A riboswitch gives rise to multi-generational phenotypic heterogeneity in an auxotrophic bacterium

Abstract

Auxotrophy, the inability to produce an organic compound essential for growth, is widespread among bacteria. Auxotrophic bacteria rely on transporters to acquire these compounds from their environment. Here, we study the expression of both low-and high-affinity transporters of the costly amino acid methionine in an auxotrophic lactic acid bacterium, Lactococcus lactis. We show that the high-affinity transporter (Met-transporter) is heterogeneously expressed at low methionine concentrations, resulting in two isogenic subpopulations that sequester methionine in different ways: one subpopulation primarily relies on the high-affinity transporter (high expression of the Met-transporter) and the other subpopulation primarily relies on the low-affinity transporter (low expression of the Met-transporter). The phenotypic heterogeneity is remarkably stable, inherited for tens of generations, and apparent at the colony level. This heterogeneity results from a T-box riboswitch in the promoter region of the met operon encoding the high-affinity Met-transporter. We hypothesize that T-box riboswitches, which are commonly found in the Lactobacillales, may play as-yet unexplored roles in the predominantly auxotrophic lifestyle of these bacteria.

Development of Lactococcus lactis Biosensors for Detection of Sulfur-Containing Amino Acids

“…This dose-curve shows that Δ met provides a linear range of growth output in the range of approximately 0.5–5 mM. Figure 1B (right scheme) illustrates the only available path for methionine uptake in this strain; the higher methionine requirements in this strain are explained by the low affinity of BcaP for methionine and its competition with the branched-chain amino acids to be taken up via this transporter ( Hernandez-Valdes et al, 2020c ).…”

“…The auxotrophic nature of L. lactis for methionine makes it an attractive bacterial host to design growth-based biosensors to detect this amino acid. When methionine is available in the environment, it can be transported by two uptake systems that have been described previously: an ABC transporter (Met) and the branched-chain amino acid permease (BcaP) ( Trip et al, 2013 ; Hernandez-Valdes et al, 2020c ). We used a wild-type L. lactis (WTmet) strain and measured the culture cell densities, when growing in chemically defined medium (CDM-met) supplemented with different methionine concentrations (0.0004–20 mM).…”

“…Hernandez-Valdes et al, 2020a met L. lactis MG1363 met deletion mutant. Hernandez-Valdes et al, 2020c WTcys Ery r , MG1363 derivative, llmg_pseudo10:P cys -gfp. This study…”

Riboswitch RS thiT as a Molecular Tool in Lactococcus lactis

“…Of the three 5′ UTRs in genes/operons involved in thiamine biosynthesis in L. lactis , the one upstream of thiT is confirmed here to be a TPP riboswitch (RS thiT ). RS thiT is the fourth confirmed riboswitch in L. lactis and the first to be used as a molecular tool ( 30 – 32 ). Bacterial TPP riboswitches control their downstream genes by either inducing transcription termination, inhibiting translation initiation, or both, while in eukaryotes, they operate by promoting alternative splicing ( 33 , 34 ).…”

Lifestyle, metabolism and environmental adaptation inLactococcus lactis

“…Nevertheless, recent studies underpin that our understanding of amino acid import can still be refined. This is exemplified by the recent discovery of the co-existence of two isogenic L. lactis subpopulations that depend on either high-or low-affinity import systems of the essential amino acid methionine, which is controlled by a riboswitch present in the promoter region of the genes encoding the high-affinity importer (Hernandez-Valdes, van Gestel and Kuipers 2020). Similarly, the transport-coordinating role of the peculiar presence of two distinct and competing substrate binding domains in the L. lactis GlnPQ importer for asparagine, glutamine and glutamic acid was only recently unravelled (Gouridis et al 2015;Fulyani et al 2016;Schuurman-Wolters et al 2018).…”