The ability of Lactococcus lactis to grow under moderate oxygen conditions enables its expansion from the food industry to microbial cell factories.While leveraging known antioxidative regulatory mechanisms represents...The ability of Lactococcus lactis to grow under moderate oxygen conditions enables its expansion from the food industry to microbial cell factories.While leveraging known antioxidative regulatory mechanisms represents a promising strategy for engineering robust strains under high oxygen conditions,the complete genetic basis of oxygen tolerance in L.lactis remains incompletely understood.Here,we identified and characterized a novel antioxidative system and its corresponding regulatory mechanism in L.lactis N8.Genetic analyses demonstrated that RmaH,a MarR-family transcriptional regulator,is essential for mediating the oxidative stress response.Transcriptomic and qPCR analysis further revealed that RmaH acts as a transcriptional repressor of dps,which encodes a DNA-binding protein from starved cells(Dps).In vitro and in vivo protein-DNA binding assays confirmed that RmaH specifically binds to the coding sequence(CDS)of dps,with DNase I footprinting precisely identifying the binding motif(TGTAAG-12nt-CTTTCA).Finally,functional investigations revealed that under oxygen conditions,RmaH expression was suppressed,leading to upregulated dps expression.Dps thereby confers cellular protection via two distinct mechanisms:physically shielding DNA from hydroxyl radicals and inhibiting the Fenton reaction through its ferroxidase activity.Collectively,our findings elucidated a previously uncharacterized RmaH-Dps regulatory pathway that enhances oxygen tolerance in L.lactis,providing both mechanistic insight and a potential target for engineering industrially robust strains.展开更多
基金supported by the National Natural Science Foundation of China(No.31770102)the Shanxi Province Science Foundation for Youths(No.202203021222007).
文摘The ability of Lactococcus lactis to grow under moderate oxygen conditions enables its expansion from the food industry to microbial cell factories.While leveraging known antioxidative regulatory mechanisms represents a promising strategy for engineering robust strains under high oxygen conditions,the complete genetic basis of oxygen tolerance in L.lactis remains incompletely understood.Here,we identified and characterized a novel antioxidative system and its corresponding regulatory mechanism in L.lactis N8.Genetic analyses demonstrated that RmaH,a MarR-family transcriptional regulator,is essential for mediating the oxidative stress response.Transcriptomic and qPCR analysis further revealed that RmaH acts as a transcriptional repressor of dps,which encodes a DNA-binding protein from starved cells(Dps).In vitro and in vivo protein-DNA binding assays confirmed that RmaH specifically binds to the coding sequence(CDS)of dps,with DNase I footprinting precisely identifying the binding motif(TGTAAG-12nt-CTTTCA).Finally,functional investigations revealed that under oxygen conditions,RmaH expression was suppressed,leading to upregulated dps expression.Dps thereby confers cellular protection via two distinct mechanisms:physically shielding DNA from hydroxyl radicals and inhibiting the Fenton reaction through its ferroxidase activity.Collectively,our findings elucidated a previously uncharacterized RmaH-Dps regulatory pathway that enhances oxygen tolerance in L.lactis,providing both mechanistic insight and a potential target for engineering industrially robust strains.
