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Structure of a Step II Catalytically Activated Spliceosome from Chlamydomonas reinhardtii

ABSTRACT: Pre-mRNA splicing, a fundamental step in eukaryotic gene expression, is executed by the spliceosomes. While there is extensive knowledge of the compositiona and structure of spliceosomes in yeasts and humans, the structural diversity of spliceosomes in non-canonical organisms remains unclear. Here, we present a cryo-EM structure of a step II catalytically activated spliceosome (C* complex) derived from the unicellular green alga Chlamydomonas reinhardtii at 2.6 Å resolution. This Chlamydomonas C* complex comprises 29 proteins and four RNA elements, creating a dynamic assembly that shares a similar overall architecture with yeast and human counterparts but also has unique features of its own. Distinctive structural characteristics include variations in protein compositions as well as some noteworthy RNA features. The splicing factor Prp17, with four fragments and a WD40 domain, is engaged in intricate interactions with multiple protein and RNA components. The structural elucidation of Chlamydomonas C* complex provides insights into the molecular mechanism of RNA splicing in plants and understanding splicing evolution in eukaryotes.

SUBMITTER: Xiechao Zhan

PROVIDER: S-SCDT-10_1038-S44318-024-00274-3 | biostudies-other |

REPOSITORIES: biostudies-other

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Structure of native photosystem II assembly intermediate from <i>Chlamydomonas reinhardtii</i>.

Project description:Chlamydomonas reinhardtii Photosystem II (PSII) is a dimer consisting of at least 13 nuclear-encoded and four chloroplast-encoded protein subunits that collectively function as a sunlight-driven oxidoreductase. In this study, we present the inaugural structure of a green alga PSII assembly intermediate (pre-PSII-int). This intermediate was isolated from chloroplast membranes of the temperature-sensitive mutant TSP4, cultivated for 14 hours at a non-permissive temperature. The assembly state comprises a monomer containing subunits A, B, C, D, E, F, H, I, K, and two novel assembly factors, Psb1 and Psb2. Psb1 is identified as a novel transmembrane helix located adjacent to PsbE and PsbF (cytochrome b559). The absence of PsbJ, typically found in mature PSII close to this position, indicates that Psb1 functions as an assembly factor. Psb2 is an eukaryotic homolog of the cyanobacterial assembly factor Psb27. The presence of iron, coupled with the absence of QA, QB, and the manganese cluster, implies a protective mechanism against photodamage and provides insights into the intricate assembly process.

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Structure of GUN4 from Chlamydomonas reinhardtii.

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Structure of the bifunctional cryptochrome aCRY from Chlamydomonas reinhardtii.

Project description:Photolyases and cryptochromes form an almost ubiquitous family of blue light photoreceptors involved in the repair and maintenance of DNA integrity or regulatory control. We found that one cryptochrome from the green alga Chlamydomonas reinhardtii (CraCRY) is capable of both, control of transcript levels and the sexual cycle of the alga in a positive (germination) and negative manner (mating ability), as well as catalyzing the repair of UV-DNA lesions. Its 1.6 Å crystal structure shows besides the FAD chromophore an aromatic tetrad that is indispensable in animal-like type I cryptochromes for light-driven change of their signaling-active redox state and formation of a stable radical pair. Given CraCRY's catalytic activity as (6-4) photolyase in vivo and in vitro, we present the first co-crystal structure of a cryptochrome with duplex DNA comprising a (6-4) pyrimidine-pyrimidone lesion. This 2.9 Å structure reveals a distinct conformation for the catalytic histidine His1, H357, that challenges previous models of a single-photon driven (6-4) photolyase mechanism.

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Selenoprotein-transgenic Chlamydomonas reinhardtii.

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Structural Analysis of the Rubisco-Assembly Chaperone RbcX-II from Chlamydomonas reinhardtii.

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Energy-dissipative supercomplex of photosystem II associated with LHCSR3 in Chlamydomonas reinhardtii.

Project description:Plants and green algae have a low pH-inducible mechanism in photosystem II (PSII) that dissipates excess light energy, measured as the nonphotochemical quenching of chlorophyll fluorescence (qE). Recently, nonphotochemical quenching 4 (npq4), a mutant strain of the green alga Chlamydomonas reinhardtii that is qE-deficient and lacks the light-harvesting complex stress-related protein 3 (LHCSR3), was reported [Peers G, et al. (2009) Nature 462(7272):518-521]. Here, applying a newly established procedure, we isolated the PSII supercomplex and its associated light-harvesting proteins from both WT C. reinhardtii and the npq4 mutant grown in either low light (LL) or high light (HL). LHCSR3 was present in the PSII supercomplex from the HL-grown WT, but not in the supercomplex from the LL-grown WT or mutant. The purified PSII supercomplex containing LHCSR3 exhibited a normal fluorescence lifetime at a neutral pH (7.5) by single-photon counting analysis, but a significantly shorter lifetime at pH 5.5, which mimics the acidified lumen of the thylakoid membranes in HL-exposed chloroplasts. The switch from light-harvesting mode to energy-dissipating mode observed in the LHCSR3-containing PSII supercomplex was sensitive to dicyclohexylcarbodiimide, a protein-modifying agent specific to protonatable amino acid residues. We conclude that the PSII-LHCII-LHCSR3 supercomplex formed in the HL-grown C. reinhardtii cells is capable of energy dissipation on protonation of LHCSR3.

| S-EPMC3683755 | biostudies-literature

Genetic structure of the mating-type locus of Chlamydomonas reinhardtii.

Project description:Portions of the cloned mating-type (MT) loci (mt(+) and mt(-)) of Chlamydomonas reinhardtii, defined as the approximately 1-Mb domains of linkage group VI that are under recombinational suppression, were subjected to Northern analysis to elucidate their coding capacity. The four central rearranged segments of the loci were found to contain both housekeeping genes (expressed during several life-cycle stages) and mating-related genes, while the sequences unique to mt(+) or mt(-) carried genes expressed only in the gametic or zygotic phases of the life cycle. One of these genes, Mtd1, is a candidate participant in gametic cell fusion; two others, Mta1 and Ezy2, are candidate participants in the uniparental inheritance of chloroplast DNA. The identified housekeeping genes include Pdk, encoding pyruvate dehydrogenase kinase, and GdcH, encoding glycine decarboxylase complex subunit H. Unusual genetic configurations include three genes whose sequences overlap, one gene that has inserted into the coding region of another, several genes that have been inactivated by rearrangements in the region, and genes that have undergone tandem duplication. This report extends our original conclusion that the MT locus has incurred high levels of mutational change.

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Small RNAs in Chlamydomonas reinhardtii

Project description:RNA populations in Chlamydomonas reinhardtii Keywords: Highly parallel pyrosequencing

2007-05-16 | GSE7308 | GEO

Phosphate Starvation in Chlamydomonas reinhardtii

Project description:Phosphorus (P) is an essential nutrient that is limiting in many environments. When P is scarce organisms employ strategies for conservation of internal stores, and to efficiently scavenge P from their external surroundings. In this study we investigated the acclimation response of Chlamydomonas reinhardtii to P deficiency, comparing the transcriptional profiles of P starved wild-type cells to the P replete condition. RNA was prepared from P-containing or P-deprived logarithmic growth phase cells and subjected to RNA-Seq analysis. During the 24 hours after the imposition of P starvation we observed that from the 407 significantly changing genes (> 2 fold change, corrected p-value < 0.05) in the wild-type 317 genes were up-regulated, in average 8.36-fold, and 90 genes were down-regulated by 3.43-fold, in average. Many of the upregulated genes encoded enzymes involved in specific responses to P starvation, including PHOX, encoding the major secreted alkaline phosphatase, and multiple putative, high-efficiency phosphate transporter genes. More general responses included the up-regulation of genes involved in photoprotective processes (LHCSR3, LHCSR1, LHCBM9, PTOX1) and genes involved in protein modification and degradation. Down-regulated mRNAs indicated an early stage of the reduction of chloroplast ribosomal proteins, which are considered to be a reservoir for P in the cell.

2014-04-05 | GSE56505 | GEO

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