Abstract
BackgroundMitogen-activated protein kinase (MAPK) cascades are conserved signaling modules in eukaryotic organisms and play essential roles in immunity and stress responses. However, the role of MAPKs in chloroplast development remains to be evidently established.ResultsIn this study, a rice chlorosis seedling lethality 1 (csl1) mutant with a Zhonghua11 (ZH11, japonica) background was isolated. Seedlings of the mutant were characterized by chlorotic leaves and death after the trefoil stage, and chloroplasts were observed to contain accumulated starch granules. Molecular cloning revealed that OsCSL1 encoded a MAPK kinase kinase22 (MKKK22) targeted to the endoplasmic reticulum (ER), and functional complementation of OsCSL1 was found to restore the normal phenotype in csl1 plants. The CRISPR/Cas9 technology was used for targeted disruption of OsCSL1, and the OsCSL1-Cas9 lines obtained therein exhibited yellow seedlings which phenocopied the csl1 mutant. CSL1/MKKK22 was observed to establish direct interaction with MKK4, and altered expression of MKK1 and MKK4 was detected in the csl1 mutant. Additionally, disruption of OsCSL1 led to reduced expression of chloroplast-associated genes, including chlorophyll biosynthetic genes, plastid-encoded RNA polymerases, nuclear-encoded RNA polymerase, and nuclear-encoded chloroplast genes.ConclusionsThe findings of this study revealed that OsCSL1 played roles in regulating the expression of multiple chloroplast synthesis-related genes, thereby affecting their functions, and leading to wide-ranging defects, including chlorotic seedlings and severely disrupted chloroplasts containing accumulated starch granules.
Highlights
Mitogen-activated protein kinase (MAPK) cascades are conserved signaling modules in eukaryotic organisms and play essential roles in immunity and stress responses
The csl1 mutation results in the development of chlorotic seedlings and a lethal phenotype To investigate the mechanisms underlying MAPK involvement in chloroplast development, a mutant, chlorosis seedling lethality 1, displaying defects in leaf pigmentation was identified among rice T-DNA insertion populations with a Zhonghua11 background
The contents of chlorophylls a and b and total chlorophyll content in both leaf blades and sheaths were significantly reduced in the csl1 mutant compared with WT seedlings (Fig. 1C), whereas no appreciable differences were detected between WT and heterozygous csl1 lines (Fig. S1)
Summary
Mitogen-activated protein kinase (MAPK) cascades are conserved signaling modules in eukaryotic organisms and play essential roles in immunity and stress responses. Chloroplasts are defined as organelles that play specific roles in the conversion of light energy to chemical energy via photosynthesis [1], and their functional and structural integrity are vital for normal plant growth and development [2, 3]. Glycinamide ribonucleotide synthetase (GARS), which catalyzes the second step in purine nucleotide biosynthesis, has been identified to be involved in chloroplast development in rice by affecting the expression of plastid-encoded genes [13]. Precise chloroplast function is coordinately mediated by two types of RNA polymerase, namely the nuclearencoded RNA polymerases (NEPs) and plastid-encoded RNA polymerases (PEPs) [15, 16], which are essential for the biogenesis of photosynthetically active chloroplast in plants [16, 17]. Apart from PEPs and NEPs, chlorophyll biosynthetic genes (CBGs) and nuclear-encoded chloroplast genes (NECGs) have been established to be involved in chloroplast development
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