The effect of UV-B radiation on flavonoid accumulation and gene expression in Marchantia polymorpha

Abstract

Marchantia polymorpha is the closest living representation of the earliest land plants and as such may give insight into the response that allow the first land plants to transition from the aquatic environment onto the terrestrial. One such adaption allowing this transition was to that of UV-B radiation and is the main focus of this study. This thesis investigates the effect of UV-B radiation on the basic land plant M. polymorpha and its responce through the production of secondary metabolites, in particular flavonoids. How flavonoid compounds may protect M. polymorpha and the gene regulation that contributes towards stress related defense and secondary compound production is also investigated.

M. polymorpha plants were subject to UV-B fluence at two seperate levels to stimulate both an acclimation and stress related response. Plants were then measured for the production of UV-B absorbing compounds, in particular the flavonoids, and also the localisation in thallus tissue of such compounds. The genetic response was also analysed using a combination of RNA-seq and nCounter techniques. Specific mutant plants with altered flavonoid amounts were also analysed to help further determine the genetic interactions that contribute to protection.

M. polymorpha was found to respond to UV-B through enhanced levels of UV-B absorbing compounds and in particular flavonoids, of which the flavones were the most predominant. Apigenin-based flavones were found in the highest amounts followed by luteolin-based flavones. The ratio of Apigenin to Luteolin-based flavones was seen to shift towards luteolin in high UV-B responses. Apigenin was suggested to be required for UV-B screening functions under low fluence while high fluence treatments, that induce ROS may require Luteolin based flavones that are able to scavenge ROS more effectively. Under low fluence conditions flavone compounds accumulated strongly in the epidermal layers suggesting a strong UV-B screening function.

RNA-seq revealed that M. polymorpha responds to UV-B through the UVR8 pathway similar to that of higher plants. We determined the UVR8 pathway in M. polymorpha contains the main regulators COP1, HY5 and RUP1 and act to enhance flavonoid biosynthetic genes such as CHS, CHI and FNS. RNA-seq and nCounter analysis also revealed differential expression in stress related genes that may offer protection independently of the UVR8 pathway and flavonoids. Mutant lines with altered flavonoid amounts were also tested under UV-B irradiance and showed that lower flavones concentrations resulted in enhanced damage to plant thallus. Elevated flavones resulted in enhanced protection and a reduction in thallus damage and loss of flavones altogether resulted in severe stunting in plants grown under UV-B.

Overall we determined that M. polymorpha responds through UVR8 for the production of flavone compounds and this production results in protection against the deleterous effects of UV-B. Such adaptive mechanisms likely assisted early land plants to transition from aquatic to land environments by reducing UV-B burden.