Role of D-aminoacyl-tRNA deacylase beyond chiral proofreading as a cellular defense against glycine mischarging by AlaRS

Pawar, Komal Ishwar; Suma, Katta; Seenivasan, Ayshwarya; Kuncha, Santosh Kumar; Routh, Satya Brata; Kruparani, Shobha P; Sankaranarayanan, Rajan

doi:10.7554/elife.24001

Cited by 32 publications

(59 citation statements)

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“…Although AlaRS misactivates both glycine and serine, misactivation of serine appears to have more serious consequences, perhaps due to the presence of D-aminoacyl-tRNA deacylases which act on Gly-tRNA Ala 12, 13, 14 . We previously showed that sticky ( sti ) mutant mice that have a point mutation (Ala734Glu) in the editing domain of AlaRS have ubiquitinated protein aggregates in cerebellar Purkinje cells and subsequent degeneration of these neurons 9 .…”

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confidence: 99%

ANKRD16 prevents neuron loss caused by an editing-defective tRNA synthetase

Terrey

Lee

et al. 2018

Nature

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Editing domains of aminoacyl tRNA synthetases correct tRNA charging errors to maintain translational fidelity. A mutation in the editing domain of alanyl tRNA synthetase (AlaRS) in Aarssti mutant mice resulted in an increased production of serine-mischarged tRNAAla and degeneration of cerebellar Purkinje cells. By positional cloning, we identified Ankrd16, which acts epistatically with the Aarssti mutation to attenuate neurodegeneration. ANKRD16, a vertebrate-specific, ankyrin repeat-containing protein, binds directly to the catalytic domain of AlaRS. Serine misactivated by AlaRS is captured by lysine side chains of ANKRD16, preventing the charging of serine adenylates to tRNAAla and precluding serine misincorporation in nascent peptides. Deletion of Ankrd16 in the Aarssti/sti brain causes widespread protein aggregation and neuron loss. These results identify a novel amino acid-accepting co-regulator of tRNA synthetase editing as a new layer of the machinery essential for preventing severe pathologies that arise from defects in editing.

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confidence: 99%

ANKRD16 prevents neuron loss caused by an editing-defective tRNA synthetase

Terrey

Lee

et al. 2018

Nature

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“…In addition to proteinogenic amino acids, misaminoacylation of tRNAs with non-protein amino acids is also monitored by cis - and trans -editing factors. For example, D-aminoacyl-tRNA deacylases target and hydrolyse both D- and L-aminoacyl-tRNAs and in doing so prevent misincorporation of a wide range of amino acids during protein synthesis 42–44 .…”

Section: Mechanisms Of Translational Fidelity and Errormentioning

confidence: 99%

Translational fidelity and mistranslation in the cellular response to stress

2017

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Faithful translation of mRNA into the corresponding polypeptide is a complex multistep process, requiring accurate amino acid selection, transfer RNA (tRNA) charging and mRNA decoding on the ribosome. Key players in this process are aminoacyl-tRNA synthetases (aaRSs), which not only catalyse the attachment of cognate amino acids to their respective tRNAs, but also selectively hydrolyse incorrectly activated non-cognate amino acids and/or misaminoacylated tRNAs. This aaRS proofreading provides quality control checkpoints that exclude non-cognate amino acids during translation, and in so doing helps to prevent the formation of an aberrant proteome. However, despite the intrinsic need for high accuracy during translation, and the widespread evolutionary conservation of aaRS proofreading pathways, requirements for translation quality control vary depending on cellular physiology and changes in growth conditions, and translation errors are not always detrimental. Recent work has demonstrated that mistranslation can also be beneficial to cells, and some organisms have selected for a higher degree of mistranslation than others. The aims of this Review Article are to summarize the known mechanisms of protein translational fidelity and explore the diversity and impact of mistranslation events as a potentially beneficial response to environmental and cellular stress.

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“…Glycylation of tRNA Gly and mutants of tRNA Gly (U73A, U73G, U73C, U73A/C70U) were done using T. thermophilus glycyl-tRNA synthetase, while tRNA Ala was glycylated using E. coli AlaRS as explained in Pawar et al, 2017. Deacylation assays and EF-Tu protection experiments were performed according to the protocol explained in Pawar et al, 2017. k obs values were derived by fitting the data points to the curve corresponding to following first-order exponential decay: [ S t ] = [ S 0 ]e − kt , where [ S t ] is the concentration of substrate at time t , [ S 0 ] is the initial substrate concentration, and k ( k obs ) is the first-order decay constant.…”

Section: Methodsmentioning

confidence: 99%

“…coli MG1655 strain, thereby creating dtd -null strain in AlaRS editing-defective background (i.e. Δ dtd Δ alaS ) (Pawar et al, 2017). sfGFP and sfGFP-G67A, cloned in pBAD18 vector, were under the control of an arabinose-inducible promoter.…”

Section: Methodsmentioning

confidence: 99%

“… Abstract D-aminoacyl-tRNA deacylase (DTD) acts on achiral glycine, in addition to D-amino acids, attached to tRNA. We have recently shown that this activity enables DTD to clear non-cognate Gly-tRNA Ala with 1000-fold higher efficiency than its activity on Gly-tRNA Gly , indicating tRNA-based modulation of DTD (Pawar et al, 2017). Here, we show that tRNA’s discriminator base predominantly accounts for this activity difference and is the key to selection by DTD.…”

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A discriminator code-based DTD surveillance ensures faithful glycine delivery for protein biosynthesis in bacteria

Sankaranarayanan

Kuncha

Suma

et al. 2018

Preprint

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10D-aminoacyl-tRNA deacylase (DTD) acts on achiral glycine, in addition to D-amino acids, 11 attached to tRNA. We have recently shown that this activity enables DTD to clear non-cognate 12 Gly-tRNA Ala with 1000-fold higher efficiency than its activity on Gly-tRNA Gly , indicating 13 tRNA-based modulation of DTD (Pawar et al., 2017). Here, we show that tRNA's 14 discriminator base predominantly accounts for this activity difference and is the key to 15 selection by DTD. Accordingly, the uracil discriminator base, serving as a negative 16 determinant, prevents Gly-tRNA Gly misediting by DTD and this protection is augmented by 17 EF-Tu. Intriguingly, eukaryotic DTD has inverted discriminator base specificity and uses only 18 G3•U70 for tRNA Gly/Ala discrimination. Moreover, DTD prevents alanine-to-glycine 19 misincorporation in proteins rather than only recycling mischarged tRNA Ala . Overall, the study 20 reveals the unique co-evolution of DTD and discriminator base, "reciprocally" in Bacteria and 21 Eukarya, and suggests DTD's strong selection pressure on bacterial tRNA Gly s to retain a 22 pyrimidine discriminator code. 23 24 Quality control during translation of the genetic code involves multiple stages and a multitude 25 of proofreading factors (Guo and Schimmel, 2012; Ibba and Söll, 2000; Ogle and 26 Ramakrishnan, 2005). A compromise in editing leads to serious pathologies including 27 neurodegeneration in mouse, and even cell death (Bacher et al., 2005; Bullwinkle et al., 2014; 28 Karkhanis et al., 2007; Korencic et al., 2004; Lee et al., 2006; Liu et al., 2014; Lu et al., 29 2014; Moghal et al., 2016; Nangle et al., 2002; Roy et al., 2004). Among these proofreading 30 factors, D-aminoacyl-tRNA deacylase (DTD) is the one that specifically decouples wrongly 31 acylated D-amino acids from tRNAs (Calendar and Berg, 1967; Soutourina et al., 1999, 32 2000). Our studies have shown that DTD is an RNA-based catalyst that uses an invariant Gly-33 cisPro motif as a "chiral selectivity filter" to achieve substrate chiral specificity only through 34 rejection of L-amino acid from the active site, thereby leading to Gly-tRNA Gly misediting 35 (Ahmad et al., 2013; Routh et al., 2016; Routh and Sankaranarayanan, 2017). Recently, 36we have also shown that DTD's activity on achiral glycine helps in clearing Gly-tRNA Ala , a 37 misaminoacylation product of alanyl-tRNA synthetase (AlaRS), thus resolving a long-standing 38 question in translational quality control (Pawar et al., 2017). 39 As DTD acts on multiple tRNAs charged with D-amino acids or glycine, tRNA's role 40 in modulating DTD's activity was previously thought to be inconsequential (Calendar and 41 Berg, 1967). However, our recent work has shown that DTD's activity on Gly-tRNA Ala is about 42 1000-fold higher than on Gly-tRNA Gly , clearly demonstrating the profound effect of tRNA 43 elements on DTD and suggesting an underlying tRNA code for DTD's action. G3•U70, the 44 universal tRNA Ala -specific determinant for AlaRS, is also a determinant for DTD. Ho...

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Role of D-aminoacyl-tRNA deacylase beyond chiral proofreading as a cellular defense against glycine mischarging by AlaRS

Cited by 32 publications

References 58 publications

ANKRD16 prevents neuron loss caused by an editing-defective tRNA synthetase

ANKRD16 prevents neuron loss caused by an editing-defective tRNA synthetase

Translational fidelity and mistranslation in the cellular response to stress

A discriminator code-based DTD surveillance ensures faithful glycine delivery for protein biosynthesis in bacteria

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