Mitogen-activated protein kinase (MAPK) cascades play important roles in regulating plant innate immune responses. In a genetic screen to search for mutants with constitutive defense responses, we identified multiple alleles of mpk4 and mekk1 that exhibit cell death and constitutive defense responses. Bimolecular fluorescence complementation (BiFC) analysis showed that both MPK4 and MEKK1 interact with MKK1 and MKK2, two closely related MAPK kinases. mkk1 and mkk2 single mutant plants do not have obvious mutant phenotypes. To test whether MKK1 and MKK2 function redundantly, mkk1 mkk2 double mutants were generated. The mkk1 mkk2 double mutant plants die at seedling stage and the seedling-lethality phenotype is temperature-dependent. Similar to the mpk4 and mekk1 mutants, the mkk1 mkk2 double mutant seedlings accumulate high levels of H2O2, display spontaneous cell death, constitutively express Pathogenesis Related (PR) genes and exhibit pathogen resistance. In addition, activation of MPK4 by flg22 is impaired in the mkk1 mkk2 double mutants, suggesting that MKK1 and MKK2 function together with MPK4 and MEKK1 in a MAP kinase cascade to negatively regulate innate immune responses in plants.

/npsi/ctrl?lang=en http://nparc.cisti-icist.nrc-cnrc.gc.ca/npsi/ctrl?lang=fr Access and use of this website and the material on it are subject to the Terms and Conditions set forth at http://nparc.cisti-icist.nrc-cnrc.gc.ca/npsi/jsp/nparc_cp.jsp?lang=en NRC Publications Archive Archives des publications du CNRCThis publication could be one of several versions: author's original, accepted manuscript or the publisher's version. / La version de cette publication peut être l'une des suivantes : la version prépublication de l'auteur, la version acceptée du manuscrit ou la version de l'éditeur. For the publisher's version, please access the DOI link below./ Pour consulter la version de l'éditeur, utilisez le lien DOI ci-dessous.http://dx.doi.org/10.1094/MPMI-09-11-0256Molecular Plant-Microbe Interactions, 25, 11, pp. 1459-1468, 2012 Arabidopsis Clade I TGA transcription factors regulate plant defenses in an NPR1-independent fashion Shearer, Heather L.; Cheng, Yu Ti; Wang, Lipu; Liu, Jinman; Boyle, Patrick; Després, Charles; Zhang, Yuelin; Li, Xin; Fobert, Pierre R.

The oral biofilm community consists of >800 microbial species, among which Streptococcus mutans is considered a primary pathogen for dental caries. The genomic island TnSmu2 of S. mutans comprises >2% of the genome. In this study, we demonstrate that TnSmu2 harbors a gene cluster encoding nonribosomal peptide synthetases (NRPS), polyketide synthases (PKS), and accessory proteins and regulators involved in nonribosomal peptide (NRP) and polyketide (PK) biosynthesis. Interestingly, the sequences of these genes and their genomic organizations and locations are highly divergent among different S. mutans strains, yet each TnSmu2 region encodes NRPS/PKS and accessory proteins. Mutagenesis of the structural genes and putative regulatory genes in strains UA159, UA140, and MT4653 resulted in colonies that were devoid of their yellow pigmentation (for strains UA140 and MT4653). In addition, these mutant strains also displayed retarded growth under aerobic conditions and in the presence of H 2 O 2 . High-performance liquid chromatography profiling of cell surface extracts identified unique peaks that were missing in the mutant strains, and partial characterization of the purified product from UA159 demonstrated that it is indeed a hybrid NRP/PK, as predicted. A genomic survey of 94 clinical S. mutans isolates suggests that the TnSmu2 gene cluster may be more prevalent than previously recognized.

The recent investigation of a gene cluster encoding for a hybrid PKS-NRPS metabolite in the oral pathogen Streptococcus mutans UA159 yielded evidence that this natural product might play an important role regulating a range of stress tolerance factors. We have now characterized the major compound generated from this gene cluster, mutanobactin A, and demonstrated that this secondary metabolite is also capable of influencing the yeast-mycelium transition of Candida albicans.A complex suite of cross-species interactions are anticipated to occur among the wide range of microorganisms that constitute the human microbiome. 1-3 While a variety of compounds such as peptides, lipids, and acyl-homoserine lactones have emerged as key players in the multifarious exchanges among microbes and their hosts, 4,5 the important contributions of many other families of secondary metabolites have been largely overlooked. We reported that the deletion of a gene cluster encoding for a putative hybrid polyketide synthasenonribosomal peptide synthetase (PKS-NRPS) derived metabolite in the human oral pathogenic bacteria Streptococcus mutans UA159 resulted in a loss of several resistance traits associated with oxygen and hydrogen peroxide tolerance, as well as biofilm formation. 6 Given that a variety of secondary metabolites are excreted into the extracellular environment, we suspected that the biosynthetic products of this gene cluster (which we have dubbed the mutanobactins 6 ) may have additional yet undefined functions related to the interaction of S. mutans with other members of the oral microbiome. This hypothesis recently gained support based on co-culture studies in which mutanobactin-competent S. mutans and a mutanobactin deletion mutant strain (Δmub) were grown in the presence of Candida albicans. Whereas the mutanobactin producing strain of S. mutans was capable of maintaining C. albicans in a perpetual yeast morphological state (Fig. 1A), deletion of the mutanobactin cluster permitted C. albicans to shift into a mycelial growth pattern, which is believed to be the invasive form of the fungus (Fig. 1B). We now describe the distinctive † Electronic supplementary information (ESI) available: General methods, physical data, MS and NMR ( 1 H, 13 C, 1 H-1 H COSY, 1 H-1 H TOCSY, 1 H-1 H NOESY, 1 H-13 C HSQC, 1 H-13 C HMBC, and 1 H-15 N HMBC) for 1. Analytical-scale HPLC comparison of the crude extracts generated from wild-type S. mutans UA159 and Δmub strains grown on brain-heart-infusion agar plates enabled us to identify two metabolites in the elution profile that were present only in the wild-type organism. Scale-up fermentation of the wild-type strain was performed in a bioreactor with 15 L of brain-heart-infusion broth under microaerobic conditions at 37 °C to enable the isolation and structure characterization of the major metabolite. After 48 h, the cells and broth were partitioned against ethyl acetate and the solvent removed in vacuo. The resulting extract was resuspended in methanol and the organic soluble material ...

In the oral biofilm, the 'mitis' streptococci are among the first group of organisms to colonize the tooth surface. Their proliferation is thought to be an important factor required for antagonizing the growth of cariogenic species such as Streptococcus mutans. In this study, we used a threespecies mixed culture to demonstrate that another ubiquitous early colonizing species, Veillonella parvula, can greatly affect the outcome of the competition between a pair of antagonists such as S. mutans and Streptococcus gordonii. Transcriptome analysis further revealed that S. mutans responds differentially to its friend (V. parvula) and foe (S. gordonii). In the mixed culture with S. gordonii, all but one of the S. mutans sugar uptake and metabolic genes were downregulated, while genes for alternative energy source utilization and H 2 O 2 tolerance were upregulated, resulting in a slower but persistent growth. In contrast, when cultured with V. parvula, S. mutans grew equally well or better than in monoculture and exhibited relatively few changes within its transcriptome. When V. parvula was introduced into the mixed culture of S. mutans and S. gordonii, it rescued the growth inhibition of S. mutans. In this three-species environment, S. mutans increased the expression of genes required for the uptake and metabolism of minor sugars, while genes required for oxidative stress tolerance were downregulated. We conclude that the major factors that affect the competition between S. mutans and S. gordonii are carbohydrate utilization and H 2 O 2 resistance. The presence of V. parvula in the tri-species culture mitigates these two major factors and allows S. mutans to proliferate, despite the presence of S. gordonii. INTRODUCTIONBiofilms in the human oral cavity are exceptionally diverse and can harbour more than 800 microbial species (Aas et al., 2005;Keijser et al., 2008;Paster et al., 2001Paster et al., , 2006Zaura et al., 2009). Dental biofilm formation is hypothesized to be a sequential process Kolenbrander et al., 2006Kolenbrander et al., , 2010. After a new tooth emerges or a surface is cleaned, the surface is colonized by a group of bacteria named the 'pioneer colonizers', which are composed mostly of the 'mitis' streptococci (i.e. Streptococcus gordonii, Streptococcus sanguinis, Streptococcus mitis, etc.). Subsequently, early colonizers such as Streptococcus mutans and veillonellae, and bridging species such as the fusobacteria, join the community through interactions with the pioneer colonizers or by adhering to the available sites on the tooth surface. Growth of the early colonizers then modifies the local environment, making it favourable for the growth of late colonizers, which consist mostly of Gram-negative, obligate anaerobic bacteria. Eventually, through cell growth and co-adhesion, a mature biofilm is formed (for reviews, see Diaz et al., 2006;Kolenbrander et al., 2010). Once formed, the biofilm composition is relatively stable, featuring high biodiversity; however, environmental perturbation can disrupt this e...