Stromule extension along microtubules coordinated with actin-mediated anchoring guides perinuclear chloroplast movement during innate immunity.

Dynamic tubular extensions from chloroplasts called stromules have recently been shown to connect with nuclei and function during innate immunity. We demonstrate that stromules extend along microtubules (MTs) and MT organization directly affects stromule dynamics since stabilization of MTs chemically or genetically increases stromule numbers and length. Although actin filaments (AFs) are not required for stromule extension, they provide anchor points for stromules. Interestingly, there is a strong correlation between the direction of stromules from chloroplasts and the direction of chloroplast movement. Stromule-directed chloroplast movement was observed in steady-state conditions without immune induction, suggesting it is a general function of stromules in epidermal cells. Our results show that MTs and AFs may facilitate perinuclear clustering of chloroplasts during an innate immune response. We propose a model in which stromules extend along MTs and connect to AF anchor points surrounding nuclei, facilitating stromule-directed movement of chloroplasts to nuclei during innate immunity

The formation of homogentisate in the biosynthesis of tocopherol and plastoquinone in spinach chloroplasts

Homogentisate is the precursor in the biosynthesis of -tocopherol and plastoquinone-9 in chloroplasts. It is formed of 4-hydroxyphenylpyruvate of the shikimate pathway by the 4-hydroxyphenylpyruvate dioxygenase. In experiments with spinach the dioxygenase was shown to be localized predominatedly in the chloroplasts. Envelope membranes exhibit the highest specific activity, however, because of the high stromal portion of chloroplasts, 60–80% of the total activity is housed in the stroma. The incorporation of 4-hydroxyphenylpyruvate into 2-methyl-6-phytylquinol as the first intermediate in the tocopherol synthesis by the two-step-reaction: 4-Hydroxyphenylpyruvate Homogentisate 2-Methyl-6-phytylquinol was demonstrated by using envelope membranes. Homogentisate originates directly from 4-hydroxyphenylpyruvate of the shikimate pathway. Additionally, a bypass exists in chloroplasts which forms 4-hydroxyphenylpyruvate from tyrosine by an L-amino-acid oxidase of the thylakoids and in peroxisomes by a transaminase reaction. Former results about the dioxygenase in peroxisomes were verified

Phytol synthesis from geranylgeraniol in spinach chloroplasts

The reduction of /2-14C/-geranylgeranylpyrophosphate to phytylpyrophosphosphate is shown for the first time in chloroplasts. The esterification of exogenous /2-14C/-geranylgeranylpyrophosphate with endogenous chlorophyllide and the stepwise reduction of the pigment bound geranylgeraniol to phytol was also proved for spinach chloroplasts for the first time

Physiological and transcriptomic evidence for a close coupling between chloroplast ontogeny and cell cycle progression in the pennate diatom Seminavis robusta

Despite the growing interest in diatom genomics, detailed time series of gene expression in relation to key cellular processes are still lacking. Here, we investigated the relationships between the cell cycle and chloroplast development in the pennate diatom Seminavis robusta. This diatom possesses two chloroplasts with a well-orchestrated developmental cycle, common to many pennate diatoms. By assessing the effects of induced cell cycle arrest with microscopy and flow cytometry, we found that division and reorganization of the chloroplasts are initiated only after S-phase progression. Next, we quantified the expression of the S. robusta FtsZ homolog to address the division status of chloroplasts during synchronized growth and monitored microscopically their dynamics in relation to nuclear division and silicon deposition. We show that chloroplasts divide and relocate during the S/G2 phase, after which a girdle band is deposited to accommodate cell growth. Synchronized cultures of two genotypes were subsequently used for a cDNA-amplified fragment length polymorphism-based genome-wide transcript profiling, in which 917 reproducibly modulated transcripts were identified. We observed that genes involved in pigment biosynthesis and coding for light-harvesting proteins were up-regulated during G2/M phase and cell separation. Light and cell cycle progression were both found to affect fucoxanthin-chlorophyll a/c-binding protein expression and accumulation of fucoxanthin cell content. Because chloroplasts elongate at the stage of cytokinesis, cell cycle-modulated photosynthetic gene expression and synthesis of pigments in concert with cell division might balance chloroplast growth, which confirms that chloroplast biogenesis in S. robusta is tightly regulated

Dating the cyanobacterial ancestor of the chloroplast

Cyanobacteria have played a pivotal role in the history of life on Earth being the first organism to carry out oxygenic photosynthesis, which changed atmospheric chemistry and allowed the evolution of Eukarya. Chloroplasts are the cellular organelles of photoautotrophic eukaryotes in which photosynthesis is conducted. Although the initial suggestion from Mereschkowsky (1905) that cyanobacteria are the ancestors of chloroplasts was greeted with skepticism, the idea is now widely accepted. Here we attempt to resolve and date the cyanobacterial ancestry of the chloroplast using phylogenetic analysis and molecular clocks, because until now, the long-standing question of, from which, among the vast diversity of cyanobacteria, did chloroplasts evolve, has not been resolved. We found that chloroplasts form a monophyletic lineage, are most closely related to subsection I, N2-fixing unicellular cyanobacteria (Order Chroococcales), and heterocyst-forming Order Nostocales cyanobacteria are their sister group. The appearance of Nostocales and of Chroococcales occurred during the Paleoproterozoic, and chloroplasts appeared in the mid-Proterozoic. The capability of N2-fixation in cyanobacteria appeared once during the late Archaen and early Proterozoic eons. Furthermore, we found that oxygen-evolving cyanobacteria could have appeared in the Archean. Our results suggest that a free-living cyanobacterium with the capacity to store starch via oxygenic CO2 fixation, and to fix atmospheric N2, would be a very important intracellular acquisition, which, as can be recounted today from several lines of evidence, would have become the chloroplast by endosymbiosis

Effects of Growth Temperatures on the Fatty Acid Composition of Isolated Chloroplasts From Two Species Differing in Heat Sensitivity

Lipid analyses of chloroplasts isolated from wheat (Triticum aestivum L. cv. Arthur) and milo (Sorghum bicolor cv. Funk\u27s hybrid 522) suggest no major heat effect on lipid class distribution. Assuming milo is more heat tolerant than wheat and that increased saturated/unsaturated fatty acid values increase thermal stability, changes in sulfoquinovosyldiglyceride (SL) appear to be more important than phosphatidylglycerol (PG) in conferring thermal stability to isolated chloroplasts

Tocopherol and plastoquinone synthesis in spinach chloroplasts subfractions

Subfractions isolated from intact purified spinach chloroplasts are able to prenylate the aromatic moiety of -tocopherol and plastoquinone-9 precursors. The biosynthesis of -tocopherol and plastoquinone-9 is a compartmentalized process. The chloroplast envelope membranes are the only site of the enzymatic prenylation in -tocopherol synthesis whereas the thylakoid membrane is also involved in the prenylation and methylation sequence of plastoquinone-9 biosynthesis. A very active kinase which forms phytyl-PP is localized in the stroma. Phytol but not geranylgeraniol is the polyprenol precursor of the side chain of -tocopherol in spinach chloroplasts