RBOH-dependent signaling is involved in He-Ne laser-induced salt tolerance and production of rosmarinic acid and carnosol in Salvia officinalis.

Background: In the past two decades, the impacts of Helium-Neon (He-Ne) laser on stress resistance and secondary metabolism in plants have been studied, but the signaling pathway which by laser regulates this process remains unclear. Therefore, the current study sought to explore the role of RBOH-dependent signaling in He-Ne laser-induced salt tolerance and elicitation of secondary metabolism in Salvia officinalis. Seeds were primed with He-Ne laser (6 J cm^- 2) and peroxide hydrogen (H₂O₂, 5 mM) and 15-old-day plants were exposed to two salinity levels (0, 75 mM NaCl).

Results: Salt stress reduced growth parameters, chlorophyll content and relative water content (RWC) and increased malodialdehyde (MDA) and H₂O₂ contents in leaves of 45-old-day plants. After 48 h of salt exposure, higher transcription levels of RBOH (encoding NADPH oxidase), PAL (phenylalanine ammonia-lyase), and RAS (rosmarinic acid synthase) were recorded in leaves of plants grown from seeds primed with He-Ne laser and/or H₂O₂. Despite laser up-regulated RBOH gene in the early hours of exposing to salinity, H₂O₂ and MDA contents were lower in leaves of these plants after 30 days. Seed pretreatment with He-Ne laser and/or H₂O₂ augmented the accumulation of anthocyanins, total phenol, carnasol, and rosmarinic acid and increased total antioxidant capacity under non-saline and more extensively at saline conditions. Indeed, these treatments improved RWC, and K⁺/Na⁺ ratio, enhanced the activities of superoxide dismutase and ascorbate peroxidase and proline accumulation, and significantly decreased membrane injury and H₂O₂ content in leaves of 45-old-day plants under salt stress. However, applying diphenylene iodonium (DPI as an inhibitor of NADPH oxidase) and N, N-dimethyl thiourea (DMTU as a H₂O₂ scavenger) after laser priming reversed the aforementioned effects which in turn resulted in the loss of laser-induced salt tolerance and secondary metabolism.

Conclusions: These findings for the first time deciphered that laser can induce a transient RBOH-dependent H₂O₂ burst, which might act as a downstream signal to promote secondary metabolism and salt stress alleviation in S. officinalis plants.