EGY3: homologue of S2P protease located in chloroplasts

Abstract

The EGY3 protein is a homologue of site‐2 proteases, which are intramembrane zinc metalloproteases. EGY3 itself lacks proteolytic activity due to the absence of a zinc‐binding motif. Plentiful evidence indicates that such intramembrane ‘pseudoproteases’ play significant roles in many diverse processes occurring within the cell. However, the physiological functions of EGY3, as well as its subcellular localization, remain unknown. The subcellular localization of EGY3 protein was investigated using Arabidopsis thaliana protoplasts transformed with EGY3‐GFP fusion protein, and immunoblot experiments using the total leaf protein extract, as well as highly purified chloroplasts and fractions of stroma, envelope and thylakoid membrane proteins. The physiological role of EGY3 was studied using two A. thaliana mutant lines devoid of EGY3 protein. Chlorophyll a fluorescence measurement was performed and the egy3 mutant sensitivity to photoinhibition was investigated. Additionally, the abundance of thylakoid membrane complexes was established using blue native gel electrophoresis. We present experimental evidence for thylakoid membrane localization of the EGY3 protein. We show that egy3 mutants display increased value of the non‐photochemical quenching parameter and significantly slower recovery rate after photoinhibitory treatment. This was associated with a decrease in the level of proteases involved in photosystem II recovery, Deg1 and FtsH2/8.

An Argon-Ion-Induced Pale Green Mutant of Arabidopsis Exhibiting Rapid Disassembly of Mesophyll Chloroplast Grana

“…A. thaliana also has two distant paralogs of EGY1 : EGY2 and EGY3 . All three Arabidopsis EGY proteins localize to the thylakoid membranes, and EGY1 and EGY2 (but not EGY3) possess ATP-independent proteolytic activities [ 17 , 73 , 74 , 75 , 76 ]. EGY2 was found to be involved in hypocotyl elongation [ 75 ], and EGY2 transcript levels were higher than those of EGY1 in several developmental and physiological conditions [ 77 ].…”

“…In the current study, Ar50-33-pg1 and egy1-4 plants exhibited accelerated chlorosis upon nitrogen deprivation ( Figure 10 ). EGY1 transcripts were abundantly detected in growing shoot tissues [ 18 , 76 ], and these mutant nutritional phenotypes might thus represent a secondary effect of the egy1 mutation. Nevertheless, EGY1 may be involved in the modulation of nitrogen and phosphate metabolism under conditions of plant nutritional stress.…”

EGY3 mediates chloroplastic ROS homeostasis and promotes retrograde signaling in response to salt stress in Arabidopsis

“…EGY2 encodes an intramembrane protease localized in the thylakoid membrane and affects chloroplast development (Adamiec et al, 2018). EGY3 encodes a ''pseudoprotease'' localized in chloroplasts (Adamiec et al, 2020); however, its biological functions, especially in response to salt stress, remain largely unknown.…”

The changes of PSII supercomplex stoichiometry in egy1 mutants are related to chlorophyll b deficiency

“…All other antibodies, namely Anti-Lhcb1, Anti-Lhcb2, Anti-Lhcb3, Anti-Lhcb4, Anti-Lhcb5, Anti-Lhcb6, anti-PsbA, anti-PsbC, anti-PsbD, FtsH2/8, and Deg1 antibodies are commercially available and were purchased from Agrisera (Vännäs, Sweden). For each primary antibody linearity of immunoresponse was determined previously (Adamiec et al 2018(Adamiec et al , 2020. The time-course photoinhibition and light stress recovery assays: The time-course analysis was performed according to Liu and Last (2017) and to our previous study (Adamiec et al 2018(Adamiec et al , 2020.…”

“…For each primary antibody linearity of immunoresponse was determined previously (Adamiec et al 2018(Adamiec et al , 2020. The time-course photoinhibition and light stress recovery assays: The time-course analysis was performed according to Liu and Last (2017) and to our previous study (Adamiec et al 2018(Adamiec et al , 2020. The detached leaves of the WT and mutant plants egy1-2 and egy1-3 were infiltrated with water or 1 mM lincomycin (Sigma-Aldrich, USA) to inhibit de novo synthesis of proteins encoded on the chloroplasts' genome (i.e., PsbA polypeptide) and exposed to the irradiance of white light of 900 μmol(photon) m -2 s -1 (white fluorescent light lamp Philips Master T-E-D 58 W/840 REFLEX Eco) for 5 h. The maximum quantum yield of the PSII parameter (Fv/Fm) was measured every hour after 30 min of leaf dark adaptation.…”