Functional Characterization of a Ferritin-Like Transcriptional Regulator in Cyanobacterial Cold Adaptation

• Knockout of either csrnf1 or its homolog csrn7 led to reduced cold resistance • Cross-complementation tests showed Csrn7 and Csrnf1 share similar functions • Transcriptomic analyses of the csrn7 mutant revealed potential target genes • Csrn7 could bind to the candidate target gene petE under cold stress • The csrn7 / petE double mutant had a greater reduction in ATP and NADPH levels Low-temperature stress is a major abiotic factor that negatively affects the growth of plants and algae. Cyanobacteria (also known as blue-green algae) exhibit remarkable adaptability to diverse thermal environments. In our previous transcriptomic analyses of the dryland cyanobacterium Nostoc flagelliforme , we identified a single-copy gene, csrnf1 , which encodes protein with a ferritin-like domain and is strongly responsive to cold stress. Heterologous expression of csrnf1 in the model cyanobacterium Nostoc sp. PCC 7120 enhanced its cold tolerance. However, the functional mechanisms of Csrnf1 and its homologs remains unclear. In this study, we constructed a csrnf1 knockout mutant (Δ csrnf1 ) of N. flagelliforme and a knockout of its homolog ( csrn7 ) in Nostoc sp. PCC 7120 (Δ csrn7 ). A functional cross-complementation assay verified the functional similarity and interchangeability between Csrnf1 and Csrn7 in both cyanobacterial strains. Furthermore, comparative transcriptomic analysis of Δ csrn7 and the wild-type (WT) Nostoc sp. PCC 7120, revealed a set of genes potentially regulated by Csrn7, including all0258 ( petE ), which encodes the photosynthetic electron transporter plastocyanin. Gel shift assay and in vivo GFP reporter analyses showed that Csrn7 binds to the promoter region of petE . Additionally, a double mutant (Δ csrn7 + Δ all0258 ) exhibited more pronounced reductions in ATP and NADPH levels and increased sensitivity to cold stress compared with the single mutants and WT. Thus, Csrnf1/Csrn7 may facilitate the maintenance of photosynthetic efficiency and energy homeostasis under cold stress. Together, our study reveals a previously unknown regulatory pathway in cyanobacterial cold adaptation through the transcriptional regulation of petE— a stably expressed gene encoding plastocyanin functioning in photosynthesis—by a novel ferritin-like protein.