The temperature-regulated DEAD-box RNA helicase CrhR interactome: autoregulation and photosynthesis-related transcripts

Abstract

RNA helicases play crucial functions in RNA biology. In plants, RNA helicases are encoded by large gene families, performing roles in abiotic stress responses, development, the post-transcriptional regulation of gene expression as well as house-keeping functions. Several of these RNA helicases are targeted to the organelles, mitochondria and chloroplasts. Cyanobacteria are the direct evolutionary ancestors of plant chloroplasts. The cyanobacterium Synechocystis 6803 encodes a single DEAD-box RNA helicase, CrhR, that is induced by a range of abiotic stresses, including low temperature. Though the ΔcrhR mutant exhibits a severe cold-sensitive phenotype, the physiological function(s) performed by CrhR have not been described. To identify transcripts interacting with CrhR, we performed RNA co-immunoprecipitation with extracts from a Synechocystis crhR deletion mutant expressing the FLAG-tagged native CrhR or a K57A mutated version with an anticipated enhanced RNA binding. The composition of the interactome was strikingly biased towards photosynthesis-associated and redox-controlled transcripts. A transcript highly enriched in all experiments was the crhR mRNA, suggesting an auto-regulatory molecular mechanism. The identified interactome explains the described physiological role of CrhR in response to the redox poise of the photosynthetic electron transport chain and characterizes CrhR as an enzyme with a diverse range of transcripts as molecular targets.

Multiple Light-Dark Signals Regulate Expression of the DEAD-Box RNA Helicase CrhR in Synechocystis PCC 6803

“…Overall, CrhR appears to play a role in maintaining homeostasis of photosynthesis in response to conditions resulting in over-reduction of the ETC. This crucial conclusion is supported by the results presented here combined with previous physiological and omics analysis, which suggest that CrhR regulates translation of protein products required to adapt to environmental conditions that perturb ETC homeostasis [ 11 , 25 , 27 , 35 ].…”

“…These results support the conclusion that a phytochrome-mediated mechanism regulates the post-transcriptional accumulation of CrhR protein via an auto-regulatory process ( Figure 9 b). This conclusion was supported by the observation that the crhR transcript is a major target for CrhR binding [ 35 ]. Together, the data suggest that a potentially conserved phytochrome-mediated mechanism imparts translational control in response to light signals in cyanobacteria, with CrhR performing an active role in the process.…”

AtpΘ is an inhibitor of F0F1 ATP synthase to arrest ATP hydrolysis during low-energy conditions in cyanobacteria

“…CuSO 4 (2 mM) was used to induce the expression of the Cu 2+ -responsive petE promoter, 51 while the petJ promoter was induced by removing Cu 2+ from the medium through centrifugation and resuspension. For high-density cultivation used for ATP synthase purification, Synechocystis 6803 overexpressing P petE -atpB-3xFLAG was cultured in the cell-DEG system as reported previously 52 using freshwater medium 53 with the following modifications: Na 2 EDTA and CuSO 4 were not included in the medium, and 10 mg/mL kanamycin or 5 mg/mL gentamicin was added.…”

A plastidial DEAD box RNA helicase plays a critical role in high light acclimation by modulating ribosome biogenesis inChlamydomonas reinhardtii

“…CrhR , the unique DEAD-box RNA helicase present in the genome of the cyanobacterium Synechocystis sp. PCC 6803, participates in redox regulation of gene expression (Kujat and Owttrim, 2000), low temperature acclimation of photosynthesis (Rosana et al, 2012; Sireesha et al, 2012), and was recently shown to interact with transcripts associated with photosynthesis (Migur et al, 2021). In plant chloroplasts, DEAD-box RNA helicases participate in ribosome biogenesis (Asakura et al, 2012; Chi et al, 2012), chloroplast differentiation (Wang et al, 2000), and responses to diverse abiotic stresses (Nawaz and Kang, 2017), but a role of these enzymes in the light-response of eukaryotic photosynthesis has not been reported so far.…”