Gut colonization by Bacteroides requires translation by an EF‐G paralog lacking GTPase activity

Abstract

Protein synthesis is crucial for cell growth and survival yet one of the most energy‐consuming cellular processes. How, then, do cells sustain protein synthesis under starvation conditions when energy is limited? To accelerate the translocation of mRNA–tRNAs through the ribosome, bacterial elongation factor G (EF‐G) hydrolyzes energy‐rich guanosine triphosphate (GTP) for every amino acid incorporated into a protein. Here, we identify an EF‐G paralog—EF‐G2—that supports translocation without hydrolyzing GTP in the gut commensal bacterium Bacteroides thetaiotaomicron. EF‐G2's singular ability to sustain protein synthesis, albeit at slow rates, is crucial for bacterial gut colonization. EF‐G2 is ~10‐fold more abundant than canonical EF‐G1 in bacteria harvested from murine ceca and, unlike EF‐G1, specifically accumulates during carbon starvation. Moreover, we uncover a 26‐residue region unique to EF‐G2 that is essential for protein synthesis, EF‐G2 dissociation from the ribosome, and responsible for the absence of GTPase activity. Our findings reveal how cells curb energy consumption while maintaining protein synthesis to advance fitness in nutrient‐fluctuating environments.

An integrated transcriptomics–functional genomics approach reveals a small RNA that modulatesBacteroides thetaiotaomicronsensitivity to tetracyclines

“…2b), encoding the canonical EF-G and not belonging to the BT_4338 regulon (Townsend et al, 2020). This corroborates previous reports (Han et al, 2022; Schwalm et al, 2016; Townsend et al, 2020) and further supports that B. thetaiotaomicron utilizes a distinct protein synthesis machinery during colonization of a sugar-deprived host niche.…”

“…Complementing protein-mediated transcriptional control, environmental and pathogenic bacteria employ small regulatory RNAs (sRNAs) (Wagner and Romby, 2015). Via binding to partially complementary sequences within target mRNAs, these riboregulators can posttranscriptionally modulate gene expression to help bacteria adapt their metabolism to the available nutrients (Bobrovskyy and Vanderpool, 2013) and cope with noxious stimuli (Holmqvist and Wagner, 2017). For example, several sRNAs are involved in antibiotic stress responses in phylogenetically unrelated pathogens, proposing these molecules as relevant drug targets to treat infectious diseases (Dersch et al, 2017;Lalaouna et al, 2014;Mediati et al, 2021).…”

Dietary sugars silence the master regulator of carbohydrate utilization in human gut Bacteroides species

“…We hypothesized that BT4338 controls Roc amounts by regulating transcription of an unknown factor(s) involved in its synthesis. The most highly induced BT4338 -dependent gene during carbon limitation conditions is fusA2 ( BT2167 ), which encodes a non-essential, alternative translation elongation factor G (EF-G2) that enables GTP-independent translation 40 and is critical for mammalian intestinal colonization. 27 Because BT4338 is required for synthesis of both EF-G2 and Roc, and BT4338-dependent control of Roc protein is indirect and mediated via the roc 5’ mRNA leader, we reasoned that BT4338-dependent EF-G2 expression may control Roc amounts by governing its translation in a leader-dependent manner.…”

“…We establish that BT4338 indirectly controls Roc amounts via the roc mRNA leader, which is necessary for and sufficient to confer BT4338 -dependent control to a heterologous gene. Furthermore, we show that BT4338 governs Roc abundance independently of an alternative translation elongation factor, EF-G2 40 , whose mRNA levels are regulated by BT4338 and increase dramatically during carbon limitation. 27 Finally, we demonstrate that glucose and fructose exert rapid, dramatic, and dominant silencing of BT4338-dependent genes in vitro and that these effects are consistent across distinct gut Bacteroides species.…”

Protein degradation by human 20S proteasomes elucidates the interplay between peptide hydrolysis and splicing

“…The final protein concentration was adjusted to 0.3 mM. Recombinant CaM, EF-G, EF-Ts, Ffh, H2A, H2B, H3, H4, HUWE1, IF2, LEDGF, LRP130, PDF, RF1, UbcH7, Ube2K, and Ube2S were prepared according to previously published protocols 112 , 116 – 123 .…”