Isolation and analysis of the KATI promoter from \u3ci\u3eArabidopsis thaliana\u3c/i\u3e

Plants that reduce water loss by transpiration present less agricultural stress to the environment. Transpiration is limited by the size of pores, or stomata, on the surfaces of leaves. Stomatal aperture is related to the ion concentration in surrounding guard cells, which varies in response to fluctuating concentrations of potassium ions, the principle counter-ion chloride, and the organic counter-ion malate. Levels of malate in guard cells may be altered by increased activity of NADP-Malic Enzyme (NADP-ME) expression. The goal of our research is to create transgenic Arabidopsis thaliana plants with increased NADP-ME expression in the guard cells, limiting transpiration through decreased stomatal aperture size. Such plants are expected to have less open stomata, be sufficient in yield, and retain the ability to respond to changing environmental conditions. Expression of NADP-ME in Arabidopsis thaliana required the isolation of a strong, guard-cell specific promoter. A promoter of a potassium channel gene, KAT1, is strongly expressed in guard cells. Analysis of the Arabidopsis genome was done to locate KAT1. The KAT1 promoter was amplified and isolated through Polymerase Chain Reaction (PCR). Sequence analysis confirmed isolation of KAT1 with minor mutations. Visualization of KAT1 expression was confirmed through the analysis of transgenic plants with the KAT1 promoter fused to the GUS reporter gene. Significant expression of KAT1 was detected exclusively in guard cells of 9-day-old seedlings. The KAT1 promoter will be inserted into a binary vector and then Agrobacterium to transform Arabidopsis thaliana and create transgenic plants. Expression of KAT1 with NADP-ME in Arabidopsis thaliana should result in lower concentrations of malate in guard cells, decreased aperture size of stomata, and a decrease in transpiration rate during gas exchange

Negative feedback control of jasmonate signaling by an alternative splice variant of JAZ10

The plant hormone jasmonate (JA) activates gene expression by promoting ubiquitin-dependent degradation of JAZ transcriptional repressor proteins. A key feature of all JAZ proteins is the highly conserved Jas motif, which mediates both JAZ degradation and JAZ binding to the transcription factor MYC2. Rapid expression of JAZ genes in response to JA is thought to attenuate JA responses, but little is known about the mechanisms by which newly synthesized JAZ proteins exert repression in the presence of the hormone. Here, we show that desensitization to JA is mediated by an alternative splice variant (JAZ10.4) of JAZ10 that lacks the Jas motif. Unbiased protein-protein interaction screens identified three related bHLH transcription factors (MYC2, MYC3, and MYC4) and the co-repressor NINJA as JAZ10.4-binding partners. We show that the N-terminal region of JAZ10.4 contains a cryptic MYC2-binding site that resembles the Jas motif, and that the ZIM motif of JAZ10.4 functions as a transferable repressor domain whose activity is associated with recruitment of NINJA. Functional studies showed that expression of JAZ10.4 from the native JAZ10 promoter complemented the JA-hypersensitive phenotype of a jaz10 mutant. Moreover, treatment of these complemented lines with JA resulted in rapid accumulation of JAZ10.4 protein. Our results provide an explanation for how the unique domain architecture of JAZ10.4 links transcription factors to a co-repressor complex, and suggest how JA-induced transcription and alternative splicing of JAZ10 pre-mRNA creates a regulatory circuit to attenuate JA responses.Fil: Moreno, Javier Edgardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentina. Michigan State University; Estados UnidosFil: Shyu, Christine. Michigan State University; Estados UnidosFil: Campos, Marcelo L.. Michigan State University; Estados UnidosFil: Patel, Lalita C.. Michigan State University; Estados UnidosFil: Chung, Hoo Sun. Michigan State University; Estados UnidosFil: Yao, Jian. Michigan State University; Estados UnidosFil: He, Sheng Hang. Michigan State University; Estados UnidosFil: Howe, Gregg A.. Michigan State University; Estados Unido

GeneXpert MTB/Rif to Diagnose Tuberculous Meningitis: Perhaps the First Test but not the Last

Tuberculous meningitis (TBM) is the most severe form of tuberculous with substantial mortality. In May 2015, 54 researchers from 10 countries met in Da Lat, Vietnam, to discuss advances in TBM. Among the attendees were researchers involved in pivotal studies on the use of Xpert MTB/Rif for TBM diagnosis. Attendees discussed the 2014 World Health Organization strong recommendation favoring the use of Xpert "in preference to conventional microscopy and culture as the initial diagnostic test for cerebrospinal fluid (CSF) if the sample volume is low or if additional specimens cannot be obtained to make a quick diagnosis." Attendees were concerned that the limitations of Xpert testing for TBM are not emphasized. Clear guidance is needed for the investigational pathway for TBM, including recommendations on the diagnostic package of investigations, which does not stop with Xpert testing. Second, emphasis on the large CSF volumes (ideally 8-10 mL) needed for Xpert testing is required. Guidelines should also emphasize that TBM is a medical emergency and early treatment reduces mortality