Co-transcriptional gene regulation in eukaryotes and prokaryotes

Shine, Morgan; Gordon, Jackson; Schärfen, Leonard; Zigackova, Dagmar; Herzel, Lydia; Neugebauer, Karla M.

doi:10.1038/s41580-024-00706-2

Cited by 16 publications

(3 citation statements)

References 267 publications

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“…In bacteria, transcription is well known to be coupled with translation, so that ribosomes engage the nascent RNA while being generated by RNA polymerase (RNAP). Similarly, a large body of genetic, biochemical, cellular, and structural evidence indicates that eukaryotic transcription is coupled to RNA processing (5′ and 3′), as first shown for mRNA 3′ end processing ( Whitelaw and Proudfoot 1986 ) and subsequently in splicing, surveillance, and export ( Bentley 2014 ; Shine et al 2024 ). Thus, in eukaryotes, as the RNA emerges from the RNAP, it is bound by different factors to produce a protein-coated export-competent RNA molecule (the mRNP particle).…”

Section: Rna Biogenesis Processing and Export Factors As Barriers To ...mentioning

confidence: 87%

RNA biogenesis and RNA metabolism factors as R-loop suppressors: a hidden role in genome integrity

Luna,

Gómez-González,

Aguilera

2024

Genes Dev.

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Genome integrity relies on the accuracy of DNA metabolism, but as appreciated for more than four decades, transcription enhances mutation and recombination frequencies. More recent research provided evidence for a previously unforeseen link between RNA and DNA metabolism, which is often related to the accumulation of DNA–RNA hybrids and R-loops. In addition to physiological roles, R-loops interfere with DNA replication and repair, providing a molecular scenario for the origin of genome instability. Here, we review current knowledge on the multiple RNA factors that prevent or resolve R-loops and consequent transcription–replication conflicts and thus act as modulators of genome dynamics.

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Section: Rna Biogenesis Processing and Export Factors As Barriers To ...mentioning

confidence: 87%

RNA biogenesis and RNA metabolism factors as R-loop suppressors: a hidden role in genome integrity

Luna,

Gómez-González,

Aguilera

2024

Genes Dev.

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“…In eukaryotes as well, the Sen1/senataxin factor has been proposed to function much like Rho in bacteria in mediating both transcription termination and prevention of R-loop occurrence from nascent transcripts (118)(119)(120)(121)(122). More generally, it is being increasingly recognized that various cotranscriptional processes are important for gene regulation in both prokaryotes and eukaryotes (123).…”

Section: Discussionmentioning

confidence: 99%

Pathological R-loops in bacteria from engineered expression of endogenous antisense RNAs whose synthesis is ordinarily terminated by Rho

Pandiyan,

Mallikarjun,

Maheshwari

et al. 2024

Preprint

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In many bacteria, the essential factors Rho and NusG mediate termination of synthesis of nascent transcripts (including antisense RNAs) which are not being simultaneously translated. It has been proposed that in Rho's absence toxic RNA-DNA hybrids (R-loops) may be generated from nascent untranslated transcripts; and genome-wide mapping studies inEscherichia colihave identified putative loci of R-loop formation from more than 100 endogenous antisense transcripts that are synthesized only in a Rho-deficient strain. Here we provide evidence that engineered expression in wild-typeE. coliof several such individual antisense regions on a plasmid or the chromosome generates R-loops that, in an RNase H-modulated manner, serve to disrupt genome integrity. Rho inhibition was associated with increased prevalence of antisense R-loops also inXanthomonas oryzaepv.oryzaeandCaulobacter crescentus. Our results confirm the essential role of Rho in several bacterial genera for prevention of toxic R-loops from pervasive yet cryptic endogenous antisense transcripts. Engineered antisense R-looped regions may be useful for studies on both site-specific impediments to bacterial chromosomal replication and the mechanisms of their resolution.

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“…After identifying an appropriate initiator codon, 80S assembly, polypeptide elongation, termination, and ribosome disassembly proceed (6). The cap-dependent nature and linear directional scanning of the preinitiation complex typically prioritizes the first start codon encountered, limiting the recognition of potential 3 downstream ORFs (7,8). Consequently, eukaryotic nuclear genes have traditionally been considered monocistronic; encoding mature mRNAs that contain a single protein-coding ORF.…”

Section: Introductionmentioning

confidence: 99%

Leaky ribosomal scanning enables tunable translation of bicistronic ORFs in green algae

Duenas,

Craig,

Gallaher

et al. 2024

Preprint

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Advances in sequencing technology have unveiled examples of nucleus-encoded polycistronic genes, once considered rare. Exclusively polycistronic transcripts are prevalent in green algae, although the mechanism by which multiple polypeptides are translated from a single transcript is unknown. Here, we used bioinformatic and in vivo mutational analyses to evaluate competing mechanistic models for polycistronic expression in green algae. High-confidence manually curated datasets of bicistronic loci from two divergent green algae,Chlamydomonas reinhardtiiandAuxenochlorella protothecoides, revealed 1) a preference for weak Kozak-like sequences for ORF 1 and 2) an underrepresentation of potential initiation codons before ORF 2, which are suitable conditions for leaky scanning to allow ORF 2 translation. We used mutational analysis inAuxenochlorella protothecoidesto test the mechanism. In vivo manipulation of the ORF 1 Kozak-like sequence and start codon altered reporter expression at ORF 2, with a weaker Kozak-like sequence enhancing expression and a stronger one diminishing it. A synthetic bicistronic dual reporter demonstrated inversely adjustable activity of green fluorescent protein expressed from ORF 1 and luciferase from ORF 2, depending on the strength of the ORF 1 Kozak-like sequence. Our findings demonstrate that translation of multiple ORFs in green algal bicistronic transcripts is consistent with episodic leaky ribosome scanning of ORF 1 to allow translation at ORF 2. This work has implications for the potential functionality of upstream open reading frames found across eukaryotic genomes and for transgene expression in synthetic biology applications.Significance StatementTextbook dogma states that nucleus-encoded genes are monocistronic, producing transcripts with a single translated open reading frame. However, highly conserved bicistronic loci are pervasive in green algae that are separated by several hundred million years of evolution, speaking to their ancestral origins and functions within the Chlorophyte lineage. A combination of bioinformatic analysis and in vivo gene manipulation supports leaky ribosomal scanning as the primary mechanism for translation of multiple ORFs from bicistronic transcripts. We have successfully tuned synthesis levels of two proteins encoded on one mRNA by modifying the ORF 1 Kozak-like sequence. These findings may have broad applications in synthetic biology.

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Co-transcriptional gene regulation in eukaryotes and prokaryotes

Cited by 16 publications

References 267 publications

RNA biogenesis and RNA metabolism factors as R-loop suppressors: a hidden role in genome integrity

RNA biogenesis and RNA metabolism factors as R-loop suppressors: a hidden role in genome integrity

Pathological R-loops in bacteria from engineered expression of endogenous antisense RNAs whose synthesis is ordinarily terminated by Rho

Leaky ribosomal scanning enables tunable translation of bicistronic ORFs in green algae

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