Linnemannia elongata (Mortierellaceae) stimulates Arabidopsis thaliana aerial growth and responses to auxin, ethylene, and reactive oxygen species

Harnessing the plant microbiome has the potential to improve agricultural yields and protect plants against pathogens and/or abiotic stresses, while also relieving economic and environmental costs of crop production. While previous studies have gained valuable insights into the underlying genetics facilitating plant-fungal interactions, these have largely been skewed towards certain fungal clades (e.g. arbuscular mycorrhizal fungi). Several different phyla of fungi have been shown to positively impact plant growth rates, including Mortierellaceae fungi. However, the extent of the plant growth promotion (PGP) phenotype(s), their underlying mechanism(s), and the impact of bacterial endosymbionts on fungal-plant interactions remain poorly understood for Mortierellaceae. In this study, we focused on the symbiosis between soil fungus Linnemannia elongata (Mortierellaceae) and Arabidopsis thaliana (Brassicaceae), as both organisms have high-quality reference genomes and transcriptomes available, and their lifestyles and growth requirements are conducive to research conditions. Further, L. elongata can host bacterial endosymbionts related to Mollicutes and Burkholderia . The role of these endobacteria on facilitating fungal-plant associations, including potentially further promoting plant growth, remains completely unexplored. We measured Arabidopsis aerial growth at early and late life stages, seed production, and used mRNA sequencing to characterize differentially expressed plant genes in response to fungal inoculation with and without bacterial endosymbionts. We found that L. elongata improved aerial plant growth, seed mass and altered the plant transcriptome, including the upregulation of genes involved in plant hormones and “response to oxidative stress”, “defense response to bacterium”, and “defense response to fungus”. Furthermore, the expression of genes in certain phytohormone biosynthetic pathways were found to be modified in plants treated with L. elongata . Notably, the presence of Mollicutes- or Burkholderia- related endosymbionts in Linnemannia did not impact the expression of genes in Arabidopsis or overall growth rates.

FgHtf1 Regulates Global Gene Expression towards Aerial Mycelium and Conidiophore Formation in the Cereal Fungal Pathogen Fusarium graminearum

ABSTRACT The homeobox gene family of transcription factors (HTF) controls many developmental pathways and physiological processes in eukaryotes. We previously showed that a conserved HTF in the plant-pathogenic fungus Fusarium graminearum, Htf1 (FgHtf1), regulates conidium morphology in that organism. This study investigated the mechanism of FgHtf1-mediated regulation and identified putative FgHtf1 target genes by a chromatin immunoprecipitation assay combined with parallel DNA sequencing (ChIP-seq) and RNA sequencing. A total of 186 potential binding peaks, including 142 genes directly regulated by FgHtf1, were identified. Subsequent motif prediction analysis identified two DNA-binding motifs, TAAT and CTTGT. Among the FgHtf1 target genes were FgHTF1 itself and several important conidiation-related genes (e.g., FgCON7), the chitin synthase pathway genes, and the aurofusarin biosynthetic pathway genes. In addition, FgHtf1 may regulate the cAMP-protein kinase A (PKA)-Msn2/4 and Ca2+-calcineurin-Crz1 pathways. Taken together, these results suggest that, in addition to autoregulation, FgHtf1 also controls global gene expression and promotes a shift to aerial growth and conidiation in F. graminearum by activation of FgCON7 or other conidiation-related genes. IMPORTANCE The homeobox gene family of transcription factors is known to be involved in the development and conidiation of filamentous fungi. However, the regulatory mechanisms and downstream targets of homeobox genes remain unclear. FgHtf1 is a homeobox transcription factor that is required for phialide development and conidiogenesis in the plant pathogen F. graminearum. In this study, we identified FgHtf1-controlled target genes and binding motifs. We found that, besides autoregulation, FgHtf1 also controls global gene expression and promotes conidiation in F. graminearum by activation of genes necessary for aerial growth, FgCON7, and other conidiation-related genes.

Changes on Potato Leaf Metabolism and Anatomy Induced by Plant Growth Regulators

The use of growth regulators in potato crop is an alternative to reduce the aerial growth of plants and redirects carbon assimilates and nutrients to the tubers. Therefore, the objective of this study was to evaluate the effects of growth regulators, paclobutrazol and trinexapac-ethyl on plant growth and changes on the anatomy of leaves of cultivar Markies in summer conditions of the southern region of Brazil. Potato plants cv. Markies were in the summer growing season of Southeast region of Brazil and 35 days after planting, the plants were sprayed with paclobutrazol (PBZ) at 0.125 and 0.250 L ha-1 and trinexapac-ethyl (TE) at 1.0 and 2.0 L ha-1. Treatment with PBZ at both doses reduced the height of potato plants, which resulted in higher index of leaf chlorophyll and reduced the content of starch and non-reducing sugars. Both PBZ and TE treated plants exhibited anatomical changes in the leaves, including larger epidermal cells and more elongated palisades cells. These data suggest that such changes in the anatomy of potato leaf in response to the use of PBZ directly influence leaf metabolism.

The Beneficial Root-Colonizing Fungus Mortierella hyalina Promotes the Aerial Growth of Arabidopsis and Activates Calcium-Dependent Responses That Restrict Alternaria brassicae–Induced Disease Development in Roots

The endophytic fungus Mortierella hyalina colonizes the roots of Arabidopsis thaliana and stimulates growth and biomass production of the aerial parts but not of roots. An exudate fraction from the fungus induces rapid and transient cytoplasmic Ca2+elevation in the roots. The Ca2+ response does not require the well-characterized (co)receptors BAK1, CERK1, and FLS2 for pathogen-associated molecular patterns, and the Ca2+ channels GLR-2.4, GLR-2.5, and GLR-3.3 or the vacuolar TWO PORE CHANNEL1, which might be involved in cytoplasmic Ca2+ elevation. We isolated an ethyl-methane-sulfonate–induced Arabidopsis mutant that is impaired in this Ca2+ response. The roots of the mutant are impaired in M. hyalina–mediated suppression of immune responses after Alternaria brassicae infection, i.e., jasmonate accumulation, generation of reactive oxygen species, as well as the activation of jasmonate-related defense genes. Furthermore, they are more colonized by M. hyalina than wild-type roots. We propose that the mutant gene product is involved in a Ca2+-dependent signaling pathway activated by M. hyalina to suppress immune responses in Arabidopsis roots.

Growth dynamics of the branching system in grasses: an architecture-based methodology for quantitative evaluation applied to two genotypes of Paspalum vaginatum

The aerial growth dynamics of the shoots of the widespread turfgrass Paspalum vaginatum was studied. With the aim of identifying quantitative differential features between two genotypes, plants were cultivated outdoors in pots during two consecutive summers. Axes of different branching order were marked for periodical observation to quantify internode production rate, axis production rate, covering rate and reproductive effort. The genotypes differed significantly in the following quantitative features: plagiotropic vs. orthotropic axes proportion, internode production rate, flowering vs. vegetative tiller ratio, vegetative cover and spatial distribution of the canopy. Knowledge of these features may help to understand the great plasticity and adaptability of this species to multiple environmental conditions.

The DyP-type peroxidase DtpA is a Tat-substrate required for GlxA maturation and morphogenesis in Streptomyces

The filamentous bacterium Streptomyces lividans depends on the radical copper oxidase GlxA for the formation of reproductive aerial structures and, in liquid environments, for the formation of pellets. Incorporation of copper into the active site is essential for the formation of a cross-linked tyrosyl-cysteine cofactor, which is needed for enzymatic activity. In this study, we show a crucial link between GlxA maturation and a group of copper-related proteins including the chaperone Sco and a novel DyP-type peroxidase hereinafter called DtpA. Under copper-limiting conditions, the sco and dtpA deletion mutants are blocked in aerial growth and pellet formation, similarly to a glxA mutant. Western blot analysis showed that GlxA maturation is perturbed in the sco and dtpA mutants, but both maturation and morphology can by rescued by increasing the bioavailability of copper. DtpA acts as a peroxidase in the presence of GlxA and is a substrate for the twin-arginine translocation (Tat) translocation pathway. In agreement, the maturation status of GlxA is also perturbed in tat mutants, which can be compensated for by the addition of copper, thereby partially restoring their morphological defects. Our data support a model wherein a copper-trafficking pathway and Tat-dependent secretion of DtpA link to the GlxA-dependent morphogenesis pathway.

Genes Required for Aerial Growth, Cell Division, and Chromosome Segregation Are Targets of WhiA before Sporulation in Streptomyces venezuelae

ABSTRACTWhiA is a highly unusual transcriptional regulator related to a family of eukaryotic homing endonucleases. WhiA is required for sporulation in the filamentous bacteriumStreptomyces, but WhiA homologues of unknown function are also found throughout the Gram-positive bacteria. To better understand the role of WhiA inStreptomycesdevelopment and its function as a transcription factor, we identified the WhiA regulon through a combination of chromatin immunoprecipitation-sequencing (ChIP-seq) and microarray transcriptional profiling, exploiting a new model organism for the genus,Streptomyces venezuelae, which sporulates in liquid culture. The regulon encompasses ~240 transcription units, and WhiA appears to function almost equally as an activator and as a repressor. Bioinformatic analysis of the upstream regions of the complete regulon, combined with DNase I footprinting, identified a short but highly conserved asymmetric sequence, GACAC, associated with the majority of WhiA targets. Construction of a null mutant showed thatwhiAis required for the initiation of sporulation septation and chromosome segregation inS. venezuelae, and several genes encoding key proteins of theStreptomycescell division machinery, such asftsZ,ftsW, andftsK, were found to be directly activated by WhiA during development. Several other genes encoding proteins with important roles in development were also identified as WhiA targets, including the sporulation-specific sigma factor σWhiGand the diguanylate cyclase CdgB. Cell division is tightly coordinated with the orderly arrest of apical growth in the sporogenic cell, andfilP, encoding a key component of the polarisome that directs apical growth, is a direct target for WhiA-mediated repression during sporulation.IMPORTANCESince the initial identification of the genetic loci required forStreptomycesdevelopment, all of thebldandwhidevelopmental master regulators have been cloned and characterized, and significant progress has been made toward understanding the cell biological processes that drive morphogenesis. A major challenge now is to connect the cell biological processes and the developmental master regulators by dissecting the regulatory networks that link the two. Studies of these regulatory networks have been greatly facilitated by the recent introduction ofStreptomyces venezuelaeas a new model system for the genus, a species that sporulates in liquid culture. Taking advantage ofS. venezuelae, we have characterized the regulon of genes directly under the control of one of these master regulators, WhiA. Our results implicate WhiA in the direct regulation of key steps in sporulation, including the cessation of aerial growth, the initiation of cell division, and chromosome segregation.

Hopanoids Are Not Essential for Growth of Streptomyces scabies 87-22

ABSTRACT Hopanoids are triterpenoic, pentacyclic compounds that are structurally similar to sterols, which are required for normal cell function in eukaryotes. Hopanoids are thought to be an important component of bacterial cell membranes because they control membrane fluidity and diminish passive diffusion of ions, and a few taxons modulate their hopanoid content in response to environmental stimuli. However, to our knowledge, mutational studies to assess the importance of hopanoids in bacterial physiology have never been performed. Genome sequencing of the potato scab pathogen, Streptomyces scabies 87-22, revealed a hopanoid biosynthetic gene cluster (HBGC) that is predicted to synthesize hopene and aminotrihydroxybacteriohopane products. Hopene was produced by fully sporulated cultures of S. scabies on solid ISP4 (International Streptomyces Project 4) medium as well as by submerged mycelia grown in liquid minimal medium. The elongated hopanoid aminotrihydroxybacteriohopane was not detected under either growth condition. Transcription of the S. scabies HBGC was upregulated during aerial growth, which suggests a link between hopanoid production and morphological development. Functional analysis of the S. scabies Δhop615-1 and Δhop615-7 mutant strains, the first hopanoid mutants created in any bacterial taxon, revealed that hopanoids are not required for normal growth or for tolerance of ethanol, osmotic and oxidative stress, high temperature, or low pH. This suggests that hopanoids are not essential for normal streptomycete physiology.