Dongzhi Lin

The rice OsV4 encoding a novel pentatricopeptide repeat protein is required for chloroplast development during the early leaf stage under cold stress

Pentatricopeptide repeat (PPR) proteins, characterized by tandem arrays of a 35 amino acid motif, have been suggested to play central and broad roles in modulating the expression of organelle genes in plants. However, the molecular mechanisms of most rice PPR genes remains unclear. In this paper, we isolated and characterized a temperature-conditional virescent mutant, OsV4, in rice (Oryza sativa cultivar Jiahua1 (WT, japonica rice variety)). The mutant displays albino phenotype and abnormal chloroplasts at the three leaf stage, which gradually turns green after the four leaf stage at a low temperature (20 °C). But the mutant always develops green leaves and well-developed chloroplasts at a high temperature (32 °C). Genetic and molecular analyses uncovered that OsV4 encodes a novel chloroplast-targeted PPR protein including four PPR motifs. Further investigations show that the mutant phenotype is associated with changes in chlorophyll content and chloroplast development. The OsV4 transcripts only accumulate to high levels in young leaves, indicating that its expression is tissue-specific. In addition, transcript levels of some ribosomal components and plastid-encoded polymerase-dependent genes are dramatically reduced in the albino mutants grown at 20 °C. These findings suggest that OsV4 plays an important role during early chloroplast development under cold stress in rice.

Importance of the rice TCD9 encoding α subunit of chaperonin protein 60 (Cpn60α) for the chloroplast development during the early leaf stage.

The chloroplast development governs plant growth and metabolism. This study characterized a novel rice thermo-sensitive chloroplast development 9 (tcd9) mutant, which exhibited the albino phenotype before the 3-leaf stage grown below 24 °C whereas displayed normal green at over 28 °C or even at 20 °C after 5-leaf stage. The obvious decrease in Chl levels, abnormal chloroplasts with few thylakoid lamella and abnormal thylakoids were observed for the albino mutant seedlings at low temperature, but the mutant was apparently normal green at high temperature, suggesting the thermo-sensitivity of albino phenotype. Genetic analysis showed that the albino phenotype was controlled by a single recessive nuclear gene (tcd9). The map-based cloning and molecular complementation tests revealed that the mutation of TCD9 encoding α subunit of Cpn60 protein (Cpn60α), localized in chloroplasts, was responsible for albino phenotype. In addition, TCD9 exhibited the high expression in all tested tissues, especially in young leaves. The transcriptional analysis indicated that all expression levels of the studied genes related to chloroplast development in tcd9 mutant were seriously affected in the albino seedlings at 20 °C, whereas some of them recovered into normal levels in green-seedlings at 32 °C. Our observations suggest that the nuclear-encoded Cpn60α protein TCD9 plays a crucial role in chloroplast development at early leaf stage of rice.

Temperature-sensitive albino gene TCD5, encoding a monooxygenase, affects chloroplast development at low temperatures

Highlight A new temperature-sensitive albino gene, TCD5, encoding a monooxygenase, affects chloroplast development at P4 stage under low temperature in rice.

Mutation of the rice ASL2 gene encoding plastid ribosomal protein L21 causes chloroplast developmental defects and seedling death

The plastid ribosome proteins (PRPs) play important roles in plastid protein biosynthesis, chloroplast differentiation and early chloroplast development. However, the specialised functions of individual protein components of the chloroplast ribosome in rice (Oryza sativa) remain unresolved. In this paper, we identified a novel rice PRP mutant named asl2 (Albino seedling lethality 2) exhibiting an albino, seedling death phenotype. In asl2 mutants, the alteration of leaf colour was associated with chlorophyll (Chl) content and abnormal chloroplast development. Through map-based cloning and complementation, the mutated ASL2 gene was isolated and found to encode the chloroplast 50S ribosome protein L21 (RPL21c), a component of the chloroplast ribosome large subunit, which was localised in chloroplasts. ASL2 was expressed at a higher level in the plumule and leaves, implying its tissue-specific expression. Additionally, the expression of ASL2 was regulated by light. The transcript levels of the majority of genes for Chl biosynthesis, photosynthesis and chloroplast development were strongly affected in asl2 mutants. Collectively, the absence of functional ASL2 caused chloroplast developmental defects and seedling death. This report establishes the important role of RPL21c in chloroplast development in rice.

The rice TCD11 encoding plastid ribosomal protein S6 is essential for chloroplast development at low temperature

Plastid ribosome proteins (PRPs) are important components for chloroplast biogenesis and early chloroplast development. Although it has been known that chloroplast ribosomes are similar to bacterial ones, the precise molecular function of ribosomal proteins remains to be elucidated in rice. Here, we identified a novel rice mutant, designated tcd11 (thermo-sensitive chlorophyll-deficient mutant 11), characterized by the albino phenotype until it died at 20°C, while displaying normal phenotype at 32°C. The alteration of leaf color in tcd11 mutants was aligned with chlorophyll (Chl) content and chloroplast development. The map-based cloning and molecular complementation showed that TCD11 encodes the ribosomal small subunit protein S6 in chloroplasts (RPS6). TCD11 was abundantly expressed in leaves, suggesting its different expressions in tissues. In addition, the disruption of TCD11 greatly reduced the transcript levels of certain chloroplasts-associated genes and prevented the assembly of ribosome in chloroplasts at low temperature (20°C), whereas they recovered to nearly normal levels at high temperature (32°C). Thus, our data indicate that TCD11 plays an important role in chloroplast development at low temperature. Upon our knowledge, the observations from this study provide a first glimpse into the importance of RPS6 function in rice chloroplast development.

The rice ALS3 encoding a novel pentatricopeptide repeat protein is required for chloroplast development and seedling growth

BackgroundPentatricopeptide repeat (PPR) proteins play essential roles in modulating the expression of organelle genes and have expanded greatly in higher plants. However, molecular mechanisms of most rice PPR genes remain unclear.ResultsIn this study, a new rice PPR mutant, asl3 (albinoseedlinglethality3) exhibits an albino lethal phenotype at the seedling stage. This albino phenotype was associated with altered photosynthetic-pigment and chloroplast development. Map-based cloning showed that ASL3 encodes a novel rice PPR protein with 10 tandem PPR motifs, which localizes to the chloroplast. ASL3 showed tissue-specific expression, as it was highly expressed in the chlorenchyma, but expressed at much lower levels in roots and panicles. RNAi of ASL3 confirmed that ASL3 plays an essential role in the early development and chloroplast development in rice. Moreover, expression analysis revealed that the asl3 mutation severely affected the transcriptional levels of important genes associated with plastid translation machinery and photosynthesis, which may impair photosynthesis and finally led to the seedling death in asl3 mutant. These results evidenced the important role of ASL3 in the early development of rice, especially chloroplast development.ConclusionsThe ASL3 gene encoded a novel chloroplast-targeted PPR protein with 10 tandem PPR motifs in rice. Disruption of the ASL3 would lead to a defective chloroplast and seedling lethality, and affected expression levels of genes associated with chloroplast development and photosynthesis at early leaf stage of rice.

The rice OsDG2 encoding a glycine-rich protein is involved in the regulation of chloroplast development during early seedling stage.

Key messageOsDG2 gene encoded a novel chloroplast-targeted GRP in rice. Disruption of the OsDG2 would lead to delayed greening phenotype and affected expression levels of genes associated with chloroplast development at early leaf stage of rice.AbstractGlycine-rich proteins (GRPs) participate in various biological processes in plants. However, the evidence of GRPs involved in chloroplast development in plants is quite limited. In this study, we identified a rice GRP gene mutant named osdg2 (Oryzasativadelayedgreening 2), which exhibits delayed greening phenotype characterized as bright yellow leaves before the three-leaf stage and thereafter turns to normal green. Further study showed that the mutant phenotype was consistent with changes in chlorophyll content and chloroplast development. The rice OsDG2 gene, encoding a novel GRP protein, was located on chromosome 2 through map-based cloning method and confirmed by molecular complementation tests. Subcellular localization results showed that OsDG2 was targeted in chloroplasts. In addition, the OsDG2 transcripts were highly expressed in leaves and undetectable in other tissues, showing the tissue-specific expression. In osdg2 mutant, the expression levels of most genes associated with chloroplast development were severely decreased in the 3rd leaves, but almost recovered to wild-type level in the 4th leaves. Our findings indicated that the nuclear-encoded OsDG2 plays important roles in chloroplast development at early leaf stage of rice.

Rice TSV3 Encoding Obg-Like GTPase Protein Is Essential for Chloroplast Development During the Early Leaf Stage Under Cold Stress

The Spo0B-associated GTP-binding (Obg) proteins are essential for the viability of nearly all bacteria. However, the detailed roles of Obg proteins in higher plants have not yet been elucidated. In this study, we identified a novel rice (Oryza sativa L.) thermo-sensitive virescent mutant (tsv3) that displayed an albino phenotype at 20° before the three-leaf stage while being a normal green at 32° or even at 20° after the four-leaf stage. The mutant phenotype was consistent with altered chlorophyll content and chloroplast structure in leaves. Map-based cloning and complementation experiments showed that TSV3 encoded a small GTP-binding protein. Subcellular localization studies revealed that TSV3 was localized to the chloroplasts. Expression of TSV3 was high in leaves and weak or undetectable in other tissues, suggesting a tissue-specific expression of TSV3. In the tsv3 mutant, expression levels of genes associated with the biogenesis of the chloroplast ribosome 50S subunit were severely decreased at the three-leaf stage under cold stress (20°), but could be recovered to normal levels at a higher temperature (32°). These observations suggest that the rice nuclear-encoded TSV3 plays important roles in chloroplast development at the early leaf stage under cold stress.

Rice TCM1 Encoding a Component of the TAC Complex is Required for Chloroplast Development under Cold Stress

Transcriptionally active chromosome (TAC) is a component of protein–DNA complexes with RNA polymerase activity, expressed in the plastid. Map‐based cloning revealed that TCM1 encodes a novel chloroplast‐targeted TAC protein in rice, in which a mutation leads to an albino phenotype and malformed chloroplasts before the three‐leaf stage at low temperatures. TAC protein TCM1 is essential for proper chloroplast development and maintaining plastid‐encoded polymerase activity under cold stress conditions.