THE EFFECTS OF UV-A RADIATION ON CIRCADIAN RHYTHMS IN SYNECHOCOCCUS ELONGATUS UTEX 2973

Cyanobacteria are among the simplest organisms to display circadian rhythms that synchronize endogenous physiological activities with a ~12-hour-light:12-hour-dark (12L:12D) cycle of the external environment. Detected by the input pathway composed of CikA and LdpA proteins, light is transduced to the central circadian oscillator encoded by the gene cluster kaiABC. While KaiC phosphorylation is primarily regulated by KaiA and KaiB proteins, two key components of the output pathway, RpaA and SasA proteins, mediate between KaiC phosphorylation, genome-wide expression, and control of cell division. In this study, Synechococcus elongatus UTEX 2973 showed similar growth patterns when subjected to white light only and white light supplemented with ultraviolet A (UV-A) radiation under 12L:12D intervals, although UV-A radiation hindered growth during light periods. Under continuous illumination, growth rates of S. elongatus UTEX 2973 were reduced by UV-A radiation but reflected intrinsic circadian rhythmicity. To elucidate the critical role of the circadian clock, a mutant void of kaiABC was generated via the CRISPR/Cpf1 system. A dysfunctional clock severely disrupted inherent growth rhythmicity, which was exacerbated by UV-A radiation. To investigate the effects of UV-A radiation on transcription patterns in S. elongatus UTEX 2973, expression levels of circadian genes, specifically kaiABC, cikA, lpdA, rpaA, and sasA, were assessed by qPCR analysis. For the UV-A-treated wild-type strain, kaiA and kaiB expression was generally downregulated, kaiC expression was upregulated during the second dark period, and rpaA expression was either upregulated or downregulated depending on the period. For the UV-A-treated ΔkaiABC strain, lpaA expression was upregulated in darkness, whereas rpaA and sasA expression was downregulated during light periods. When ΔkaiABC and wild-type strains were examined in the presence and absence of UV-A radiation, expression of lpaA, rpaA, and sasA was universally downregulated, yet cikA expression was upregulated in the dark. This study was the first to evaluate the impact of UV-A radiation on cyanobacterial circadian rhythms, in which UV-A radiation negatively affected cyanobacterial growth and strongly altered gene expression patterns over time. Without the circadian clock, rhythmicity of growth and transcription was demolished, such that the consequences were aggravated for the output pathway that relayed signals downstream from the central oscillator.