The DnaJ-Like Zinc-Finger Protein HCF222 Is Required for Thylakoid Membrane Biogenesis in Plants

Abstract

To understand the biogenesis of the thylakoid membrane in higher plants and to identify auxiliary proteins required to build up this highly complex membrane system, we have characterized the allelic nuclear mutants high chlorophyll fluorescence222-1 (hcf222-1) and hcf222-2 and isolated the causal gene by map-based cloning. In the ethyl methanesulfonate-induced mutant hcf222-1, the accumulation of the cytochrome b 6 f (Cytb6f) complex was reduced to 30% compared with the wild type. Other thylakoid membrane complexes accumulated to normal levels. The T-DNA knockout mutant hcf222-2 showed a more severe defect with respect to thylakoid membrane proteins and accumulated only 10% of the Cytb6f complex, accompanied by a reduction in photosystem II, the photosystem II light-harvesting complex, and photosystem I. HCF222 encodes a protein of 99 amino acids in Arabidopsis (Arabidopsis thaliana) that has similarities to the cysteine-rich zinc-binding domain of DnaJ chaperones. The insulin precipitation assay demonstrated that HCF222 has disulfide reductase activity in vitro. The protein is conserved in higher plants and bryophytes but absent in algae and cyanobacteria. Confocal fluorescence microscopy showed that a fraction of HCF222-green fluorescent protein was detectable in the endoplasmic reticulum but that it also could be recognized in chloroplasts. A fusion construct of HCF222 containing a plastid transit peptide targets the protein into chloroplasts and was able to complement the mutational defect. These findings indicate that the chloroplast-targeted HCF222 is indispensable for the maturation and/or assembly of the Cytb6f complex and is very likely involved in thiol-disulfide biochemistry at the thylakoid membrane.

GreenCut protein CPLD49 of Chlamydomonas reinhardtii associates with thylakoid membranes and is required for cytochrome b6f complex accumulation

“…Assembly of the photosynthetic machinery involves coordinated synthesis of nucleus‐ and chloroplast‐encoded polypeptides along with the pigment molecules with which they associate (Eberhard et al ., ; Lyska et al ., ). Recently, several factors necessary for assembly of PSII, PSI and Cyt b 6 f were identified in cyanobacteria, algae and plants (Rochaix, , ; Schöttler et al ., ; Chi et al ., ; Heinnickel et al ., , ; Nickelsen and Rengstl, ; Hartings et al ., ; Wittenberg et al ., ). For the unicellular green alga Chlamydomonas reinhardtii (Chlamydomonas throughout), Cyt b 6 f assembly requires a class of proteins that facilitates integration of heme into apocytochrome b 6 and apocytochrome f (Inoue et al ., ; Xie et al ., ; Kuras et al ., ; Gabilly et al ., ; Cline et al ., ).…”

Novel DNAJ‐related proteins in Arabidopsis thaliana

“…DNAJE1.11/ANGULATA7 is involved in chloroplast development and leaf development (Munoz‐Nortes et al ., ). Recently, DNAJE1.14/HCF222 was found to be essential for seedling establishment and photosynthesis (Hartings et al ., ). Of the GRL‐containing DNAJE proteins, only DNAJE2.1/THRUMIN1 has yet been characterized, and is reported to bind actin and be involved in chloroplast motility (Whippo et al ., ).…”

>i<FIP>/i< (>i<>u<F>/u<tsH5 Interacting >u<P>/u<rotein>/i<): uma proteína dedo-de-zinco envolvida no mecanismo de resposta a estresse abiótico em >i<Arabidopsis thaliana>/i<

“…Muitos membros dessa família foram reportados por estarem envolvidos em diferentes processos na biogênese e/ou manutenção dos cloroplastos de arabidopsis como B2D2 (Brutnell et al, 1999), ORANGE (Lu et al, 2006), PTAC5 (Zhong et al, 2013) e ANGULATA7 (Munoz-Nortes et al, 2017). Além disso, LQY1 (Lu et al, 2011), 2011), PSA2 (Fristedt et al, 2014), CYO1 (Shimada et al, 2007) HCF222 (Hartings et al, 2017) (Brutnell et al, 1999). ANGULATA7 está envolvido na acumulação de pigmentos fotossintetizantes em cloroplastos de arabidopsis participando do mecanismo de crescimento e desenvolvimento foliar (Munoz-Nortes et al, 2017).…”

“…Mutantes psa2 tiveram severa redução no acúmulo de PSI e severa foto-inibição (Fristedt et al, 2014). LQY1 (Lu et al, 2011) e CYO1 (Shimada et al, 2007) (Hartings et al, 2017).…”

“…O ensaio de redução de insulina foi usado para monitorar a atividade dissulfeto redutase de 6xHis-FIP, onde foi medida a taxa de redução da insulina, devido ao aumento da turbidez provocada pela precipitação da cadeia B livre da insulina, liberada após a redução da ligação estrutural dissulfeto que liga as cadei as A e B (Holmgren, 1979). Este método foi usado com sucesso anteriormente para mostrar a atividade redutase das proteínas da família DNAJE1 de arabidopsis HCF222 (Hartings et al, 2017), CYO1 (Shimada et al, 2007) e PSA2 (Fristedt et al, 2014). A atividade redutase da Tiorredoxina é conhecida desde a década de 1960 (Thelander, 1968).…”