Bradyrhizobium japonicum HmuP is an RNA‐binding protein that positively controls hmuR operon expression by suppression of a negative regulatory RNA element in the 5′ untranslated region

• Published in: Molecular microbiology Vol. 121; no. 6; pp. 1217 - 1227
• Main Authors: Wu, Peipei, Ong, Alasteir, O'Brian, Mark R.
• Format: Journal Article
• Language: English
• Published: England Blackwell Publishing Ltd 01-06-2024
• Subjects: 5' Untranslated Regions; 5' Untranslated Regions - genetics; bacterial gene expression; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bradyrhizobium - genetics; Bradyrhizobium - metabolism; Bradyrhizobium japonicum; Clonal deletion; Down-regulation; Gene expression; Gene Expression Regulation, Bacterial; Heme; Heme - metabolism; heme transport; iron acquisition; Mutants; Operon - genetics; post‐transcriptional regulation; Promoter Regions, Genetic; Protein Binding; Protein structure; Proteins; Ribonucleic acid; RNA; RNA binding protein; RNA, Bacterial - genetics; RNA, Bacterial - metabolism; RNA-Binding Proteins - genetics; RNA-Binding Proteins - metabolism; Secondary structure; bacterial gene expression; post‐transcriptional regulation; heme transport; iron acquisition; RNA binding protein
• ISSN: 0950-382X; 1365-2958; 1365-2958
• DOI: 10.1111/mmi.15274

The hmuR operon encodes proteins for the uptake and utilization of heme as a nutritional iron source in Bradyrhizobium japonicum. The hmuR operon is transcriptionally activated by the Irr protein and is also positively controlled by HmuP by an unknown mechanism. An hmuP mutant does not express the hmuR operon genes nor does it grow on heme. Here, we show that hmuR expression from a heterologous promoter still requires hmuP, suggesting that HmuP does not regulate at the transcriptional level. Replacement of the 5′ untranslated region (5′UTR) of an HmuP‐independent gene with the hmuR 5′UTR conferred HmuP‐dependent expression on that gene. Recombinant HmuP bound an HmuP‐responsive RNA element (HPRE) within the hmuR 5′UTR. A 2 nt substitution predicted to destabilize the secondary structure of the HPRE abolished both HmuP binding activity in vitro and hmuR expression in cells. However, deletion of the HPRE partially restored hmuR expression in an hmuP mutant, and it rescued growth of the hmuP mutant on heme. These findings suggest that the HPRE is a negative regulatory RNA element that is suppressed when bound by HmuP to express the hmuR operon.
The hmuR operon encodes proteins needed for heme transport into cells. The 5′ untranslated region of the hmuR operon contains a negative regulatory RNA element (HPRE) that inhibits its expression. Binding of HmuP to the HPRE suppresses HPRE activity, resulting in expression of the hmuR operon.