Hiding in plain sight: Genome-wide recombination and a dynamic accessory genome drive diversity in Fusarium oxysporum f.sp. ciceris.

Understanding the origins of variation in agricultural pathogens is of fundamental interest and practical importance, especially for diseases that threaten food security. Fusarium oxysporum is among the most important of soil-borne pathogens, with a global distribution and an extensive host range. The pathogen is considered to be asexual, with horizontal transfer of chromosomes providing an analog of assortment by meiotic recombination. Here, we challenge those assumptions based on the results of population genomic analyses, describing the pathogen's diversity and inferring its origins and functional consequences in the context of a single, long-standing agricultural system. We identify simultaneously low nucleotide distance among strains, and unexpectedly high levels of genetic and genomic variability. We determine that these features arise from a combination of genome-scale recombination, best explained by widespread sexual reproduction, and presence-absence variation consistent with chromosomal rearrangement. Pangenome analyses document an accessory genome more than twice the size of the core genome, with contrasting evolutionary dynamics. The core genome is stable, with low diversity and high genetic differentiation across geographic space, while the accessory genome is paradoxically more diverse and unstable but with lower genetic differentiation and hallmarks of contemporary gene flow at local scales. We suggest a model in which episodic sexual reproduction generates haplotypes that are selected and then maintained through clone-like dynamics, followed by contemporary genomic rearrangements that reassort the accessory genome among sympatric strains. Taken together, these processes contribute unique genome content, including reassortment of virulence determinants that may explain observed variation in pathogenic potential.

Deep Sequencing of the Medicago truncatula Root Transcriptome Reveals a Massive and Early Interaction between Nodulation Factor and Ethylene Signals.

The legume-rhizobium symbiosis is initiated through the activation of the Nodulation (Nod) factor-signaling cascade, leading to a rapid reprogramming of host cell developmental pathways. In this work, we combine transcriptome sequencing with molecular genetics and network analysis to quantify and categorize the transcriptional changes occurring in roots of Medicago truncatula from minutes to days after inoculation with Sinorhizobium medicae. To identify the nature of the inductive and regulatory cues, we employed mutants with absent or decreased Nod factor sensitivities (i.e. Nodulation factor perception and Lysine motif domain-containing receptor-like kinase3, respectively) and an ethylene (ET)-insensitive, Nod factor-hypersensitive mutant (sickle). This unique data set encompasses nine time points, allowing observation of the symbiotic regulation of diverse biological processes with high temporal resolution. Among the many outputs of the study is the early Nod factor-induced, ET-regulated expression of ET signaling and biosynthesis genes. Coupled with the observation of massive transcriptional derepression in the ET-insensitive background, these results suggest that Nod factor signaling activates ET production to attenuate its own signal. Promoter:ß-glucuronidase fusions report ET biosynthesis both in root hairs responding to rhizobium as well as in meristematic tissue during nodule organogenesis and growth, indicating that ET signaling functions at multiple developmental stages during symbiosis. In addition, we identified thousands of novel candidate genes undergoing Nod factor-dependent, ET-regulated expression. We leveraged the power of this large data set to model Nod factor- and ET-regulated signaling networks using MERLIN, a regulatory network inference algorithm. These analyses predict key nodes regulating the biological process impacted by Nod factor perception. We have made these results available to the research community through a searchable online resource.

The TORMOZ gene encodes a nucleolar protein required for regulated division planes and embryo development in Arabidopsis.

Embryogenesis in Arabidopsis thaliana is marked by a predictable sequence of oriented cell divisions, which precede cell fate determination. We show that mutation of the TORMOZ (TOZ) gene yields embryos with aberrant cell division planes and arrested embryos that appear not to have established normal patterning. The defects in toz mutants differ from previously described mutations that affect embryonic cell division patterns. Longitudinal division planes of the proembryo are frequently replaced by transverse divisions and less frequently by oblique divisions, while divisions of the suspensor cells, which divide only transversely, appear generally unaffected. Expression patterns of selected embryo patterning genes are altered in the mutant embryos, implying that the positional cues required for their proper expression are perturbed by the misoriented divisions. The TOZ gene encodes a nucleolar protein containing WD repeats. Putative TOZ orthologs exist in other eukaryotes including Saccharomyces cerevisiae, where the protein is predicted to function in 18S rRNA biogenesis. We find that disruption of the Sp TOZ gene results in cell division defects in Schizosaccharomyces pombe. Previous studies in yeast and animal cells have identified nucleolar proteins that regulate the exit from M phase and cytokinesis, including factors involved in pre-rRNA processing. Our study suggests that in plant cells, nucleolar functions might interact with the processes of regulated cell divisions and influence the selection of longitudinal division planes during embryogenesis.

APETALA1 and SEPALLATA3 interact with SEUSS to mediate transcription repression during flower development.

The transcriptional repression of key regulatory genes is crucial for plant and animal development. Previously, we identified and isolated two Arabidopsis transcription co-repressors LEUNIG (LUG) and SEUSS (SEU) that function together in a putative co-repressor complex to prevent ectopic AGAMOUS (AG) transcription in flowers. Because neither LUG nor SEU possesses a recognizable DNA-binding motif, how they are tethered to specific target promoters remains unknown. Using the yeast two-hybrid assay and a co-immunoprecipitation assay, we showed that APETALA1 (AP1) and SEPALLATA3 (SEP3), both MADS box DNA-binding proteins, interacted with SEU. The AP1-SEU protein-protein interaction was supported by synergistic genetic interactions between ap1 and seu mutations. The role of SEU proteins in bridging the interaction between AP1/SEP3 and LUG to repress target gene transcription was further demonstrated in yeast and plant cells, providing important mechanistic insights into co-repressor function in plants. Furthermore, a direct in vivo association of SEU proteins with the AG cis-regulatory element was shown by chromatin immunoprecipitation. Accordingly, a reporter gene driven by the AG cis-element was able to respond to AP1- and SEP3-mediated transcriptional repression in a transient plant cell system when supplied with SEU and LUG. These results suggest that AP1 and SEP3 may serve as the DNA-binding partners of SEU/LUG. Our demonstration of the direct physical interaction between SEU and the C-terminal domain of SEP3 and AP1 suggests that AP1 and SEP3 MADS box proteins may interact with positive, as well as negative, regulatory proteins via their C-terminal domains, to either stimulate or repress their regulatory targets.

Transcriptional repression of target genes by LEUNIG and SEUSS, two interacting regulatory proteins for Arabidopsis flower development.

Transcription repression plays important roles in preventing crucial regulatory proteins from being expressed in inappropriate temporal or spatial domains. LEUNIG (LUG) and SEUSS (SEU) normally act to prevent ectopic expression of the floral homeotic gene AGAMOUS in flowers. LUG encodes a protein with sequence similarities to the yeast Tup1 corepressor. SEU encodes a plant-specific regulatory protein with sequence similarity in a conserved dimerization domain to the LIM-domain binding 1/Chip proteins in mouse and Drosophila. Despite the molecular isolation of LUG and SEU, the biochemical function of these two proteins remains uncharacterized, and the mechanism of AGAMOUS repression remains unknown. Here, we report that LUG and SEU interact directly in vitro and in vivo. Furthermore, LUG exhibits a strong repressor activity on several heterologous promoters in yeast and plant cells. SEU, in contrast, does not exhibit any direct repressor activity, but can repress reporter gene expression only in the presence of LUG, indicating a possible role of SEU as an adaptor protein for LUG. Our results demonstrate that LUG encodes a functional homologue of Tup1 and that SEU may function similarly to Ssn6, an adaptor protein of Tup1. We have defined the LUG/LUH, Flo8, single-strand DNA-binding protein domain of LUG as both necessary and sufficient for the interaction with SEU and two domains of LUG as important for its repressor function. Our work provides functional insights into plant transcriptional corepressors and reveals both conservation and distinctions between plant corepressors and those of yeast and animals.