A genetic analysis of the biological clock in Chlamydomonas reinhardi has been initiated. Of six wild-type strains tested (3 mt(+) and 3 mt(-)), five had periods close to 24 hr whereas one had a 21-hr period. Mutants with altered clock period have been isolated. The periods of 3 of these variant strains are temperature compensated. Genetic crosses involving a long-period mutant suggest that a single gene confers the long-period character, and in general clock-period length seems to be a useful phenotypic measure of alterations in the clock due to genetic differences. One phase mutant was found but its behavior was variable and the phase of the rhythm, relative to a light-dark transition which initiates the rhythm, does not seem to be reliable as a parameter of clock differences. No markers have yet been mapped.

The alignment of sequencing reads against a protein reference database is a major computational bottleneck in metagenomics and data-intensive evolutionary projects. Although recent tools offer improved performance over the gold standard BLASTX, they exhibit only a modest speedup or low sensitivity. We introduce DIAMOND, an open-source algorithm based on double indexing that is 20,000 times faster than BLASTX on short reads and has a similar degree of sensitivity.

Chlamydomonas reinhardtii expresses a well-documented circadian rhythm of phototaxis, which peaks in the subjective daytime. We find that vegetative cells also express circadian rhythms of chemotaxis to ammonium and ammonium uptake (as gauged by uptake of [(14)C]methylammonium). The chemotaxis rhythm peaks in the subjective night. Methylammonium uptake is light dependent, and its rhythm peaks at subjective dawn. Unlike vegetative cells, gametes are not attracted to ammonium. We believe this to be the first report of a circadian rhythm of chemotaxis.

Correct circadian regulation increases plant productivity, and photosynthesis is circadian-regulated. Here, we discuss the regulatory basis for the circadian control of photosynthesis. We discuss candidate mechanisms underpinning circadian oscillations of light harvesting and consider how the circadian clock modulates CO2 fixation by Rubisco. We show that new techniques may provide a platform to better understand the signalling pathways that couple the circadian clock with the photosynthetic apparatus. Finally, we discuss how understanding circadian regulation in model systems is underpinning research into the impact of circadian regulation in crop species.

Here, we present a major advance of the OrthoFinder method. This extends OrthoFinder's high accuracy orthogroup inference to provide phylogenetic inference of orthologs, rooted gene trees, gene duplication events, the rooted species tree, and comparative genomics statistics. Each output is benchmarked on appropriate real or simulated datasets, and where comparable methods exist, OrthoFinder is equivalent to or outperforms these methods. Furthermore, OrthoFinder is the most accurate ortholog inference method on the Quest for Orthologs benchmark test. Finally, OrthoFinder's comprehensive phylogenetic analysis is achieved with equivalent speed and scalability to the fastest, score-based heuristic methods. OrthoFinder is available at https://github.com/davidemms/OrthoFinder.

Circadian clocks affect a large proportion of differentially expressed genes in many organisms. Tissue-specific hierarchies in circadian networks in mammals have been contentiously debated, whereas little attention has been devoted to the concept in plants, owing to technical difficulties. Recently, several studies have demonstrated tissue-specific circadian clocks and their coupling in plants, suggesting that plants possess a hierarchical network of circadian clocks. The following review summarizes recent studies describing the tissue-specific functions and properties of these circadian clocks and discusses the network structure and potential messengers that might share temporal information on such a network. Copyright © 2015 Elsevier Ltd. All rights reserved.

Plants have adapted to the diurnal light-dark cycle by establishing elaborate transcriptional programs that coordinate many metabolic, physiological, and developmental responses to the external environment. These transcriptional programs have been studied in only a few species, and their function and conservation across algae and plants is currently unknown. We performed a comparative transcriptome analysis of the diurnal cycle of nine members of Archaeplastida, and we observed that, despite large phylogenetic distances and dramatic differences in morphology and lifestyle, diurnal transcriptional programs of these organisms are similar. Expression of genes related to cell division and the majority of biological pathways depends on the time of day in unicellular algae but we did not observe such patterns at the tissue level in multicellular land plants. Hence, our study provides evidence for the universality of diurnal gene expression and elucidates its evolutionary history among different photosynthetic eukaryotes.

Circadian oscillators are known to regulate the timing of cell division in many organisms. In the case of Chlamydomonas reinhardtii, however, this conclusion has been challenged by several investigators. We have reexamined this issue and find that the division behavior of Chlamydomonas meets all the criteria for circadian rhythmicity: persistence of a cell division rhythm (a) with a period of approximately 24 h under free-running conditions, (b) that is temperature compensated, and (c) which can entrain to light/dark signals. In addition, a mutation that lengthens the circadian period of the phototactic rhythm similarly affects the cell division rhythm. We conclude that a circadian mechanism determines the timing of cell division in Chlamydomonas reinhardtii.

Massively parallel sequencing of cDNA has enabled deep and efficient probing of transcriptomes. Current approaches for transcript reconstruction from such data often rely on aligning reads to a reference genome, and are thus unsuitable for samples with a partial or missing reference genome. Here we present the Trinity method for de novo assembly of full-length transcripts and evaluate it on samples from fission yeast, mouse and whitefly, whose reference genome is not yet available. By efficiently constructing and analyzing sets of de Bruijn graphs, Trinity fully reconstructs a large fraction of transcripts, including alternatively spliced isoforms and transcripts from recently duplicated genes. Compared with other de novo transcriptome assemblers, Trinity recovers more full-length transcripts across a broad range of expression levels, with a sensitivity similar to methods that rely on genome alignments. Our approach provides a unified solution for transcriptome reconstruction in any sample, especially in the absence of a reference genome.

The plant circadian clock coordinates the responses to multiple and often simultaneous environmental challenges that the sessile plant cannot avoid. These responses must be integrated efficiently into dynamic metabolic and physiological networks essential for growth and reproduction. Many of the output pathways regulated by the circadian clock feed back to modulate clock function, leading to the appreciation of the clock as a central hub in a sophisticated regulatory network. In this Review, we discuss the circadian regulation of growth, flowering time, abiotic and biotic stress responses, and metabolism, as well as why temporal 'gating' of these processes is important to plant fitness.

Background: RNA-Seq is revolutionizing the way transcript abundances are measured. A key challenge in transcript quantification from RNA-Seq data is the handling of reads that map to multiple genes or isoforms. This issue is particularly important for quantification with de novo transcriptome assemblies in the absence of sequenced genomes, as it is difficult to determine which transcripts are isoforms of the same gene. A second significant issue is the design of RNA-Seq experiments, in terms of the number of reads, read length, and whether reads come from one or both ends of cDNA fragments.;Results: We present RSEM, an user-friendly software package for quantifying gene and isoform abundances from single-end or paired-end RNA-Seq data. RSEM outputs abundance estimates, 95% credibility intervals, and visualization files and can also simulate RNA-Seq data. In contrast to other existing tools, the software does not require a reference genome. Thus, in combination with a de novo transcriptome assembler, RSEM enables accurate transcript quantification for species without sequenced genomes. On simulated and real data sets, RSEM has superior or comparable performance to quantification methods that rely on a reference genome. Taking advantage of RSEM's ability to effectively use ambiguously-mapping reads, we show that accurate gene-level abundance estimates are best obtained with large numbers of short single-end reads. On the other hand, estimates of the relative frequencies of isoforms within single genes may be improved through the use of paired-end reads, depending on the number of possible splice forms for each gene.;Conclusions: RSEM is an accurate and user-friendly software tool for quantifying transcript abundances from RNA-Seq data. As it does not rely on the existence of a reference genome, it is particularly useful for quantification with de novo transcriptome assemblies. In addition, RSEM has enabled valuable guidance for cost-efficient design of quantification experiments with RNA-Seq, which is currently relatively expensive.

Cell division often occurs at specific times of the day in animal and photosynthetic organisms. Studies in unicellular photosynthetic algae, such as Chlamydomonas or Euglena, have shown that the photoperiodic control of cell division is mediated through the circadian clock. However, the underlying mechanisms remain unknown. We have studied the molecular basis of light-dependent control of cell division in the unicellular green alga Ostreococcus. We found that cell division obeys a circadian oscillator in Ostreococcus. We provide evidence suggesting that the clock may, at least in part, regulate directly cell division independently of the metabolism. Combined microarray and quantitative real-time reverse transcription-polymerase chain reaction analysis of the main core cell cycle gene expression revealed an extensive transcriptional regulation of cell division by the photoperiod in Ostreococcus. Finally, transcription of the main core cell cycle genes, including cyclins and cyclin-dependent kinases, was shown to be under circadian control in Ostreococcus, suggesting that these genes are potential targets of the circadian clock in the control of cell division.

Circadian timekeeping in plants increases photosynthesis and productivity. There are circadian oscillations in the abundance of many chloroplast-encoded transcripts, but it is not known how the circadian clock regulates chloroplast transcription or the photosynthetic apparatus. We show that, in Arabidopsis, nuclear-encoded SIGMA FACTOR5 (SIG5) controls circadian rhythms of transcription of several chloroplast genes, revealing one pathway by which the nuclear-encoded circadian oscillator controls rhythms of chloroplast gene expression. We also show that SIG5 mediates the circadian gating of light input to a chloroplast-encoded gene. We have identified an evolutionarily conserved mechanism that communicates circadian timing information between organelles with distinct genetic systems and have established a new level of integration between eukaryotic circadian clocks and organelles of endosymbiotic origin.

The evolution of circadian clocks in land plants is not understood, because circadian rhythms have received little attention in plants other than angiosperms. We have characterized two genes, PpCCA1a and PpCCA1b, homologs of the Arabidopsis thaliana clock genes CCA1/LHY, from the moss Physcomitrella patens. PpCCA1a and PpCCA1b, together with angiosperm CCA1/LHY homologs, belong to the clock-associated single-myb gene family of green plants (including green algae and land plants). The accumulation of PpCCA1a and PpCCA1b mRNA showed rhythms with a period of approximately 1 day, phased as are those of angiosperm homologs, under 24 h light/dark cycles or in continuous dark. However, in marked contrast to angiosperm homologs, both genes showed arrhythmic profiles in continuous light. The timing of the PpCCA1b peak is determined by the time of the last light to dark transition, suggesting that the arrhythmicity in continuous light is due to dysfunction of the core clock. We generated single and double disruptants for PpCCA1a and PpCCA1b, and found that the double disruptants showed: (i) short periodicity and damped amplitude in the PpCCA1b rhythm, (ii) similar changes in the rhythmically expressed genes PpSIG5 and PpPRRa, and (iii) de-repression of PpCCA1b transcription levels, indicating negative feedback regulation. These observations indicate that the two genes are not merely structural homologs but also functional counterparts of CCA1/LHY. Together, our results illustrate similarities as well as divergence of the clock machineries between P. patens and A. thaliana, two distantly placed species in land plant phylogeny.

We have investigated the role of the circadian clock in the regulation of expression of genes required for ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) synthesis, assembly, and activation. Circadian oscillations in RCA (the gene encoding Rubisco activase) and RBCS (the gene encoding Rubisco small subunit) mRNA accumulation, with peak abundance occurring soon after dawn, occur in Arabidopsis thaliana grown in a light-dark (LD) photoperiod. These oscillations persist in plants that have been transferred from LD to either continuous darkness (DD) or continuous light (LL). In contrast, CPN60[alpha] (the gene encoding [alpha]-chaperonin) and CPN60[beta] (the gene encoding [beta]-chaperonin) mRNA abundance oscillates in a diurnal, but not in a circadian, fashion. Although rapid damping of the circadian oscillation in RCA mRNA abundance is observed in Arabidopsis that have been grown in LD and then transferred to DD for 2 d, the circadian oscillations in RCA and RBCS mRNA abundance persist for at least five continuous cycles in LL, demonstrating the robustness of the circadian oscillator.

Molecular clocks drive oscillations in leaf photosynthesis, stomatal conductance, and other cell and leaf-level processes over ~24 h under controlled laboratory conditions. The influence of such circadian regulation over whole-canopy fluxes remains uncertain; diurnal CO and HO vapor flux dynamics in the field are currently interpreted as resulting almost exclusively from direct physiological responses to variations in light, temperature and other environmental factors. We tested whether circadian regulation would affect plant and canopy gas exchange at the Montpellier European Ecotron. Canopy and leaf-level fluxes were constantly monitored under field-like environmental conditions, and under constant environmental conditions (no variation in temperature, radiation, or other environmental cues).We show direct experimental evidence at canopy scales of the circadian regulation of daytime gas exchange: 20-79 % of the daily variation range in CO and HO fluxes occurred under circadian entrainment in canopies of an annual herb (bean) and of a perennial shrub (cotton). We also observed that considering circadian regulation improved performance by 8-17 % in commonly used stomatal conductance models.Our results show that circadian controls affect diurnal CO and HO flux patterns in entire canopies in field-like conditions, and its consideration significantly improves model performance. Circadian controls act as a 'memory' of the past conditions experienced by the plant, which synchronizes metabolism across entire plant canopies.

Stomata are pores that regulate plant gas exchange [1]. They evolved more than 400 million years ago [2, 3], but the origin of their active physiological responses to endogenous and environmental cues is unclear [2-6]. Recent research suggests that the stomata of lycophytes and ferns lack pore closure responses to abscisic acid (ABA) and CO(2). This evidence led to the hypothesis that a fundamental transition from passive to active control of plant water balance occurred after the divergence of ferns 360 million years ago [7, 8]. Here we show that stomatal responses of the lycophyte Selaginella [9] to ABA and CO(2) are directly comparable to those of the flowering plant Arabidopsis [10]. Furthermore, we show that the underlying intracellular signaling pathways responsible for stomatal aperture control are similar in both basal and modern vascular plant lineages. Our evidence challenges the hypothesis that acquisition of active stomatal control of plant carbon and water balance represents a critical turning point in land plant evolution [7, 8]. Instead, we suggest that the critical evolutionary development is represented by the innovation of stomata themselves and that physiologically active stomatal control originated at least as far back as the emergence of the lycophytes (circa 420 million years ago) [11].Copyright © 2011 Elsevier Ltd. All rights reserved.

Genome sequences from over 200 plant species have already been published, with this number expected to increase rapidly due to advances in sequencing technologies. Once a new genome has been assembled and the genes identified, the functional annotation of their putative translational products, proteins, using ontologies is of key importance as it places the sequencing data in a biological context. Furthermore, to keep pace with rapid production of genome sequences, this functional annotation process must be fully automated. Here we present a redesigned and significantly enhanced MapMan4 framework, together with a revised version of the associated online Mercator annotation tool. Compared with the original MapMan, the new ontology has been expanded almost threefold and enforces stricter assignment rules. This framework was then incorporated into Mercator4, which has been upgraded to reflect current knowledge across the land plant group, providing protein annotations for all embryophytes with a comparably high quality. The annotation process has been optimized to allow a plant genome to be annotated in a matter of minutes. The output results continue to be compatible with the established MapMan desktop application.Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.