scholarly journals Four translation initiation pathways employed by the leaderless mRNA in eukaryotes

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Kseniya A. Akulich ◽  
Dmitry E. Andreev ◽  
Ilya M. Terenin ◽  
Victoria V. Smirnova ◽  
Aleksandra S. Anisimova ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Leaderless Mrna

scholarly journals ΔGunfold leaderless, a package for high-throughput analysis of translation initiation regions (TIRs) at the transcriptome scale and for leaderless mRNA optimization

2021 ◽  
Author(s):  
Mohammed Husain Bharmal ◽  
Jared M Schrader
Keyword(s):  
Gene Expression ◽  
Binding Site ◽  
High Throughput ◽  
Translation Initiation ◽  
High Throughput Analysis ◽  
Translation Initiation Region ◽  
Throughput Analysis ◽  
Ribosome Binding ◽  
Leaderless Mrna ◽  
Essential Step

Translation initiation is an essential step for fidelity of gene expression, in which the ribosome must bind to the translation initiation region (TIR) and position the initiator tRNA in the P-site (1). For this to occur correctly, the TIR encompassing the ribosome binding site (RBS) needs to be highly accessible (2-5). ΔGunfold is a metric for computing accessibility of the TIR, but there is no automated way to compute it manually with existing software/tools limiting throughput. ΔGunfold leaderless allows users to automate the ΔGunfold calculation to perform high-throughput analysis. Importantly, ΔGunfold leaderless allows calculation of TIRs of both leadered mRNAs and leaderless mRNAs which lack a 5' UTR and which are abundant in bacterial, archaeal, and mitochondrial transcriptomes (4, 6, 7). The ability to analyze leaderless mRNAs also allows one additional feature where users can computationally optimize leaderless mRNA TIRs to maximize their gene expression (8, 9). The ΔGunfold leaderless package facilitates high-throughput calculations of TIR accessibility, is designed to calculate TIR accessibility for leadered and leaderless mRNA TIRs which are abundant in bacterial/archaeal/organellar transcriptomes and allows optimization of leaderless mRNA TIRs for biotechnology.

scholarly journals A combination of mRNA features influence the efficiency of leaderless mRNA translation initiation

2021 ◽  
Vol 3 (3) ◽  
Author(s):  
Mohammed-Husain M Bharmal ◽  
Alisa Gega ◽  
Jared M Schrader
Keyword(s):  
16S Rrna ◽  
Translation Initiation ◽  
Untranslated Region ◽  
Mrna Translation ◽  
Start Codon ◽  
Leaderless Mrna ◽  
Shine Dalgarno Sequence ◽  
Relationship Of ◽  
Bacterial Translation

Abstract Bacterial translation is thought to initiate by base pairing of the 16S rRNA and the Shine–Dalgarno sequence in the mRNA’s 5′ untranslated region (UTR). However, transcriptomics has revealed that leaderless mRNAs, which completely lack any 5′ UTR, are broadly distributed across bacteria and can initiate translation in the absence of the Shine–Dalgarno sequence. To investigate the mechanism of leaderless mRNA translation initiation, synthetic in vivo translation reporters were designed that systematically tested the effects of start codon accessibility, leader length, and start codon identity on leaderless mRNA translation initiation. Using these data, a simple computational model was built based on the combinatorial relationship of these mRNA features that can accurately classify leaderless mRNAs and predict the translation initiation efficiency of leaderless mRNAs. Thus, start codon accessibility, leader length, and start codon identity combine to define leaderless mRNA translation initiation in bacteria.

scholarly journals Evidence against an Interaction between the mRNA Downstream Box and 16S rRNA in Translation Initiation

2001 ◽  
Vol 183 (11) ◽  
pp. 3499-3505 ◽  
Author(s):  
Isabella Moll ◽  
Michael Huber ◽  
Sonja Grill ◽  
Pooneh Sairafi ◽  
Florian Mueller ◽  
...  
Keyword(s):  
16S Rrna ◽  
Translation Initiation ◽  
Base Pair ◽  
Translational Efficiency ◽  
Coding Region ◽  
Reporter Construct ◽  
Leaderless Mrna ◽  
30S Subunit ◽  
Downstream Box ◽  
Ribosomal P

ABSTRACT Based on the complementarity of the initial coding region (downstream box [db]) of several bacterial and phage mRNAs to bases 1469 to 1483 in helix 44 of 16S rRNA (anti-downstream box [adb]), it has been proposed that db-adb base pairing enhances translation in a way that is similar to that of the Shine-Dalgarno (SD)/anti-Shine-Dalgarno (aSD) interaction. Computer modeling of helix 44 on the 30S subunit shows that the topography of the 30S ribosome does not allow a simultaneous db-adb interaction and placement of the initiation codon in the ribosomal P site. Thus, the db-adb interaction cannot substitute for the SD-aSD interaction in translation initiation. We have always argued that any contribution of the db-adb interaction should be most apparent on mRNAs devoid of an SD sequence. Here, we show that 30S ribosomes do not bind to leaderless mRNA in the absence of initiator tRNA, even when the initial coding region shows a 15-nucleotide complementarity (optimal fit) with the putative adb. In addition, an optimized db did not affect the translational efficiency of a leaderless λ cI-lacZ reporter construct. Thus, the db-adb interaction can hardly serve as an initial recruitment signal for ribosomes. Moreover, we show that different leaderless mRNAs are translated in heterologous systems although the sequence of the putative adb's within helix 44 of the 30S subunits of the corresponding bacteria differ largely. Taken our data together with those of others (M. O'Connor, T. Asai, C. L. Squires, and A. E. Dahlberg, Proc. Natl. Acad. Sci. USA 96:8973–8978, 1999; A. La Teana, A. Brandi, M. O'Connor, S. Freddi, and C. L. Pon, RNA 6:1393–1402, 2000), we conclude that the db does not base pair with the adb.

scholarly journals A combinatorial model predicts leaderless mRNA start codon selection in C. crescentus

2020 ◽  
Author(s):  
Mohammed-Husain M. Bharmal ◽  
Jared M. Schrader
Keyword(s):  
Translation Initiation ◽  
Mrna Translation ◽  
Start Codon ◽  
Leaderless Mrna ◽  
Combinatorial Model ◽  
Shine Dalgarno Sequence ◽  
Codon Selection ◽  
Relationship Of ◽  
Bacterial Translation

AbstractBacterial translation is thought to initiate by base-pairing of the 16S rRNA and the Shine-Dalgarno sequence in the mRNA’s 5’ UTR. However, transcriptomics has revealed that leaderless mRNAs, which completely lack any 5’ UTR, are broadly distributed across bacteria and can initiate translation in the absence of the Shine-Dalgarno sequence. To investigate the mechanism of leaderless mRNA translation initiation, synthetic in vivo translation reporters were designed that systematically tested the effects of start codon accessibility, leader length, and start codon identity on leaderless mRNA translation initiation. Using this data, a simple computational model was built based on the combinatorial relationship of these mRNA features which can accurately classify leaderless mRNAs and predict the translation initiation efficiency of leaderless mRNAs. Thus, start codon accessibility, leader length, and start codon identity combine to define leaderless mRNA translation initiation in bacteria.

scholarly journals Isolation and Characterization of Ribosomes and Translation Initiation Factors from the Gram-Positive Soil Bacterium Streptomyces lividans

2004 ◽  
Vol 186 (20) ◽  
pp. 6864-6875 ◽  
Author(s):  
J. Michael Day ◽  
Gary R. Janssen
Keyword(s):  
Translation Initiation ◽  
Ternary Complex ◽  
Streptomyces Lividans ◽  
Ternary Complexes ◽  
Binding Assays ◽  
Ribosomal Subunits ◽  
Initiation Factors ◽  
Leaderless Mrna ◽  
Isolation And Characterization ◽  
Translation Initiation Factors

ABSTRACT A primer extension inhibition (toeprint) assay was developed using ribosomes and ribosomal subunits from Streptomyces lividans. This assay allowed the study of ribosome binding to streptomycete leaderless and leadered mRNA. Purified 30S subunits were unable to form a ternary complex on aph leaderless mRNA, whereas 70S ribosomes could form ternary complexes on this mRNA. 30S subunits formed ternary complexes on leadered aph and malE mRNA. The translation initiation factors (IF1, IF2, and IF3) from S. lividans were isolated and included in toeprint and filter binding assays with leadered and leaderless mRNA. Generally, the IFs reduced the toeprint signal on leadered mRNA; however, incubation of IF1 and IF2 with 30S subunits that had been washed under high-salt conditions promoted the formation of a ternary complex on aph leaderless mRNA. Our data suggest that, as reported for Escherichia coli, initiation complexes with leaderless mRNAs might use a novel pathway involving 70S ribosomes or 30S subunits bound by IF1 and IF2 but not IF3. Some mRNA-ribosome-initiator tRNA reactions that yielded weak or no toeprint signals still formed complexes in filter binding assays, suggesting the occurrence of interactions that are not stable in the toeprint assay.

scholarly journals A Leaderless mRNA Can Bind to Mammalian 80S Ribosomes and Direct Polypeptide Synthesis in the Absence of Translation Initiation Factors

2006 ◽  
Vol 26 (8) ◽  
pp. 3164-3169 ◽  
Author(s):  
Dmitri E. Andreev ◽  
Ilya M. Terenin ◽  
Yan E. Dunaevsky ◽  
Sergei E. Dmitriev ◽  
Ivan N. Shatsky
Keyword(s):  
Translation Initiation ◽  
Striking Similarity ◽  
Alternative Mode ◽  
80S Ribosome ◽  
Eukaryotic Translation ◽  
Initiation Factors ◽  
Leaderless Mrna ◽  
Translation Initiation Factors ◽  
Polypeptide Synthesis ◽  
Multistep Process

ABSTRACT Translation initiation in eukaryotic cells is known to be a complex multistep process which involves numerous protein factors. Here we demonstrate that leaderless mRNAs with initiator Met-tRNA can bind directly to 80S mammalian ribosomes in the absence of initiation factors and that the complexes thus formed are fully competent for the subsequent steps of polypeptide synthesis. We show that the canonical 48S pathway of eukaryotic translation initiation has no obvious advantage over the 80S pathway of translation initiation on leaderless mRNAs and suggest that, in the presence of competing mRNAs containing a leader, the latter mechanism will be preferred. The direct binding of the leaderless mRNA to the 80S ribosome was precluded when such an mRNA was supplied with a 5′ leader, irrespective of whether it was in a totally single-stranded conformation or was prone to base pairing. The striking similarity between the mechanisms of binding of leaderless mRNAs with mammalian 80S or bacterial 70S ribosomes gives support to the idea that the alternative mode of translation initiation used by leaderless mRNAs represents a relic from early steps in the evolution of the translation apparatus.

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