P-body components, Dhh1 and Pat1, are involved in tRNA nuclear-cytoplasmic dynamics

Hurto, Rebecca L.; Hopper, Anita K.

doi:10.1261/rna.2558511

Cited by 17 publications

(30 citation statements)

References 48 publications

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“…To ensure that our results were dependent on the contribution of Mtr10 to retrograde tRNA transport and not on other Mtr10 functions, we also disrupted the retrograde pathway by a mechanism that is independent of a β-importin protein. Dhh1 and Pat1 affect retrograde nuclear accumulation, and neither is a member of the β-importin family (48). The 5′ endextended spliced tRNA Ile UAU accumulates in both mtr10Δ and dhh1Δ pat1Δ mutant strains in which retrograde tRNA nuclear accumulation is independently blocked (Fig.…”

Section: Resultsmentioning

confidence: 95%

Retrograde transfer RNA nuclear import provides a new level of tRNA quality control in Saccharomyces cerevisiae

Kramer

Hopper

2013

Proc. Natl. Acad. Sci. U.S.A.

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In eukaryotes, transfer RNAs (tRNAs) are transcribed in the nucleus yet function in the cytoplasm; thus, tRNA movement within the cell was believed to be unidirectional-from the nucleus to the cytoplasm. It is now known that mature tRNAs also move in a retrograde direction from the cytoplasm to the nucleus via retrograde tRNA nuclear import, a process that is conserved from yeast to vertebrates. The biological significance of this tRNA nuclear import is not entirely clear. We hypothesized that retrograde tRNA nuclear import might function in proofreading tRNAs to ensure that only proper tRNAs reside in the cytoplasm and interact with the translational machinery. Here we identify two major types of aberrant tRNAs in yeast: a 5′, 3′ end-extended, spliced tRNA and hypomodified tRNAs. We show that both types of aberrant tRNAs accumulate in mutant cells that are defective in tRNA nuclear traffic, suggesting that they are normally imported into the nucleus and are repaired or degraded. The retrograde pathway functions in parallel with the cytoplasmic rapid tRNA decay pathway previously demonstrated to monitor tRNA quality, and cells are not viable if they lack both pathways. Our data support the hypothesis that the retrograde process provides a newly discovered level of tRNA quality control as a pathway that monitors both end processing of pre-tRNAs and the modification state of mature tRNAs.tRNA retrograde pathway | tRNA processing errors | quality control pathways | Mtr10 | Los1 T ransfer RNAs (tRNAs) are well known for their role as adaptor molecules during the translation of mRNAs into proteins, delivering amino acids to the ribosome for incorporation into a polypeptide chain. In eukaryotes, tRNAs are transcribed in the nucleus as primary transcripts that undergo extensive processing, chemical modification, and subcellular trafficking to produce the mature tRNAs that function in cytoplasmic translation. In the yeast Saccharomyces cerevisiae, tRNA transcription in the nucleolus (1), with one exception (2), is followed by removal of the 5′ leader sequence by RNase P (3). The 3′ trailer is subsequently removed endonucleolytically, presumably by tRNase Z (4-6), and/or exonucleolytically by Rex1 (7,8). Following removal of the 3′ trailer, the nucleotides C 74 C 75 A 76 , necessary for tRNA aminoacylation, are added to the 3′ terminus. Twenty percent of yeast tRNAs are transcribed with an intron (9). Splicing of tRNA introns occurs in the cytoplasm in yeast, on the outer surface of mitochondria where the tRNA splicing endonuclease complex resides (10-12). Thus, end-processed intron-containing tRNAs are exported from the nucleus to the cytoplasm before removal of the intron. This primary export of intron-containing pre-tRNA is mediated by the β-importin family member Los1 (13-15). Because LOS1 is unessential and because it is essential to have tRNAs delivered to the cytoplasm, there is at least one additional unidentified nuclear exporter for end-processed intron-containing pre-tRNA (16).tRNAs are subject to numerous pos...

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Section: Resultsmentioning

confidence: 95%

Retrograde transfer RNA nuclear import provides a new level of tRNA quality control in Saccharomyces cerevisiae

Kramer

Hopper

2013

Proc. Natl. Acad. Sci. U.S.A.

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“…It is now known that tRNAs can be aminoacylated in the nucleus (Lund and Dahlberg 1998;Sarkar et al 1999;Grosshans et al 2000a), that tRNAs can traffic from the cytoplasm to the nucleus via the tRNA retrograde process (Shaheen and Hopper 2005;Takano et al 2005;Zaitseva et al 2006), and that tRNAs imported into the nucleus can be reexported to the cytoplasm (Whitney et al 2007;Eswara et al 2009) (Figure 3). tRNA subcellular traffic is conserved (Zaitseva et al 2006;Shaheen et al 2007;Barhoom et al 2011), responsive to nutrient availability (Shaheen and Hopper 2005;Hurto et al 2007;Shaheen et al 2007;Whitney et al 2007;Murthi et al 2010), and, in yeast, is coordinated with the formation of P-bodies in the cytoplasm (Hurto and Hopper 2011). In addition to the trafficking of tRNAs between the nucleus and the cytoplasm, some tRNAs encoded by the nucleus are imported into mitochondria (reviewed in Rubio and Hopper 2011;Schneider 2011).…”

Section: Trna Subcellular Traffickingmentioning

confidence: 99%

“…P-body formation is dependent upon Dhh1 and Pat1 as dhh1D pat1D cells lack P-bodies when deprived of glucose and cells overexpressing Dhh1 or Pat1 generate P-bodies in fed conditions (Parker 2012). Likewise, tRNAs fail to accumulate in nuclei in dhh1D pat1D cells subjected to glucose deprivation and constitutively accumulate in nuclei in cells overexpressing Dhh1 or Pat1 in fed conditions (Hurto and Hopper 2011). The results demonstrate that there is coordinate regulation between mRNA cytoplasmic dynamics and tRNA nuclear/cytoplasmic trafficking and selection for this coordination may be key to cell survival upon stress.…”

Section: Regulation Of the Trna Retrograde Processmentioning

confidence: 99%

Transfer RNA Post-Transcriptional Processing, Turnover, and Subcellular Dynamics in the YeastSaccharomyces cerevisiae

2013

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Transfer RNAs (tRNAs) are essential for protein synthesis. In eukaryotes, tRNA biosynthesis employs a specialized RNA polymerase that generates initial transcripts that must be subsequently altered via a multitude of post-transcriptional steps before the tRNAs beome mature molecules that function in protein synthesis. Genetic, genomic, biochemical, and cell biological approaches possible in the powerful Saccharomyces cerevisiae system have led to exciting advances in our understandings of tRNA post-transcriptional processing as well as to novel insights into tRNA turnover and tRNA subcellular dynamics. tRNA processing steps include removal of transcribed leader and trailer sequences, addition of CCA to the 39 mature sequence and, for tRNA His , addition of a 59 G. About 20% of yeast tRNAs are encoded by intron-containing genes. The three-step splicing process to remove the introns surprisingly occurs in the cytoplasm in yeast and each of the splicing enzymes appears to moonlight in functions in addition to tRNA splicing. There are 25 different nucleoside modifications that are added post-transcriptionally, creating tRNAs in which 15% of the residues are nucleosides other than A, G, U, or C. These modified nucleosides serve numerous important functions including tRNA discrimination, translation fidelity, and tRNA quality control. Mature tRNAs are very stable, but nevertheless yeast cells possess multiple pathways to degrade inappropriately processed or folded tRNAs. Mature tRNAs are also dynamic in cells, moving from the cytoplasm to the nucleus and back again to the cytoplasm; the mechanism and function of this retrograde process is poorly understood. Here, the state of knowledge for tRNA post-transcriptional processing, turnover, and subcellular dynamics is addressed, highlighting the questions that remain. TABLE OF CONTENTS Abstract 43Introduction 44

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“…Both Lsm1 and Pat1 are nucleocytoplasmic shuttling proteins that get exported by Crm1 (Marnef et al 2012;Haimovich et al 2013). Yeast Pat1 affects tRNA subcellular distribution dynamics in a manner that is different from that of Lsm1 (Hurto and Hopper 2011). Yeast Pat1 associates with topoisomerase-II and CEN DNA and has been implicated in rDNA locus stability and chromosome transmission fidelity (Wang et al 1996(Wang et al , 1999Mishra et al 2013).…”

Section: Introductionmentioning

confidence: 99%

Pat1 contributes to the RNA binding activity of the Lsm1-7–Pat1 complex

Chowdhury

Kalurupalle

Tharun

2014

RNA

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A major mRNA decay pathway in eukaryotes is initiated by deadenylation followed by decapping of the oligoadenylated mRNAs and subsequent 5 ′ -to-3 ′ exonucleolytic degradation of the capless mRNA. In this pathway, decapping is a rate-limiting step that requires the hetero-octameric Lsm1-7-Pat1 complex to occur at normal rates in vivo. This complex is made up of the seven Sm-like proteins, Lsm1 through Lsm7, and the Pat1 protein. It binds RNA and has a unique binding preference for oligoadenylated RNAs over polyadenylated RNAs. Such binding ability is crucial for its mRNA decay function in vivo. In order to determine the contribution of Pat1 to the function of the Lsm1-7-Pat1 complex, we compared the RNA binding properties of the Lsm1-7 complex purified from pat1Δ cells and purified Pat1 fragments with that of the wild-type Lsm1-7-Pat1 complex. Our studies revealed that both the Lsm1-7 complex and purified Pat1 fragments have very low RNA binding activity and are impaired in the ability to recognize the oligo(A) tail on the RNA. However, reconstitution of the Lsm1-7-Pat1 complex from these components restored these abilities. We also observed that Pat1 directly contacts RNA in the context of the Lsm1-7-Pat1 complex. These studies suggest that the unique RNA binding properties and the mRNA decay function of the Lsm1-7-Pat1 complex involve cooperation of residues from both Pat1 and the Lsm1-7 ring. Finally our studies also revealed that the middle domain of Pat1 is essential for the interaction of Pat1 with the Lsm1-7 complex in vivo.

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P-body components, Dhh1 and Pat1, are involved in tRNA nuclear-cytoplasmic dynamics

Cited by 17 publications

References 48 publications

Retrograde transfer RNA nuclear import provides a new level of tRNA quality control in Saccharomyces cerevisiae

Retrograde transfer RNA nuclear import provides a new level of tRNA quality control in Saccharomyces cerevisiae

Transfer RNA Post-Transcriptional Processing, Turnover, and Subcellular Dynamics in the YeastSaccharomyces cerevisiae

Pat1 contributes to the RNA binding activity of the Lsm1-7–Pat1 complex

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