Cycads are the only early seed plants that have evolved a specialized root to host endophytic bacteria that fix nitrogen. To provide evolutionary and functional insights into this million-year old symbiosis, we investigate endophytic bacterial sub-communities isolated from coralloid roots of species from Dioon (Zamiaceae) sampled from their natural habitats. We employed a sub-community co-culture experimental strategy to reveal both predominant and rare bacteria, which were characterized using phylogenomics and detailed metabolic annotation. Diazotrophic plant endophytes, including Bradyrhizobium, Burkholderia, Mesorhizobium, Rhizobium, and Nostoc species, dominated the epiphyte-free sub-communities. Draft genomes of six cyanobacteria species were obtained after shotgun metagenomics of selected sub-communities. These data were used for whole-genome inferences that suggest two Dioon-specific monophyletic groups, and a level of specialization characteristic of co-evolved symbiotic relationships. Furthermore, the genomes of these cyanobacteria were found to encode unique biosynthetic gene clusters, predicted to direct the synthesis of specialized metabolites, mainly involving peptides. After combining genome mining with detection of pigment emissions using multiphoton excitation fluorescence microscopy, we also show that Caulobacter species co-exist with cyanobacteria, and may interact with them by means of a novel indigoidine-like specialized metabolite. We provide an unprecedented view of the composition of the cycad coralloid root, including phylogenetic and functional patterns mediated by specialized metabolites that may be important for the evolution of ancient symbiotic adaptations.

Long noncoding RNAs (lncRNAs) are essential regulatory elements of sex chromosomes that act to equalize gene expression levels between males and females.XIST,RSX, androX2regulate X chromosomes in placental mammals, marsupials, andDrosophila, respectively. Because the green anole (Anolis carolinensis) shows complete dosage compensation of its X chromosome, we tested whether a lncRNA was involved. We found an ancient lncRNA,MAYEX, that gained male-specific expression more than 89 million years ago.MAYEXevolved a notable association with the acetylated histone 4 lysine 16 (H4K16ac) epigenetic mark and the ability to loop its locus to the totality of the X chromosome to increase expression levels.MAYEXis the first lncRNA in reptiles linked to a dosage compensation mechanism that balances the expression of sex chromosomes.

LncRNAs are essential regulators of eukaryotic gene expression. They exert their gene regulatory functions by interacting with DNA, RNA, and protein. These functions are considered at least in part associated with their capacity to fold into complex three-dimensional structures. The conservation of lncRNA structure in mammalian genomes has been assessed in several studies, however, very little is known about the conservation of lncRNA structures in plants. Here, we analyze the structural conservation of lncRNAs in Brassicaceae , using a whole genome alignment of 16 Brassicaceae species. We found that 44.2% (1925 of 4354) of the intergenic lncRNAs (lincRNAs) and 75.1% (1549 of 2060) of the natural antisense transcripts (NATs) of Arabidopsis thaliana have conserved structural motifs in at least 2 of the 16 species. Also, 3612 lncRNAs have conserved structural motifs in multiple species; 2264 of which are tissue-specific, and 841 can be associated with a function by a co-expression network in A. thaliana . Indeed, we find evidence for the conservation of structural motifs in several lncRNAs with known functions, including, lncCOBRA1, FLORE, IPS1, ELENA1 and COOLAIR. The latter was shown previously to have a conserved structure. Overall, we have identified numerous lncRNAs with conserved structures in Brassicaceae that warrant further experimental exploration in vivo to understand whether these lncRNAs and their conserved structures are of biological significance.

Long non-coding RNAs (lncRNAs) have recently emerged as prominent regulators of gene expression in eukaryotes. LncRNAs often drive the modification and maintenance of gene activation or gene silencing states via chromatin conformation rearrangements. In plants, lncRNAs have been shown to participate in gene regulation, and are essential to processes such as vernalization and photomorphogenesis. Despite their prominent functions only over a dozen lncRNAs have been experimentally and functionally characterized. Similar to its animal counterparts, the rates of sequence divergence are much higher in plant lncRNAs than in protein coding mRNAs, making it difficult to identify lncRNA conservation using traditional sequence comparison methods. Beyond this, little is known about the evolutionary patterns of lncRNAs in plants. Here, we characterized the splicing conservation of lncRNAs in Brassicaceae. We generated a whole-genome alignment of 16 Brassica species and used it to identify synthenic lncRNA orthologues. Using a scoring system trained on transcriptomes from A. thaliana and B. oleracea , we identified splice sites across the whole alignment and measured their conservation. Our analysis revealed that 17.9% (112/627) of all intergenic lncRNAs display splicing conservation in at least one exon, an estimate that is substantially higher to previous estimates of lncRNA conservation in this group. Our findings agree with similar studies in vertebrates, demonstrating that splicing conservation can be evidence of stabilizing selection. We provide conclusive evidence for the existence of evolutionary deeply conserved lncRNAs in plants and describe a generally applicable computational workflow to identify functional lncRNAs in plants.

Cycads are the only early seed plants that have evolved a specialized root to host endophytic bacteria that fix nitrogen. To provide evolutionary and functional insights into this million-year old symbiosis, we investigate endophytic bacterial sub-communities isolated from coralloid roots of species from Dioon (Zamiaceae) sampled from their natural habitats. We employed a sub-community co-culture experimental strategy to reveal both predominant and rare bacteria, which were characterized using phylogenomics and detailed metabolic annotation. Diazotrophic plant endophytes, including Bradyrhizobium, Burkholderia, Mesorhizobium, Nostoc , and Rhizobium species, dominated the epiphyte-free sub-communities. Draft genomes of six cyanobacteria species were obtained after shotgun metagenomics of selected sub-communities and used for whole-genome inferences that suggest two Dioon- specific monophyletic groups and a level of specialization characteristic of co-evolved symbiotic relationships. In agreement with this, the genomes of these cyanobacteria were found to encode unique biosynthetic gene clusters, predicted to direct the synthesis of specialized metabolites, mainly involving peptides. After combining genome mining with metabolite profiling using multiphoton excitation fluorescence microscopy, we also show that Caulobacter species co-exist with cyanobacteria, and may interact with them by means of a novel indigoidine-like specialized metabolite. We provide an unprecedented view of the composition of the cycad coralloid root, including phylogenetic and functional patterns mediated by specialized metabolites that may be important for the evolution of ancient symbiotic adaptations.

Lasiodiplodia theobromae is a pathogenic fungus associated with tropical perennial fruit plants worldwide. In apple trees, L. theobromae causes dieback and canker, a disease that affects the architecture of the wood producing the progressive death of branches and stems, from the tips to the base, invading the vascular tissue, manifesting necrotic lesions in the bark, impeding the flow of nutrients and water. The present work reports the whole genome de novo sequencing (WGS) of L. theobromae strain Bot-2018-LT45 isolated from apple trees with dieback symptoms. Genomic DNA of L. theobromae was sequenced using Illumina paired-end short-read technology (NovaSeq6000) and PacBio SMRTbell^TM (Single Molecule, Real-Time) long-read technology. The genome size was 44.17 Mb. Then, assembly and annotation revealed a total of 12,948 genes of which 11,634 encoded proteins. The genome was assembled into 34 contigs with an N50 (Mb) value of 3.23. This study is the first report of the L. theobromae genome de novo obtained from apple trees with dieback and canker symptoms in the Maule Region, Chile. This genetic information may set the basis for future study of the mechanisms of L. theobromae and establish the possibility of specific molecular improvements for the control of dieback and canker.