Abstract
Oxygenic photosynthetic organisms evolved mechanisms for thermal dissipation of energy absorbed in excess to prevent formation of reactive oxygen species. The major and fastest component, called non-photochemical quenching, occurs within the photosystem II antenna system by the action of two essential light-harvesting complex (LHC)-like proteins, photosystem II subunit S (PSBS) in plants and light-harvesting complex stress-related (LHCSR) in green algae and diatoms. In the evolutionary intermediate Physcomitrella patens, a moss, both gene products are active. These proteins, which are present in low amounts, are difficult to purify, preventing structural and functional analysis. Here, we report on the overexpression of the LHCSR1 protein from P. patens in the heterologous systems Nicotiana benthamiana and Nicotiana tabacum using transient and stable nuclear transformation. We show that the protein accumulated in both heterologous systems is in its mature form, localizes in the chloroplast thylakoid membranes, and is correctly folded with chlorophyll a and xanthophylls but without chlorophyll b, an essential chromophore for plants and algal LHC proteins. Finally, we show that recombinant LHCSR1 is active in quenching in vivo, implying that the recombinant protein obtained is a good material for future structural and functional studies.
Highlights
light-harvesting complex stressrelated (LHCSR) protein in algae and mosses is essential for non-photochemical quenching (NPQ)
In Physcomitrella patens, LHCSR is responsible for most of the quenching activity [21], which is strongly enhanced by zeaxanthin binding [22], making LHCSR the ideal system for structure-function studies aimed at elucidating the properties of this molecular switch that regulates photon use efficiency in unicellular algae and mosses resuming the two functions of pH detection and quenching of the chlorophyll excited states [14, 20]
Expression of P. patens LHCSR1 in N. benthamiana and N. tabacum Plants—The full-length ORFs of LHCSR1 with or without thrombin-cleavable histidine tail (His tag) at the C terminus of the protein were introduced by LR recombination into the binary GATEWAY vector pK7WG2 under the control of the constitutive cauliflower mosaic virus 35S promoter
Summary
LHCSR protein in algae and mosses is essential for NPQ. Results: Expression and characterization of Physcomitrella patens LHCSR1 protein upon heterologous expression in N. benthamiana and N. tabacum was obtained. Forward genetic analysis showed that qE requires one of two lhc-like gene products, PSBS and LHCSR, respectively, in plants and algae These are essential for sensing pH of the thylakoid lumen [13, 14] and transduction into quenching events. In Physcomitrella patens, LHCSR is responsible for most of the quenching activity [21], which is strongly enhanced by zeaxanthin binding [22], making LHCSR the ideal system for structure-function studies aimed at elucidating the properties of this molecular switch that regulates photon use efficiency in unicellular algae and mosses resuming the two functions of pH detection and quenching of the chlorophyll excited states [14, 20]. We show that LHCSR is active in quenching in vivo, suggesting that the recombinant product obtained closely features the protein responsible for NPQ in mosses, providing a solid basis for future structural and functional studies
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
