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1.
Plant Physiol ; 191(3): 1818-1835, 2023 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-36635853

RESUMO

Understanding the regulation of photosynthetic light harvesting and electron transfer is of great importance to efforts to improve the ability of the electron transport chain to supply downstream metabolism. A central regulator of the electron transport chain is ATP synthase, the molecular motor that harnesses the chemiosmotic potential generated from proton-coupled electron transport to synthesize ATP. ATP synthase is regulated both thermodynamically and post-translationally, with proposed phosphorylation sites on multiple subunits. In this study we focused on two N-terminal serines on the catalytic subunit ß in tobacco (Nicotiana tabacum), previously proposed to be important for dark inactivation of the complex to avoid ATP hydrolysis at night. Here we show that there is no clear role for phosphorylation in the dark inactivation of ATP synthase. Instead, mutation of one of the two phosphorylated serine residues to aspartate to mimic constitutive phosphorylation strongly decreased ATP synthase abundance. We propose that the loss of N-terminal phosphorylation of ATPß may be involved in proper ATP synthase accumulation during complex assembly.


Assuntos
ATPases de Cloroplastos Translocadoras de Prótons , Fotossíntese , ATPases de Cloroplastos Translocadoras de Prótons/genética , ATPases de Cloroplastos Translocadoras de Prótons/metabolismo , Fosforilação , Fotossíntese/genética , Transporte de Elétrons , Trifosfato de Adenosina/metabolismo
2.
J Exp Bot ; 72(7): 2544-2569, 2021 03 29.
Artigo em Inglês | MEDLINE | ID: mdl-33484250

RESUMO

Carotenoids are important isoprenoids produced in the plastids of photosynthetic organisms that play key roles in photoprotection and antioxidative processes. ß-Carotene is generated from lycopene by lycopene ß-cyclase (LCYB). Previously, we demonstrated that the introduction of the Daucus carota (carrot) DcLCYB1 gene into tobacco (cv. Xanthi) resulted in increased levels of abscisic acid (ABA) and especially gibberellins (GAs), resulting in increased plant yield. In order to understand this phenomenon prior to exporting this genetic strategy to crops, we generated tobacco (Nicotiana tabacum cv. Petit Havana) mutants that exhibited a wide range of LCYB expression. Transplastomic plants expressing DcLCYB1 at high levels showed a wild-type-like growth, even though their pigment content was increased and their leaf GA1 content was reduced. RNA interference (RNAi) NtLCYB lines showed different reductions in NtLCYB transcript abundance, correlating with reduced pigment content and plant variegation. Photosynthesis (leaf absorptance, Fv/Fm, and light-saturated capacity of linear electron transport) and plant growth were impaired. Remarkably, drastic changes in phytohormone content also occurred in the RNAi lines. However, external application of phytohormones was not sufficient to rescue these phenotypes, suggesting that altered photosynthetic efficiency might be another important factor explaining their reduced biomass. These results show that LCYB expression influences plant biomass by different mechanisms and suggests thresholds for LCYB expression levels that might be beneficial or detrimental for plant growth.


Assuntos
Liases Intramoleculares , Nicotiana , Carotenoides , Regulação da Expressão Gênica de Plantas , Liases Intramoleculares/genética , Liases Intramoleculares/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Nicotiana/genética , Nicotiana/metabolismo
3.
Nat Plants ; 5(3): 282-289, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30778165

RESUMO

The development of technologies for the stable genetic transformation of plastid (chloroplast) genomes has been a boon to both basic and applied research. However, extension of the transplastomic technology to major crops and model plants has proven extremely challenging, and the species range of plastid transformation is still very much limited in that most species currently remain recalcitrant to plastid genome engineering. Here, we report an efficient plastid transformation technology for the model plant Arabidopsis thaliana that relies on root-derived microcalli as a source tissue for biolistic transformation. The method produces fertile transplastomic plants at high frequency when combined with a clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9)-generated knockout allele of a nuclear locus that enhances sensitivity to the selection agent used for isolation of transplastomic events. Our work makes the model organism of plant biology amenable to routine engineering of the plastid genome, facilitates the combination of plastid engineering with the power of Arabidopsis nuclear genetics, and informs the future development of plastid transformation protocols for other recalcitrant species.


Assuntos
Arabidopsis/fisiologia , Sistemas CRISPR-Cas , Plantas Geneticamente Modificadas , Plastídeos/genética , Arabidopsis/citologia , Arabidopsis/genética , Proteínas de Bactérias/genética , Biolística/métodos , Técnicas de Cultura de Células , Cloroplastos/genética , Edição de Genes , Técnicas de Inativação de Genes , Vetores Genéticos , Proteínas Luminescentes/genética , Raízes de Plantas/citologia , Raízes de Plantas/genética , Transformação Genética
4.
Mol Plant ; 2(6): 1223-32, 2009 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-19995727

RESUMO

The chloroplasts genome (plastome) occurs at high copy numbers per cell. Several chloroplast genome copies are densely packed into nucleoprotein particles called nucleoids. How genome packaging occurs and which proteins organize chloroplast nucleoids are largely unknown. Here, we have analyzed the Chlamydomonas reinhardtii homolog of the bacterial architectural DNA-binding protein HU, the histone-like protein HLP. We show that the Chlamydomonas HLP protein is targeted to chloroplasts and associates with nucleoids. Knockdown of HLP gene expression by RNA interference (RNAi) alters the structure of chloroplast nucleoids and appears to reduce the level of compaction of chloroplast DNA. Unexpectedly, also chloroplast genome copy numbers are significantly decreased in the RNAi strains, suggesting that, in addition to its architectural role in nucleoid formation, the HLP protein is also involved in chloroplast genome maintenance.


Assuntos
Núcleo Celular/genética , Chlamydomonas reinhardtii/genética , Genoma de Planta , Sequência de Aminoácidos , Sequência de Bases , Cloroplastos/genética , Cloroplastos/fisiologia , Clonagem Molecular , Cianobactérias/genética , Dados de Sequência Molecular , Proteínas de Plantas/genética , Proteínas de Plantas/fisiologia , Mapeamento por Restrição , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos
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