TY - JOUR
T1 - NAD+ capping of sibD transcripts in E. coli is mediated by its minimal promoter and enhanced by ppGpp
AU - Liu, Wuzhen
AU - Hu, Kefan
AU - Nie, Shiqi
AU - Zhang, Feng
AU - Li, Qiongfang
AU - Zhang, Hailei
AU - Liang, Shumin
AU - Peng, Jialin
AU - Zhang, Shoudong
AU - Cai, Zongwei
AU - Xia, Yiji
N1 - This study was supported by the Research Grants Council of Hong Kong through the following grants to Yiji Xia: GRF grants 12102220, 12102022, 12103425, CRS grant CRS_HKBU201/22, CRF grant 2003-22WF, and AoE grant AoE/M-402/25-N. We thank Dr George Church for depositing the plasmidpJ251-GERC (plasmid ID: #47441) to Addgene, which provides the sfGFP gene used in this study.
Publisher Copyright:
© The Author(s) 2026. Published by Oxford University Press.
PY - 2026/2/24
Y1 - 2026/2/24
N2 - Recently, nicotinamide adenine dinucleotide (NAD+) and other nucleotide analogs have been identified as non-canonical RNA caps in both prokaryotes and eukaryotes. In Escherichia coli, NAD capping has been shown to be influenced by environmental conditions in a gene-specific manner, yet its regulatory mechanisms remain poorly understood. We previously reported that most transcripts produced by sibD are NAD-capped during the stationary phase. In this study, we found that the 35-bp minimal promoter of sibD is sufficient for its NAD capping. When this minimal promoter was applied to express genes not typically producing NAD-RNAs, their transcripts could also be NAD-capped. These findings strongly support that NAD capping in E. coli occurs during transcription initiation mediated by the promoter and RNA polymerase. Additionally, the bacterial alarmone ppGpp and the small protein DksA, both transcription initiation regulators, synergistically enhance transcription of both NAD-capped and uncapped RNAs from sibD and its homologous genes, sibC and sibE, in vitro. In contrast, the ppGpp⁰ mutant, deficient in ppGpp synthesis, showed a significant reduction in NAD-RNA production from these genes. Our findings elucidate the cis-elements and trans-acting factors mediating NAD capping during transcription initiation, offering novel insight into this regulatory process.
AB - Recently, nicotinamide adenine dinucleotide (NAD+) and other nucleotide analogs have been identified as non-canonical RNA caps in both prokaryotes and eukaryotes. In Escherichia coli, NAD capping has been shown to be influenced by environmental conditions in a gene-specific manner, yet its regulatory mechanisms remain poorly understood. We previously reported that most transcripts produced by sibD are NAD-capped during the stationary phase. In this study, we found that the 35-bp minimal promoter of sibD is sufficient for its NAD capping. When this minimal promoter was applied to express genes not typically producing NAD-RNAs, their transcripts could also be NAD-capped. These findings strongly support that NAD capping in E. coli occurs during transcription initiation mediated by the promoter and RNA polymerase. Additionally, the bacterial alarmone ppGpp and the small protein DksA, both transcription initiation regulators, synergistically enhance transcription of both NAD-capped and uncapped RNAs from sibD and its homologous genes, sibC and sibE, in vitro. In contrast, the ppGpp⁰ mutant, deficient in ppGpp synthesis, showed a significant reduction in NAD-RNA production from these genes. Our findings elucidate the cis-elements and trans-acting factors mediating NAD capping during transcription initiation, offering novel insight into this regulatory process.
UR - https://www.scopus.com/pages/publications/105030898806
U2 - 10.1093/nar/gkag102
DO - 10.1093/nar/gkag102
M3 - Journal article
C2 - 41732914
SN - 0305-1048
VL - 54
JO - Nucleic Acids Research
JF - Nucleic Acids Research
IS - 5
M1 - gkag102
ER -