The RNA Helicase Mtr4p Modulates Polyadenylation in the TRAMP Complex

Jia, Huijue; Wang, Xuying; Liu, Fei; Guenther, Ulf-Peter; Srinivasan, Sukanya; Anderson, James T.; Jankowsky, Eckhard

doi:10.1016/j.cell.2011.05.010

Cited by 93 publications

(120 citation statements)

References 37 publications

Supporting

Mentioning

115

Contrasting

Order By: Relevance

“…Of the 25 DEAD-box proteins in Saccharomyces cerevisiae, five (eIF4A, Fal1, Ded1, Dbp5, and Dbp8) are known to be regulated by cofactors (12,19,21,28,34,35,(65)(66)(67). Similarly, three related SF2-helicases from the DEAH and Ski2-like subfamilies (Prp43, Mtr4, and Brr2) are also regulated by cofactors (22,23,25,(68)(69)(70)(71). These cofactors can increase or decrease the rates of ATP hydrolysis, RNA unwinding, or phosphate or RNA release.…”

Section: Discussionmentioning

confidence: 99%

Cofactor-dependent specificity of a DEAD-box protein

Young

Khoshnevis

Karbstein

2013

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

View full text Add to dashboard Cite

DEAD-box proteins, a large class of RNA-dependent ATPases, regulate all aspects of gene expression and RNA metabolism. They can facilitate dissociation of RNA duplexes and remodeling of RNA-protein complexes, serve as ATP-dependent RNA-binding proteins, or even anneal duplexes. These proteins have highly conserved sequence elements that are contained within two RecA-like domains; consequently, their structures are nearly identical. Furthermore, crystal structures of DEAD-box proteins with bound RNA reveal interactions exclusively between the protein and the RNA backbone. Together, these findings suggest that DEAD-box proteins interact with their substrates in a nonspecific manner, which is confirmed in biochemical experiments. Nevertheless, this contrasts with the need to target these enzymes to specific substrates in vivo. Using the DEAD-box protein Rok1 and its cofactor Rrp5, which both function during maturation of the small ribosomal subunit, we show here that Rrp5 provides specificity to the otherwise nonspecific biochemical activities of the Rok1 DEAD-domain. This finding could reconcile the need for specific substrate binding of some DEAD-box proteins with their nonspecific binding surface and expands the potential roles of cofactors to specificity factors. Identification of helicase cofactors and their RNA substrates could therefore help define the undescribed roles of the 19 DEAD-box proteins that function in ribosome assembly.RNA helicases | annealing D EAD-box proteins are RNA-binding ATPases that are involved in all aspects of RNA metabolism: translation initiation, pre-mRNA splicing, mRNA export and decay, and ribosome biogenesis. In these processes, their functions include RNA duplex unwinding, RNA-protein complex remodeling, RNA duplex annealing, and ATP-dependent RNA binding (1-5).In vivo, these enzymes have specific functions that often involve the recognition of specific RNAs and the discrimination against a myriad of nonsubstrates. Nevertheless, only DbpA, a DEAD-box protein involved in bacterial ribosome assembly (6-8), has sequence-specific ATPase activity (6, 9), which arises from unique sequences in its C terminus (10). Such sequence specificity has not been demonstrated for any other DEAD-box protein and is consistent with their highly conserved structures and the almost universal conservation of residues that contact the RNA. Furthermore, analysis of the structures of RNA-bound DEAD-box proteins reveals that RNA contacts are made with the sugarphosphate backbone (11-17), largely precluding sequence-specific interactions between DEAD-box proteins and RNA.Many DEAD-box and related DEAH-box proteins function in conjunction with cofactors (ref. 18 and references therein). These cofactors can regulate the activity of DEAD-box or DEAH-box proteins by stimulating or inhibiting their ATPase, helicase, RNAbinding, or nucleotide-binding activities. Interestingly, cofactors are often RNA-binding proteins (19)(20)(21)(22)(23)(24)(25). We and others therefore postulated that these cofactors could als...

show abstract

Section: Discussionmentioning

confidence: 99%

Cofactor-dependent specificity of a DEAD-box protein

Young

Khoshnevis

Karbstein

2013

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

View full text Add to dashboard Cite

show abstract

“…Trf4-mediated poly(A) addition is also involved in the turnover of other types of aberrant transcripts (Kadaba et al 2006). Turnover requires Mtr4, a RNA-dependent helicase (Wang et al 2008;Jia et al 2011), and other proteins comprising the TRAMP complex including the two poly(A) polymerases, Trf4 and Trf5, which have overlapping as well as nonoverlapping substrate specificities (San Paolo et al 2009), and either Air1 or Air2, RNA binding proteins also with both overlapping and nonoverlapping specificities (Schmidt et al 2012). The activated substrate-containing TRAMP complex interacts with the nuclear exosome that contains two nucleases, Rrp6 and Rrp44, and numerous other proteins (Parker 2012).…”

Section: -59 Exonucleolytic Degradation By the Nuclear Exosomementioning

confidence: 99%

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

2013

View full text Add to dashboard Cite

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

show abstract

“…Moreover, Mtr4p controls the lengths of poly(A) tails appended by TRAMP (9). This role of Mtr4p requires ATP, but does not involve unwinding (9).…”

mentioning

confidence: 99%

“…The TRAMP complex consists of three subunits that are highly conserved in eukaryotes: a noncanonical poly(A) polymerase (Trf4p or Trf5p in Saccharomyces cerevisiae), a Zn-knuckle protein (Air2p or Air1p), and the RNA helicase Mtr4p (4)(5)(6)(7). TRAMP assists RNA degradation by the nuclear exosome, most notably by appending short (∼4-5 nt) oligo(A) tails at the 3′ ends of RNAs slated for exosome-mediated degradation (4,5,(8)(9)(10). In addition, it has been speculated that TRAMP enables the nuclear exosome to efficiently degrade structured RNA through unwinding activity associated with Mtr4p (1, 2, 11-13).…”

mentioning

confidence: 99%

RNA unwinding by the Trf4/Air2/Mtr4 polyadenylation (TRAMP) complex

Jia

Wang

Anderson

et al. 2012

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

Self Cite

View full text Add to dashboard Cite

Many RNA-processing events in the cell nucleus involve the Trf4/ Air2/Mtr4 polyadenylation (TRAMP) complex, which contains the poly(A) polymerase Trf4p, the Zn-knuckle protein Air2p, and the RNA helicase Mtr4p. TRAMP polyadenylates RNAs designated for processing by the nuclear exosome. In addition, TRAMP functions as an exosome cofactor during RNA degradation, and it has been speculated that this role involves disruption of RNA secondary structure. However, it is unknown whether TRAMP displays RNA unwinding activity. It is also not clear how unwinding would be coordinated with polyadenylation and the function of the RNA helicase Mtr4p in modulating poly(A) addition. Here, we show that TRAMP robustly unwinds RNA duplexes. The unwinding activity of Mtr4p is significantly stimulated by Trf4p/Air2p, but the stimulation of Mtr4p does not depend on ongoing polyadenylation. Nonetheless, polyadenylation enables TRAMP to unwind RNA substrates that it otherwise cannot separate. Moreover, TRAMP displays optimal unwinding activity on substrates with a minimal Mtr4p binding site comprised of adenylates. Our results suggest a model for coordination between unwinding and polyadenylation activities by TRAMP that reveals remarkable synergy between helicase and poly(A) polymerase.he Trf4/Air2/Mtr4 polyadenylation (TRAMP) complex is involved in nuclear RNA surveillance, 3′-end processing of rRNA, small nucleolar RNAs (snoRNAs), and snRNAs, and in gene silencing and chromatin maintenance (1-3). The TRAMP complex consists of three subunits that are highly conserved in eukaryotes: a noncanonical poly(A) polymerase (Trf4p or Trf5p in Saccharomyces cerevisiae), a Zn-knuckle protein (Air2p or Air1p), and the RNA helicase Mtr4p (4-7). TRAMP assists RNA degradation by the nuclear exosome, most notably by appending short (∼4-5 nt) oligo(A) tails at the 3′ ends of RNAs slated for exosome-mediated degradation (4,5,(8)(9)(10). In addition, it has been speculated that TRAMP enables the nuclear exosome to efficiently degrade structured RNA through unwinding activity associated with Mtr4p (1, 2, 11-13).However, RNA helicase activity has not been demonstrated for TRAMP. Though Mtr4p alone has been shown to unwind RNA duplexes in vitro (12, 13), it is not known whether binding of Trf4p/Air2p abolishes, decreases, increases, or otherwise alters this helicase activity. This question is important for TRAMP function, because Mtr4p and Trf4p operate with opposite polarities. Mtr4p only unwinds duplexes with a 3′ unpaired region, i.e., with a 3′ to 5′ polarity (12, 13). Trf4p polyadenylates the 3′ end of RNA and thus possesses 5′ to 3′ polarity (4, 5). How unwinding and polyadenylation with opposite polarities are coordinated in one complex is not readily apparent.Moreover, Mtr4p controls the lengths of poly(A) tails appended by TRAMP (9). This role of Mtr4p requires ATP, but does not involve unwinding (9). Instead, Mtr4p binds to the 3′ end of the RNA, detects the number of 3′-terminal adenosines, and, in response, modulates ATP affinity and adenylatio...

show abstract

The RNA Helicase Mtr4p Modulates Polyadenylation in the TRAMP Complex

Cited by 93 publications

References 37 publications

Cofactor-dependent specificity of a DEAD-box protein

Cofactor-dependent specificity of a DEAD-box protein

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

RNA unwinding by the Trf4/Air2/Mtr4 polyadenylation (TRAMP) complex

Contact Info

Product

Resources

About