Abstract
Chloroplasts are semi-autonomous organelles governed by the precise coordination between the genomes of their own and the nucleus for functioning correctly in response to developmental and environmental cues. Under stressed conditions, various plastid-to-nucleus retrograde signals are generated to regulate the expression of a large number of nuclear genes for acclimation. Among these retrograde signaling pathways, the chloroplast protein GENOMES UNCOUPLED 1 (GUN1) is the first component identified. However, in addition to integrating aberrant physiological signals when chloroplasts are challenged by stresses such as photooxidative damage or the inhibition of plastid gene expression, GUN1 was also found to regulate other developmental processes such as flowering. Several partner proteins have been found to interact with GUN1 and facilitate its different regulatory functions. In this study, we report 15 possible interacting proteins identified through yeast two-hybrid (Y2H) screening, among which 11 showed positive interactions by pair-wise Y2H assay. Through the bimolecular fluorescence complementation assay in Arabidopsis protoplasts, two candidate proteins with chloroplast localization, DJC31 and HCF145, were confirmed to interact with GUN1 in planta. Genes for these GUN1-interacting proteins showed different fluctuations in the WT and gun1 mutant under norflurazon and lincomycin treatments. Our results provide novel clues for a better understanding of molecular mechanisms underlying GUN1-mediated regulations.
Highlights
Chloroplasts are semi-autonomous organelles of photosynthetic eukaryotes
Genetic evidence for the retrograde communication was first demonstrated in Arabidopsis thaliana by the characterization of the genomes uncoupled 1 mutants, which showed partially rescued expression of photosynthesis-associated nuclear genes (PhANGs), compared with the wild type, when chloroplasts were photodamaged under treatment by norflurazon (NF) [5]
GENOMES UNCOUPLED 1 (GUN1) modulates protein homeostasis by controlling the accumulations of plastid ribosomal protein S1 (PRPS1), plays a direct role in RNA editing through its interaction with MORF2, and regulates chloroplast protein import through its interaction with the import-related chaperone cpHSC70-1 [14,15,16]
Summary
Chloroplasts are semi-autonomous organelles of photosynthetic eukaryotes. More than 95% of chloroplast proteins are encoded by nuclear genes [1]. GUN1 modulates protein homeostasis by controlling the accumulations of plastid ribosomal protein S1 (PRPS1), plays a direct role in RNA editing through its interaction with MORF2, and regulates chloroplast protein import through its interaction with the import-related chaperone cpHSC70-1 [14,15,16]. GUN1 has been demonstrated to interact with several partner proteins. In addition to PRPS1, MORF2, and cpHSC70-1 mentioned above, GUN1 was demonstrated to interact with enzymes for tetrapyrrole biosynthesis (such as ChlD) and a cytosolic protein. The molecular mechanisms underlying a plethora of GUN1mediated regulations remain elusive, GUN1 was proposed to function, at least partially, through its physical interaction with different protein partners [14,19,20]. We report the confirmation of the interactions between GUN1 and two chloroplast proteins, DJC31 and HCF145, by Y2H and biomolecular fluorescence complementation (BiFC) assays
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