Computational prediction and experimental validation of microRNAs in the brown alga Ectocarpus siliculosus.

We used an in silico approach to predict microRNAs (miRNAs) genome-wide in the brown alga Ectocarpus siliculosus. As brown algae are phylogenetically distant from both animals and land plants, our approach relied on features shared by all known organisms, excluding sequence conservation, genome localization and pattern of base-pairing with the target. We predicted between 500 and 1500 miRNAs candidates, depending on the values of the energetic parameters used to filter the potential precursors. Using quantitative polymerase chain reaction assays, we confirmed the existence of 22 miRNAs among 72 candidates tested, and of 8 predicted precursors. In addition, we compared the expression of miRNAs and their precursors in two life cycle states (sporophyte, gametophyte) and under salt stress. Several miRNA precursors, Argonaute and DICER messenger RNAs were differentially expressed in these conditions. Finally, we analyzed the gene organization and the target functions of the predicted candidates. This showed that E. siliculosus miRNA genes are, like plant miRNA genes, rarely clustered and, like animal miRNA genes, often located in introns. Among the predicted targets, several widely conserved functional domains are significantly overrepresented, like kinesin, nucleotide-binding/APAF-1, R proteins and CED-4 (NB-ARC) and tetratricopeptide repeats. The combination of computational and experimental approaches thus emphasizes the originality of molecular and cellular processes in brown algae.

Genome structure and metabolic features in the red seaweed Chondrus crispus shed light on evolution of the Archaeplastida.

Red seaweeds are key components of coastal ecosystems and are economically important as food and as a source of gelling agents, but their genes and genomes have received little attention. Here we report the sequencing of the 105-Mbp genome of the florideophyte Chondrus crispus (Irish moss) and the annotation of the 9,606 genes. The genome features an unusual structure characterized by gene-dense regions surrounded by repeat-rich regions dominated by transposable elements. Despite its fairly large size, this genome shows features typical of compact genomes, e.g., on average only 0.3 introns per gene, short introns, low median distance between genes, small gene families, and no indication of large-scale genome duplication. The genome also gives insights into the metabolism of marine red algae and adaptations to the marine environment, including genes related to halogen metabolism, oxylipins, and multicellularity (microRNA processing and transcription factors). Particularly interesting are features related to carbohydrate metabolism, which include a minimalistic gene set for starch biosynthesis, the presence of cellulose synthases acquired before the primary endosymbiosis showing the polyphyly of cellulose synthesis in Archaeplastida, and cellulases absent in terrestrial plants as well as the occurrence of a mannosylglycerate synthase potentially originating from a marine bacterium. To explain the observations on genome structure and gene content, we propose an evolutionary scenario involving an ancestral red alga that was driven by early ecological forces to lose genes, introns, and intergenetic DNA; this loss was followed by an expansion of genome size as a consequence of activity of transposable elements.

Selection drives convergent gene expression changes during transitions to co-sexuality in haploid sexual systems.

Co-sexuality has evolved repeatedly from unisexual (dioicous) ancestors across a wide range of taxa. However, the molecular changes underpinning this important transition remain unknown, particularly in organisms with haploid sexual systems such as bryophytes, red algae and brown algae. Here we explore four independent events of emergence of co-sexuality from unisexual ancestors in brown algal clades to examine the nature, evolution and degree of convergence of gene expression changes that accompany the breakdown of dioicy. The amounts of male versus female phenotypic differences in dioicous species were not correlated with the extent of sex-biased gene expression, in stark contrast to what is observed in animals. Although sex-biased genes exhibited a high turnover rate during brown alga diversification, some of their predicted functions were conserved across species. Transitions to co-sexuality consistently involved adaptive gene expression shifts and rapid sequence evolution, particularly for male-biased genes. Gene expression in co-sexual species was more similar to that in females rather than males of related dioicous species, suggesting that co-sexuality may have arisen from ancestral females. Finally, extensive convergent gene expression changes, driven by selection, were associated with the transition to co-sexuality. Together, our observations provide insights on how co-sexual systems arise from ancestral, haploid UV sexual systems.

Localization of causal locus in the genome of the brown macroalga Ectocarpus: NGS-based mapping and positional cloning approaches.

Mutagenesis is the only process by which unpredicted biological gene function can be identified. Despite that several macroalgal developmental mutants have been generated, their causal mutation was never identified, because experimental conditions were not gathered at that time. Today, progresses in macroalgal genomics and judicious choices of suitable genetic models make mutated gene identification possible. This article presents a comparative study of two methods aiming at identifying a genetic locus in the brown alga Ectocarpus siliculosus: positional cloning and Next-Generation Sequencing (NGS)-based mapping. Once necessary preliminary experimental tools were gathered, we tested both analyses on an Ectocarpus morphogenetic mutant. We show how a narrower localization results from the combination of the two methods. Advantages and drawbacks of these two approaches as well as potential transfer to other macroalgae are discussed.

Space-time decoupling in the branching process in the mutant étoile of the filamentous brown alga Ectocarpus siliculosus.

Ectocarpus siliculosus is being developed as a model organism for brown algal genetics and genomics. Brown algae are phylogenetically distant from the other multicellular phyla (green lineage, red algae, fungi and metazoan) and therefore might offer the opportunity to study novel and alternative developmental processes that lead to the establishment of multicellularity. E. siliculosus develops as uniseriate filaments, thereby displaying one of the simplest architectures among multicellular organisms. The young sporophyte grows as a primary filament and then branching occurs, preferentially at the center of the filament. We recently described the first morphogenetic mutant étoile (etl) in a brown alga, produced by UVB mutagenesis in E. siliculosus. We showed that a single recessive mutation was responsible for a defect in both cell differentiation and the very early branching pattern (first and second branch emergences). Here, we supplement this study by reporting the branching defects observed subsequently, i.e. for the later stages corresponding to the emergence of up to the first six secondary filaments, and we show that the branching process is composed of at least two distinct components: time and position.