Microalgae have potential to help meet energy and food demands without exacerbating environmental problems. There is interest in the unicellular green alga Chromochloris zofingiensis, because it produces lipids for biofuels and a highly valuable carotenoid nutraceutical, astaxanthin. To advance understanding of its biology and facilitate commercial development, we present a C. zofingiensis chromosome-level nuclear genome, organelle genomes, and transcriptome from diverse growth conditions. The assembly, derived from a combination of short-and long-read sequencing in conjunction with optical mapping, revealed a compact genome of ∼58 Mbp distributed over 19 chromosomes containing 15,274 predicted protein-coding genes. The genome has uniform gene density over chromosomes, low repetitive sequence content (∼6%), and a high fraction of protein-coding sequence (∼39%) with relatively long coding exons and few coding introns. Functional annotation of gene models identified orthologous families for the majority (∼73%) of genes. Synteny analysis uncovered localized but scrambled blocks of genes in putative orthologous relationships with other green algae. Two genes encoding beta-ketolase (BKT), the key enzyme synthesizing astaxanthin, were found in the genome, and both were up-regulated by high light. Isolation and molecular analysis of astaxanthin-deficient mutants showed that BKT1 is required for the production of astaxanthin. Moreover, the transcriptome under high light exposure revealed candidate genes that could be involved in critical yet missing steps of astaxanthin biosynthesis, including ABC transporters, cytochrome P450 enzymes, and an acyltransferase. The high-quality genome and transcriptome provide insight into the green algal lineage and carotenoid production.Chlorophyceae | carotenoid biosynthesis | de novo genome | genome mapping | RNA-Seq

DNA methylation is a heritable epigenetic mark that controls gene expression, is responsive to environmental stresses, and, in plants, may also play a role in heterosis. To determine the degree to which DNA methylation is inherited in rice, and how it both influences and is affected by transcription, we performed genome-wide measurements of these patterns through an integrative analysis of bisulfite-sequencing, RNA-sequencing, and siRNA-sequencing data in two inbred parents of the Nipponbare (NPB) and indica (93-11) varieties of rice and their hybrid offspring. We show that SNPs occur at a rate of about 1∕253 bp between the two parents and that these are faithfully transmitted into the hybrids. We use the presence of these SNPs to reconstruct the two chromosomes in the hybrids according to their parental origin. We found that, unlike genetic inheritance, epigenetic heritability is quite variable. Cytosines were found to be differentially methylated (epimutated) at a rate of 7.48% (1∕15 cytosines) between the NPB and 93-11 parental strains. We also observed that 0.79% of cytosines were epimutated between the parent and corresponding hybrid chromosome. We found that these epimutations are often clustered on the chromosomes, with clusters representing 20% of all epimutations between parental ecotypes, and 2-5% in F1 plants. Epimutation clusters are also strongly associated with regions where the production of siRNA differs between parents. Finally, we identified genes with both allele-specific expression patterns that were strongly inherited as well as those differentially expressed between hybrids and the corresponding parental chromosome. We conclude that much of the misinheritance of expression levels is likely caused by epimutations and trans effects.bioinformatics | plant biology D NA methylation is an epigenetic mark that can often lead to the repression of gene expression (1, 2). It is enriched in heterochromatin and, when present at regulatory sites, usually acts as a repressor of expression, most notably in transposons (3). However, it is also found over coding regions, where it likely does not directly affect transcription and is associated with moderately expressed genes (2, 4-6). In plants, DNA methylation occurs in three different contexts: CG, CHG, and CHH (where H is any nucleotide but G). In Arabidopsis, each context is maintained by different enzymes: MET1 for CG sites, CMT3 for CHG sites and DRM2 for CHH sites. CG and CHG sites are symmetric across the two DNA strands, which is thought to be important for the maintenance of methylation at these sites following DNA replication. In contrast, CHH sites are not symmetric, and their methylation is mediated by RNA-directed DNA methylation pathways (RdDM), which use siRNAs to initiate de novo methylation (3). Cellular methylation states tend to persist during cell division, and recent studies in Arabidopsis have also shown that DNA methylation is faithfully inherited across generations (7,8). Nonetheless, we are only beginning to understand how different...

Background Lichens, encompassing 20,000 known species, are symbioses between specialized fungi (mycobionts), mostly ascomycetes, and unicellular green algae or cyanobacteria (photobionts). Here we describe the first parallel genomic analysis of the mycobiont Cladonia grayi and of its green algal photobiont Asterochloris glomerata . We focus on genes/predicted proteins of potential symbiotic significance, sought by surveying proteins differentially activated during early stages of mycobiont and photobiont interaction in coculture, expanded or contracted protein families, and proteins with differential rates of evolution. Results A) In coculture, the fungus upregulated small secreted proteins, membrane transport proteins, signal transduction components, extracellular hydrolases and, notably, a ribitol transporter and an ammonium transporter, and the alga activated DNA metabolism, signal transduction, and expression of flagellar components. B) Expanded fungal protein families include heterokaryon incompatibility proteins, polyketide synthases, and a unique set of G-protein α subunit paralogs. Expanded algal protein families include carbohydrate active enzymes and a specific subclass of cytoplasmic carbonic anhydrases. The alga also appears to have acquired by horizontal gene transfer from prokaryotes novel archaeal ATPases and Desiccation-Related Proteins. Expanded in both symbionts are signal transduction components, ankyrin domain proteins and transcription factors involved in chromatin remodeling and stress responses. The fungal transportome is contracted, as are algal nitrate assimilation genes. C) In the mycobiont, slow-evolving proteins were enriched for components involved in protein translation, translocation and sorting. Conclusions The surveyed genes affect stress resistance, signaling, genome reprogramming, nutritional and structural interactions. The alga carries many genes likely transferred horizontally through viruses, yet we found no evidence of inter-symbiont gene transfer. The presence in the photobiont of meiosis-specific genes supports the notion that sexual reproduction occurs in Asterochloris while they are free-living, a phenomenon with implications for the adaptability of lichens and the persistent autonomy of the symbionts. The diversity of the genes affecting the symbiosis suggests that lichens evolved by accretion of many scattered regulatory and structural changes rather than through introduction of a few key innovations. This predicts that paths to lichenization were variable in different phyla, which is consistent with the emerging consensus that ascolichens could have had a few independent origins. Electronic supplementary material The online version of this article (10.1186/s12864-019-5629-x) contains supplementary material, which is available to authorized users.

; 0000-0001-9355-9564 (M.P.).The green alga Chlamydomonas reinhardtii undergoes gametogenesis and mating upon nitrogen starvation. While the steps involved in its sexual reproductive cycle have been extensively characterized, the genome-wide transcriptional and epigenetic changes underlying different life cycle stages have yet to be fully described. Here, we performed transcriptome and methylome sequencing to quantify expression and DNA methylation from vegetative and gametic cells of each mating type and from zygotes. We identified 361 gametic genes with mating type-specific expression patterns and 627 genes that are specifically induced in zygotes; furthermore, these sex-related gene sets were enriched for secretory pathway and alga-specific genes. We also examined the C. reinhardtii nuclear methylation map with base-level resolution at different life cycle stages. Despite having low global levels of nuclear methylation, we detected 23 hypermethylated loci in gene-poor, repeat-rich regions. We observed mating type-specific differences in chloroplast DNA methylation levels in plus versus minus mating type gametes followed by chloroplast DNA hypermethylation in zygotes. Lastly, we examined the expression of candidate DNA methyltransferases and found three, DMT1a, DMT1b, and DMT4, that are differentially expressed during the life cycle and are candidate DNA methylases. The expression and methylation data we present provide insight into cell type-specific transcriptional and epigenetic programs during key stages of the C. reinhardtii life cycle.Chlamydomonas reinhardtii is a unicellular, biflagellate species of green alga found primarily in fresh water and soil (Harris et al., 2009). C. reinhardtii is an important reference organism for diverse eukaryotic cellular and metabolic processes, including photosynthetic biology (Rochaix, 2001), flagellar function and biogenesis (Silflow and Lefebvre, 2001), nutrient homeostasis (Grossman, 2000;Merchant et al., 2006;Glaesener et al., 2013), and sexual cycles (Goodenough et al., 2007). The nuclear and chloroplast genomes of C. reinhardtii have been fully sequenced, enabling genomic and epigenomic analyses (Maul et al., 2002;Merchant et al., 2007). The approximately 112-Mb haploid C. reinhardtii nuclear genome comprises 17 chromosomes. The circular chloroplast DNA (cpDNA) genome is 203 kb and present in 80 to 100 copies per cell that are organized into eight to 10 nucleoprotein complexes called nucleoids, which are distributed through the stroma.Like many unicellular eukaryotes, C. reinhardtii has a biphasic life cycle where haploid cells can reproduce vegetatively by mitotic division or, alternatively, undergo a sexual cycle. Vegetative cells can propagate indefinitely when provided with nutrients and light. Upon nitrogen starvation, however, cells stop dividing and differentiate into gametes whose mating type (plus or minus) is determined genetically by an approximately 300-kb mating type locus on chromosome 6, with two haplotypes, MT+ and MT2 (Umen, 2011; De Hoff et al.,...

Summary Regulatory T cells (T reg cells) deficient in the transcription factor Foxp3 lack suppressor function and manifest a T effector (T eff ) cell-like phenotype. We demonstrate that Foxp3 deficiency dysregulates metabolic checkpoint kinase mTORC2 signaling and gives rise to augmented aerobic glycolysis and oxidative phosphorylation. Specific deletion of the mTORC2 adaptor gene Rictor in Foxp3-deficient T reg cells ameliorated disease in a Foxo1 transcription factor-dependent manner. Rictor deficiency reestablished a subset of T reg cell genetic circuits and suppressed the T eff cell-like glycolytic and respiratory programs, which contributed to immune dysregulation. Treatment of T reg cells from patients with FOXP3 deficiency with mTOR inhibitors similarly antagonized their T eff cell-like program and restored suppressive function. Thus, regulatory function can be reestablished in Foxp3-deficient T reg cells by targeting their metabolic pathways, providing opportunities to restore tolerance in T reg cell disorders.