A singular PpaA/AerR-like protein in Rhodospirillum rubrum rules beyond the boundaries of photosynthesis in response to the intracellular redox state

Abstract

Photosynthesis (PS) in purple non-Sulfur bacteria (PNSB) is strictly controlled to avoid energy consumption and to prevent oxidative stress when oxygen is present. PpsR, a transcriptional repressor encoded in the photosynthesis gene cluster (PGC), is one of the main players in the regulation of PS-related genes. In some cases, PpsR does not sense the environmental cue by itself but delegates this task to other proteins like PpaA/AerR anti-repressors, which can ultimately affect PpsR affinity to some promoter regions. In this work, the effects of locus Rru_A0625 product (HP1) on PS regulation of Rhodospirillum rubrum were studied by mutation/complementation and transcriptomic strategies. Rru_A0625 is located next to ppsR gene, just like other PpaA/AerR members, and its deletion annuls pigment synthesis in dark micro/anaerobic growth conditions. HP1 shows similarity to PpaA/AerR anti-repressors family, although it does not possess their typical cobalamin binding domain. A transcriptomic analysis of Rru_A0625 deletion mutant showed that HP1 not only has effects on bacteriochlorophyll and carotenoid biosynthesis, but also many other biological processes in the cell. The most notorious is the impact on the transcription of the nitrogenase complex components and accessory proteins. Our results suggest that this new member of the PpaA/AerR family has evolved losing the canonical cobalamin binding domain, but not the redox sensing capability, conserving a not fully understood mechanism of PS regulation. IMPORTANCE Rhodospirillum rubrum vast metabolic versatility places it as a remarkable model bacterium and an excellent biotechnological chassis. The key component of photosynthesis (PS) studied in this work (HP1) stands out among the other members of PpaA/AerR anti-repressor family since it lacks the motif they all share: the cobalamin B-12 binding motif. Despite being reduced and poorly conserved, HP1 stills controls PS as the other members of the family, allowing a fast response to changes in the redox state of the cell. This work also shows that HP1 absence affects genes from relevant biological processes other than PS, including nitrogen fixation and stress response. From a biotechnological perspective, HP1 could be manipulated in approaches where PS is not necessary, such as hydrogen or polyhydroxyalkanoates production, to save energy.

Exploring Rhodospirillum rubrum response to high doses of carbon monoxide under light and dark conditions

“…This locus codifies a particular version of the anti-repressor PpaA, which in PNSB modifies PpsR repressor activity. PpaA in R. rubrum has been recently proved to affect various biological processes related to adaptation to microarobiosis/anaerobiosis (Godoy et al 2023b ). Notably, the same mutation at the Rru_A0697 locus present in stain 1.4-2B was also identified in 1.7-3A.…”

“…We specifically refer to the three indels in G/C repetitive sequences impacting on loci Rru_A0933 and Rru_A1264 from strain 1.4-2B, Rru_A0625 from strain 1.7-3A, and Rru_A0697 from both strains. This hypothetical mechanism could be responsible for both the emergence of the spontaneous mutant R02_01 by annulling the Rru_A0625 locus (Godoy et al 2023b ) and restoring the same gene after the UVa-ALE protocol. This hints at the possibility that not all mutations present in both strains were induced solely by UV radiation, but may have arisen from other adaptive mechanisms during the mutagenesis experiment.…”

“…467T, here referred to as strain S1). Rr02_01 was isolated and characterized previously in our laboratory due to its impaired pigment production under low oxygen levels (Godoy et al 2023b ). Strains were stored at −80°C in Luria-Bertani (LB) medium supplemented with 15% v/v glycerol.…”

“…DNA samples were sent to MicrobesNG (Birmingham, UK) for genomic sequencing. The resulting reads, including those from the genome of strain Rr02_01 previously sequenced (Godoy et al 2023b ), were aligned and compared with the reference genome of the ATCC 11170 using Geneious Prime software. Discrepancies with the reference genome were filtered by the coverage value (CV>15 reads) and the frequency of variation (FV>70%), and corroborated by Sanger sequencing (Secugen®, Madrid) using primers listed in Table S 2 .…”