches for the identification of tRNA species that are reduced as a consequence of a modification defect, methods for analysis of the rate of tRNA loss and analysis of its aminoacylation, and methods for initial characterization of tRNA turnover.

Degradation of several hypomodified mature tRNA species in Saccharomyces cerevisiae is mediated by Met22 and the 5'-3' exonucleases Rat1 and Xrn1.

Mature tRNA is normally extensively modified and extremely stable. Recent evidence suggests that hypomodified mature tRNA in yeast can undergo a quality control check by a rapid tRNA decay (RTD) pathway, since mature tRNA(Val(AAC)) lacking 7-methylguanosine and 5-methylcytidine is rapidly degraded and deacylated at 37 degrees C in a trm8-Delta trm4-Delta strain, resulting in temperature-sensitive growth. We show here that components of this RTD pathway include the 5'-3' exonucleases Rat1 and Xrn1, and Met22, which likely acts indirectly through Rat1 and Xrn1.

Rapid tRNA decay can result from lack of nonessential modifications.

The biological role of many nonessential tRNA modifications outside of the anticodon remains elusive despite their evolutionary conservation. We show here that m7G46 methyltransferase Trm8p/Trm82p acts as a hub of synthetic interactions with several tRNA modification enzymes, resulting in temperature-sensitive growth. Analysis of three double mutants indicates reduced levels of tRNA(Val(AAC)), consistent with a role of the corresponding modifications in maintenance of tRNA levels.