The brown alga Ectocarpus siliculosus has a haploid–diploid life cycle that involves an alternation between two distinct generations, the sporophyte and the gametophyte. We describe a mutant, ouroboros ( oro ), in which the sporophyte generation is converted into a functional, gamete-producing gametophyte. The life history of the mutant thus consists of a continuous reiteration of the gametophyte generation. The oro mutant exhibited morphological features typical of the gametophyte generation and accumulated transcripts of gametophyte generation marker genes. Genetic analysis showed that oro behaved as a single, recessive, Mendelian locus that was unlinked to the IMMEDIATE UPRIGHT locus, which has been shown to be necessary for full expression of the sporophyte developmental program. The data presented here indicate that ORO is a master regulator of the gametophyte-to-sporophyte life cycle transition and, moreover, that oro represents a unique class of homeotic mutation that results in switching between two developmental programs that operate at the level of the whole organism.

Rhizobia can infect roots of host legume plants and induce new organs called nodules, in which they fix atmospheric nitrogen. Infection generally starts with root hair curling, then proceeds inside newly formed, intracellular tubular structures called infection threads. A successful symbiotic interaction relies on infection threads advancing rapidly at their tips by polar growth through successive cell layers of the root toward developing nodule primordia. To identify a plant component that controls this tip growth process, we characterized a symbiotic mutant of Medicago truncatula, called rpg for rhizobium-directed polar growth. In this mutant, nitrogen-fixing nodules were rarely formed due to abnormally thick and slowly progressing infection threads. Root hair curling was also abnormal, indicating that the RPG gene fulfils an essential function in the process whereby rhizobia manage to dominate the process of induced tip growth for root hair infection. Map-based cloning of RPG revealed a member of a previously unknown plant-specific gene family encoding putative long coiled-coil proteins we have called RRPs (RPG-related proteins) and characterized by an ''RRP domain'' specific to this family. RPG expression was strongly associated with rhizobial infection, and the RPG protein showed a nuclear localization, indicating that this symbiotic gene constitutes an important component of symbiotic signaling.genetics ͉ symbiosis ͉ coiled-coil I n the symbiotic interaction between legumes and soil bacteria called rhizobia, nitrogen-fixing nodules are formed that allow plant growth to be independent of added combined nitrogen, and the plant provides rhizobia with a carbon source derived from photosynthesis. During initial signal exchange in the rhizosphere, rhizobia respond to plant flavonoids by producing lipochito-oligosaccharidic molecules called Nod factors (NFs). Host-specific recognition of NFs triggers a controlled infection leading to rhizobial internalization, and the induction of a new plant organ, the nodule, in which nitrogen fixation occurs (1).Rhizobial infection of host legumes is generally via root hairs (RHs) that undergo marked curling. Compared with normal RH tip growth in which vesicles, containing cell wall and membrane material, travel in an actin-and microtubule-dependent fashion to the RH tip, where they fuse with the cell membrane (2), this rhizobium-induced growth reorientation involves alterations to the plant cytoskeleton and the redirection of vesicle traffic away from the RH tip to a new site (3, 4). Inside a closed chamber formed by root hair curling (RHC), the plant cell wall is locally degraded and the plasma membrane becomes invaginated. Rhizobia enter a newly formed, plant-derived structure, the infection thread (IT), that undergoes inward tip growth within the RH. Underlying outer cortical cells change into highly polarized pre-IT cells, which guide IT passage to the nodule primordium formed, in M. truncatula, in the inner root cortex (3). Here, bacteria are released into plant cells an...

Summary 24• The genome of the filamentous brown alga Ectocarpus was the first to be completely 25 sequenced from within the brown algal group and has served as a key reference genome both 26 for this lineage and for the stramenopiles. 27• We present a complete structural and functional reannotation of the Ectocarpus genome. 28• The large-scale assembly of the Ectocarpus genome was significantly improved and genome-29 wide gene re-annotation using extensive RNA-seq data improved the structure of 11,108 30 existing protein-coding genes and added 2,030 new loci. A genome-wide analysis of splicing 31 isoforms identified an average of 1.6 transcripts per locus. A large number of previously 32

Three amino acid loop extension homeodomain transcription factors (TALE HD TFs) act as life cycle regulators in green algae and land plants. In mosses these regulators are required for the deployment of the sporophyte developmental program. We demonstrate that mutations in either of two TALE HD TF genes, OUROBOROS or SAMSARA, in the brown alga Ectocarpus result in conversion of the sporophyte generation into a gametophyte. The OUROBOROS and SAMSARA proteins heterodimerise in a similar manner to TALE HD TF life cycle regulators in the green lineage. These observations demonstrate that TALE-HD-TF-based life cycle regulation systems have an extremely ancient origin, and that these systems have been independently recruited to regulate sporophyte developmental programs in at least two different complex multicellular eukaryotic supergroups, Archaeplastida and Chromalveolata.

Brown algae are one of the most developmentally complex groups within the eukaryotes. As in many land plants and animals, their main body axis is established early in development, when the initial cell gives rise to two daughter cells that have apical and basal identities, equivalent to shoot and root identities in land plants, respectively. We show here that mutations in the Ectocarpus DISTAG (DIS) gene lead to loss of basal structures during both the gametophyte and the sporophyte generations. Several abnormalities were observed in the germinating initial cell in dis mutants, including increased cell size, disorganization of the Golgi apparatus, disruption of the microtubule network, and aberrant positioning of the nucleus. DIS encodes a TBCCd1 protein, which has a role in internal cell organization in animals, Chlamydomonas reinhardtii, and trypanosomes. Our study highlights the key role of subcellular events within the germinating initial cell in the determination of apical/basal cell identities in a brown alga and emphasizes the remarkable functional conservation of TBCCd1 in regulating internal cell organization across extremely distant eukaryotic groups.

The life cycle of an organism is one of its fundamental features, influencing many aspects of its biology. The brown algae exhibit a diverse range of life cycles indicating that transitions between life cycle types may have been key adaptive events in the evolution of this group. Life cycle mutants, identified in the model organism Ectocarpus, are providing information about how life cycle progression is regulated at the molecular level in brown algae. We explore some of the implications of the phenotypes of the life cycle mutants described to date and draw comparisons with recent insights into life cycle regulation in the green lineage. Given the importance of coordinating growth and development with life cycle progression, we suggest that the co-option of ancient life cycle regulators to control key developmental events may be a common feature in diverse groups of multicellular eukaryotes.

Hi-C exploits contact frequencies between pairs of loci to bridge and order contigs during genome assembly, resulting in chromosome-level assemblies. Because few robust programs are available for this type of data, we developed instaGRAAL, a complete overhaul of the GRAAL program, which has adapted the latter to allow efficient assembly of large genomes. instaGRAAL features a number of improvements over GRAAL, including a modular correction approach that optionally integrates independent data. We validate the program using data for two brown algae, and human, to generate near-complete assemblies with minimal human intervention.

The Medicago truncatula DMI3 gene encodes a calcium- and calmodulin-dependent protein kinase (CCaMK) that is necessary for the establishment of both rhizobial and mycorrhizal symbioses. The two symbiotic signaling pathways diverge downstream of DMI3; therefore, it has been proposed that legumes have evolved a particular form of CCaMK, acting like a switch able both to discriminate between rhizobial and mycorrhizal calcium signatures and to trigger the appropriate downstream signaling pathway. To test this hypothesis, we examined whether a CCaMK gene from a nonlegume species was able to restore the rhizobial symbiotic properties of a M. truncatula dmi3 mutant. Our results show that a CCaMK gene from rice can restore nodule formation, indicating that CCaMKs from nonlegumes can interpret the calcium signature elicited by rhizobial Nod factors and activate the appropriate downstream target. The nodules did not contain bacteria, which suggests that DMI3 is also involved in the control of the infection process.