Salicylic acid (SA) is a plant immune signal produced upon pathogen challenge to induce systemic acquired resistance (SAR). It is the only major plant hormone for which the receptor has not been firmly identified. SAR in Arabidopsis requires the transcription cofactor NPR1 (nonexpresser of PR genes 1), whose degradation serves as a molecular switch for SAR. Here we show that NPR1 paralogues, NPR3 and NPR4, are SA receptors that bind SA with different affinities and function as adaptors of the Cullin 3 ubiquitin E3 ligase to mediate NPR1 degradation in an SA-regulated manner. Accordingly, the npr3 npr4 mutant accumulates higher levels of NPR1 and is insensitive to SAR induction. Moreover, this mutant is defective in pathogen effector-triggered programmed cell death and immunity. Our study reveals the mechanism of SA perception in determining cell death and survival in response to pathogen challenge.

Congenital heart disease (CHD) is the most frequent birth defect, affecting 0.8% of live births1. Many cases occur sporadically and impair reproductive fitness, suggesting a role for de novo mutations. By analysis of exome sequencing of parent-offspring trios, we compared the incidence of de novo mutations in 362 severe CHD cases and 264 controls. CHD cases showed a significant excess of protein-altering de novo mutations in genes expressed in the developing heart, with an odds ratio of 7.5 for damaging mutations. Similar odds ratios were seen across major classes of severe CHD. We found a marked excess of de novo mutations in genes involved in production, removal or reading of H3K4 methylation (H3K4me), or ubiquitination of H2BK120, which is required for H3K4 methylation2–4. There were also two de novo mutations in SMAD2; SMAD2 signaling in the embryonic left-right organizer induces demethylation of H3K27me5. H3K4me and H3K27me mark `poised' promoters and enhancers that regulate expression of key developmental genes6. These findings implicate de novo point mutations in several hundred genes that collectively contribute to ~10% of severe CHD.

The principal immune mechanism against biotrophic pathogens in plants is the resistance (R)-gene-mediated defence. It was proposed to share components with the broad-spectrum basal defence machinery. However, the underlying molecular mechanism is largely unknown. Here we report the identification of novel genes involved in R-gene-mediated resistance against downy mildew in Arabidopsis and their regulatory control by the circadian regulator, CIRCADIAN CLOCK-ASSOCIATED 1 (CCA1). Numerical clustering based on phenotypes of these gene mutants revealed that programmed cell death (PCD) is the major contributor to resistance. Mutants compromised in the R-gene-mediated PCD were also defective in basal resistance, establishing an interconnection between these two distinct defence mechanisms. Surprisingly, we found that these new defence genes are under circadian control by CCA1, allowing plants to 'anticipate' infection at dawn when the pathogen normally disperses the spores and time immune responses according to the perception of different pathogenic signals upon infection. Temporal control of the defence genes by CCA1 differentiates their involvement in basal and R-gene-mediated defence. Our study has revealed a key functional link between the circadian clock and plant immunity.

Transposon systems are widely used for generating mutations in various model organisms. PiggyBac (PB) has recently been shown to transpose efficiently in the mouse germ line and other mammalian cell lines. To facilitate PB's application in mammalian genetics, we characterized the properties of the PB transposon in mouse embryonic stem (ES) cells. We first measured the transposition efficiencies of PB transposon in mouse embryonic stem cells. We next constructed a PB/SB hybrid transposon to compare PB and Sleeping Beauty (SB) transposon systems and demonstrated that PB transposition was inhibited by DNA methylation. The excision and reintegration rates of a single PB from two independent genomic loci were measured and its ability to mutate genes with gene trap cassettes was tested. We examined PB's integration site distribution in the mouse genome and found that PB transposition exhibited local hopping. The comprehensive information from this study should facilitate further exploration of the potential of PB and SB DNA transposons in mammalian genetics. ES cell ͉ Sleeping Beauty ͉ transposon DNA transposons are genetic elements that can relocate between genomic sites by a ''cut and paste'' mechanism. Since the discovery of the first DNA transposon in maize by Barbara McClintock (1), these elements have been extensively used for genetics and functional genomics in different organisms (2-7).Sleeping Beauty was the first DNA transposon system shown to be functional in mammalian cells. It has been tested for insertional mutagenesis in the mouse and rat germ lines (8-13), but its relatively low transposition efficiency and strong ''local hopping'' tendency limit its application in genome-wide screens. However, SB has successfully been used to screen for new cancer genes (14,15).PiggyBac was isolated from Trichoplusia ni (16) and subsequently found to transpose efficiently in many different species (5,(17)(18)(19). One important feature of the PB transposon is that it nearly always excises itself precisely and leaves no footprint behind (5,20,21). It was demonstrated that PB was very efficient for germ-line mutagenesis in the mouse (22, 23). It was subsequently confirmed that PB has significantly higher transposition activity in mammalian cell lines than SB and Tol2 (24). These studies suggest that PB has wide applications in dissecting gene functions (25).To facilitate PB's application in mammals, we have characterized further the properties of the PB transposon. We measured the transposition efficiencies of PB in mouse ES cells. We also constructed a PB/SB hybrid transposon to compare the PB and SB transposon systems in ES cells. Our data demonstrated that PB transposition was inhibited by DNA methylation. We then measured the excision and reintegration rates of a single PB transposon from two independent genomic loci and tested its ability to mutate a genomic locus with gene trap cassettes. Finally, we investigated PB's integration site distribution in the mouse genome and found that at least 9% of transposition eve...

Summary Background Induction of plant immune responses involves significant transcription reprogramming that prioritizes defense over growth-related cellular functions. Despite intensive forward genetic screens and genome-wide expression-profiling studies, a limited number of transcription factors have been found that regulate this transition. Results Using the endoplasmic-reticulum-resident genes required for antimicrobial protein secretion as markers, we identified a heat-shock factor-like transcription factor that specifically binds to the TL1 (GAAGAAGAA) cis element required for the induction of these genes. Surprisingly, plants lacking this TL1-binding factor, TBF1, respond normally to heat stress but are compromised in immune responses induced by salicylic acid and by microbe-associated molecular pattern, elf18. Genome-wide expression profiling indicates that TBF1 plays a key role in the growth-to-defense transition. Moreover, the expression of TBF1 itself is tightly regulated at both the transcriptional and translational levels. Two upstream open reading frames encoding multiple aromatic amino acids were found 5′ of the translation initiation codon of TBF1 and shown to affect its translation. Conclusions Through this unique regulatory mechanism, TBF1 can sense the metabolic changes upon pathogen invasion and trigger the specific transcriptional reprogramming through its target genes expression.

Dynamic changes in the expression of transcription factors (TFs) can influence specification of distinct CD8+ T cell fates, but the observation of equivalent expression of TF among differentially-fated precursor cells suggests additional underlying mechanisms. Here, we profiled genome-wide histone modifications, open chromatin and gene expression of naive, terminal-effector, memory-precursor and memory CD8+ T cell populations induced during the in vivo response to bacterial infection. Integration of these data suggested that TF expression and binding contributed to establishment of subset-specific enhancers during differentiation. We developed a new bioinformatics method using the PageRank algorithm to reveal novel TFs influencing the generation of effector and memory populations. The TFs YY1 and Nr3c1, both constitutively expressed during CD8+ T cell differentiation, regulated the formation of terminal-effector and memory-precursor cell-fates, respectively. Our data define the epigenetic landscape of differentiation intermediates, facilitating identification of TFs with previously unappreciated roles in CD8+ T cell differentiation.

Enhancers act to regulate cell type specific gene expression by facilitating the transcription of target genes. In mammalian cells active or primed enhancers are commonly marked by monomethylation of Histone H3 at lysine 4 (H3K4me1) in a cell-type specific manner. Whether and how this histone modification regulates enhancer-dependent transcription programs in mammals is unclear. In this study, we conducted SILAC Mass-spec experiments with mono-nucleosomes and identified multiple H3K4me1 associated proteins, including many involved in chromatin remodeling. We demonstrate that H3K4me1 augments the association of the chromatin remodeling complex BAF to enhancers in vivo and that in vitro, H3K4me1 nucleosomes are more efficiently remodeled by the BAF complex. Crystal structures of BAF component BAF45c reveal that monomethylation, but not trimethylation, is accommodated by BAF45c’s H3K4 binding site. Our results suggest that H3K4me1 plays an active role at enhancers by facilitating the binding of the BAF complex and possibly other chromatin regulators.

Although mesenchymal progenitor cells can be isolated from periodontal ligament (PDL) tissues using stem cell markers STRO-1 and CD146, the proportion of these cells that have the capacity to differentiate into multiple cell lineages remains to be determined. This study was designed to quantify the proportions of primary human PDL cells that can undergo multi-lineage differentiation, and to compare the magnitude of these capabilities relative to bone marrow-derived mesenchymal stem cells (MSC) and parental PDL (PPDL) cells. PDL mesenchymal progenitor (PMP) cells were isolated from PPDL cells using the markers STRO-1 and CD146. The colony-forming efficiency and multi-lineage differentiation potential of PMP, PPDL, and MSCs under chondrogenic, osteogenic and adipogenic conditions were determined. Flow cytometry revealed that on average 2.6% of PPDL cells were STRO-1+/CD146+, whereas more than 63% were STRO-1-/CD146-. Colony-forming efficiency of STRO-1+/CD146+ PMP cells (19.3%) and MSCs (16.7%) was significantly higher than that of PPDL cells (6.8%). Cartilage specific genes, early markers of osteoblastic differentiation, and adipogenic markers were significantly up-regulated under appropriate conditions in PMP cells and MSCs compared to either their non-induced counterparts or induced PPDL cells. Consistent with these findings, immunohistochemistry revealed substantial accumulation of cartilaginous macromolecules, mineralized calcium nodules, and lipid vacuoles under chondrogenic, osteogenic or adipogenic conditions in PMP and MSC cultures, respectively, compared to non-induced controls or induced PPDL cells. Thus, STRO-1+/CD146+ PMP cells demonstrate multi-lineage differentiation capacity comparable in magnitude to MSCs, and could potentially be utilized for regeneration of the periodontium and other tissues.