HydroxyprolineO-arabinosyltransferase mutants oppositely alter tip growth inArabidopsis thalianaandPhyscomitrella patens

Hydroxyproline O-arabinosyltransferases (HPATs) are members of a small, deeply conserved family of plant-specific glycosyltransferases that add arabinose sugars to diverse proteins including cell wall-associated extensins and small signaling peptides. Recent genetic studies in flowering plants suggest that different HPAT homologs have been co-opted to function in diverse species-specific developmental contexts. However, nothing is known about the roles of HPATs in basal plants. We show that complete loss of HPAT function in Arabidopsis thaliana and the moss Physcomitrella patens results in a shared defect in gametophytic tip cell growth. Arabidopsis hpat1/2/3 triple knockout mutants suffer from a strong male sterility defect as a consequence of pollen tubes that fail to fully elongate following pollination. Knocking out the two HPAT genes of Physcomitrella results in larger multicellular filamentous networks due to increased elongation of protonemal tip cells. Physcomitrella hpat mutants lack cell-wall associated hydroxyproline arabinosides and can be rescued with exogenous cellulose, while global expression profiling shows that cell wall-associated genes are severely misexpressed, implicating a defect in cell wall formation during tip growth. Our findings point to a major role for HPATs in influencing cell elongation during tip growth in plants.

The Sinorhizobium meliloti EmrR Regulator Is Required for Efficient Colonization of Medicago sativa Root Nodules

The nitrogen-fixing bacterium Sinorhizobium meliloti must adapt to diverse conditions encountered during its symbiosis with leguminous plants. We characterized a new symbiotically relevant gene, emrR (SMc03169), whose product belongs to the TetR family of repressors and is divergently transcribed from emrAB genes encoding a putative major facilitator superfamily-type efflux pump. An emrR deletion mutant produced more succinoglycan, displayed increased cell-wall permeability, and exhibited higher tolerance to heat shock. It also showed lower tolerance to acidic conditions, a reduced production of siderophores, and lower motility and biofilm formation. The simultaneous deletion of emrA and emrR genes restored the mentioned traits to the wild-type phenotype, except for survival under heat shock, which was lower than that displayed by the wild-type strain. Furthermore, the ΔemrR mutant as well as the double ΔemrAR mutant was impaired in symbiosis with Medicago sativa; it formed fewer nodules and competed poorly with the wild-type strain for nodule colonization. Expression profiling of the ΔemrR mutant showed decreased expression of genes involved in Nod-factor and rhizobactin biosynthesis and in stress responses. Expression of genes directing the biosynthesis of succinoglycan and other polysaccharides were increased. EmrR may therefore be involved in a regulatory network targeting membrane and cell wall modifications in preparation for colonization of root hairs during symbiosis.

The Trk Potassium Transporter Is Required for RsmB-Mediated Activation of Virulence in the Phytopathogen Pectobacterium wasabiae

Pectobacterium wasabiae (previously known as Erwinia carotovora) is an important plant pathogen that regulates the production of plant cell wall-degrading enzymes through an N-acyl homoserine lactone-based quorum sensing system and through the GacS/GacA two-component system (also known as ExpS/ExpA). At high cell density, activation of GacS/GacA induces the expression of RsmB, a noncoding RNA that is essential for the activation of virulence in this bacterium. A genetic screen to identify regulators of RsmB revealed that mutants defective in components of a putative Trk potassium transporter (trkH and trkA) had decreased rsmB expression. Further analysis of these mutants showed that changes in potassium concentration influenced rsmB expression and consequent tissue damage in potato tubers and that this regulation required an intact Trk system. Regulation of rsmB expression by potassium via the Trk system occurred even in the absence of the GacS/GacA system, demonstrating that these systems act independently and are both required for full activation of RsmB and for the downstream induction of virulence in potato infection assays. Overall, our results identified potassium as an essential environmental factor regulating the Rsm system, and the consequent induction of virulence, in the plant pathogen P. wasabiae.

Expression-based and co-localization detection of arabinogalactan protein 6 and arabinogalactan protein 11 interactors in Arabidopsis pollen and pollen tubes

Arabinogalactan proteins (AGPs) are cell wall proteoglycans that have been shown to be important for pollen development. An Arabidopsis double null mutant for two pollen-specific AGPs (agp6 agp11) showed reduced pollen tube growth and compromised response to germination cues in vivo. A microarray experiment was performed on agp6 agp11 pollen tubes to search for genetic interactions in the context of pollen tube growth. A yeast two-hybrid experiment for AGP6 and AGP11 was also designed.

Whole Genome Analysis of Gene Expression Reveals Coordinated Activation of Signaling and Metabolic Pathways during Pollen-Pistil Interactions in Arabidopsis

Plant reproduction depends on the concerted activation of many genes to ensure correct communication between pollen and pistil. Here, we queried the whole transcriptome of Arabidopsis (Arabidopsis thaliana) in order to identify genes with specific reproductive functions. We used the Affymetrix ATH1 whole genome array to profile wild-type unpollinated pistils and unfertilized ovules. By comparing the expression profile of pistils at 0.5, 3.5, and 8.0 h after pollination and applying a number of statistical and bioinformatics criteria, we found 1,373 genes differentially regulated during pollen-pistil interactions. Robust clustering analysis grouped these genes in 16 time-course clusters representing distinct patterns of regulation. Coregulation within each cluster suggests the presence of distinct genetic pathways, which might be under the control of specific transcriptional regulators. A total of 78% of the regulated genes were expressed initially in unpollinated pistil and/or ovules, 15% were initially detected in the pollen data sets as enriched or preferentially expressed, and 7% were induced upon pollination. Among those, we found a particular enrichment for unknown transcripts predicted to encode secreted proteins or representing signaling and cell wall-related proteins, which may function by remodeling the extracellular matrix or as extracellular signaling molecules. A strict regulatory control in various metabolic pathways suggests that fine-tuning of the biochemical and physiological cellular environment is crucial for reproductive success. Our study provides a unique and detailed temporal and spatial gene expression profile of in vivo pollen-pistil interactions, providing a framework to better understand the basis of the molecular mechanisms operating during the reproductive process in higher plants.