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vegetative function [keywords]
- Expression and enzymatic properties of rice (Oryza sativa L.) monolignol β-glucosidases. [Journal Article]
- Plant Sci 2014 Oct.:101-9.
Monolignol glucosides and their β-glucosidases are found in monocots, but their biological roles are unclear. Phylogenetic analysis of rice (Oryza sativa L.) glycoside hydrolase family GH1 β-glucosidases indicated that Os4BGlu14, Os4BGlu16, and Os4BGlu18 are closely related to known monolignol β-glucosidases. An optimized Os4BGlu16 cDNA and cloned Os4BGlu18 cDNA were used to express fusion proteins with His6 tags in Pichia pastoris and Escherichia coli, respectively. The secreted Os4BGlu16 fusion protein was purified from media by immobilized metal affinity chromatography (IMAC), while Os4BGlu18 was extracted from E. coli cells and purified by anion exchange chromatography, hydrophobic interaction chromatography and IMAC. Os4BGlu16 and Os4BGlu18 hydrolyzed the monolignol glucosides coniferin (kcat/KM, 21.6mM(-1)s(-1) for Os4BGlu16 and for Os4BGlu18) and syringin (kcat/KM, 22.8mM(-1)s(-1) for Os4BGlu16 and 24.0mM(-1)s(-1) for Os4BGlu18) with much higher catalytic efficiencies than other substrates. In quantitative RT-PCR, highest Os4BGlu14 mRNA levels were detected in endosperm, embryo, lemma, panicle and pollen. Os4BGlu16 was detected highest in leaf from 4 to 10 weeks, endosperm and lemma, while Os4BGlu18 mRNA was most abundant in vegetative stage from 1 week to 4 weeks, pollen and lemma. These data suggest a role for Os4BGlu16 and Os4BGlu18 monolignol β-glucosidases in both vegetative and reproductive rice tissues.
- Arabidopsis RRP6L1 and RRP6L2 Function in FLOWERING LOCUS C Silencing via Regulation of Antisense RNA Synthesis. [Journal Article]
- PLoS Genet 2014 Sep; 10(9):e1004612.
The exosome complex functions in RNA metabolism and transcriptional gene silencing. Here, we report that mutations of two Arabidopsis genes encoding nuclear exosome components AtRRP6L1 and AtRRP6L2, cause de-repression of the main flowering repressor FLOWERING LOCUS C (FLC) and thus delay flowering in early-flowering Arabidopsis ecotypes. AtRRP6L mutations affect the expression of known FLC regulatory antisense (AS) RNAs AS I and II, and cause an increase in Histone3 K4 trimethylation (H3K4me3) at FLC. AtRRP6L1 and AtRRP6L2 function redundantly in regulation of FLC and also act independently of the exosome core complex. Moreover, we discovered a novel, long non-coding, non-polyadenylated antisense transcript (ASL, for Antisense Long) originating from the FLC locus in wild type plants. The AtRRP6L proteins function as the main regulators of ASL synthesis, as these mutants show little or no ASL transcript. Unlike ASI/II, ASL associates with H3K27me3 regions of FLC, suggesting that it could function in the maintenance of H3K27 trimethylation during vegetative growth. AtRRP6L mutations also affect H3K27me3 levels and nucleosome density at the FLC locus. Furthermore, AtRRP6L1 physically associates with the ASL transcript and directly interacts with the FLC locus. We propose that AtRRP6L proteins participate in the maintenance of H3K27me3 at FLC via regulating ASL. Furthermore, AtRRP6Ls might participate in multiple FLC silencing pathways by regulating diverse antisense RNAs derived from the FLC locus.
- Functional Characterisation of Germinant Receptors in Clostridium botulinum and Clostridium sporogenes Presents Novel Insights into Spore Germination Systems. [Journal Article]
- PLoS Pathog 2014 Sep; 10(9):e1004382.
Clostridium botulinum is a dangerous pathogen that forms the highly potent botulinum toxin, which when ingested causes a deadly neuroparalytic disease. The closely related Clostridium sporogenes is occasionally pathogenic, frequently associated with food spoilage and regarded as the non-toxigenic equivalent of Group I C. botulinum. Both species form highly resistant spores that are ubiquitous in the environment and which, under favourable growth conditions germinate to produce vegetative cells. To improve the control of botulinum neurotoxin-forming clostridia, it is imperative to comprehend the mechanisms by which spores germinate. Germination is initiated following the recognition of small molecules (germinants) by a specific germinant receptor (GR) located in the spore inner membrane. The present study precisely defines clostridial GRs, germinants and co-germinants. Group I C. botulinum ATCC3502 contains two tricistronic and one pentacistronic GR operons, while C. sporogenes ATCC15579 has three tricistronic and one tetracistronic GR operons. Insertional knockout mutants, allied with characterisation of recombinant GRs shows for the first time that amino acid stimulated germination in C. botulinum requires two tri-cistronic encoded GRs which act in synergy and cannot function individually. Spore germination in C. sporogenes requires one tri-cistronic GR. Two other GRs form part of a complex involved in controlling the rate of amino-acid stimulated germination. The suitability of using C. sporogenes as a substitute for C. botulinum in germination studies and food challenge tests is discussed.
- The rice receptor-like cytoplasmic kinase GUDK is required for drought tolerance, and grain yield under normal and drought stress conditions. [JOURNAL ARTICLE]
- Plant Physiol 2014 Sep 10.
Rice is the primary food source for more than half of the world's population, and since rice cultivation is dependent on water availability, drought during flowering severely affects grain yield. Here, we show that the function of a drought inducible receptor-like cytoplasmic kinase (RLCK), named GROWTH UNDER DROUGHT KINASE (GUDK), is required for grain yield under drought and well-watered conditions. Loss-of-function gudk mutant lines exhibit sensitivity to salinity, osmotic stress and ABA treatment at the seedling stage, and reduction in photosynthesis and plant biomass under controlled drought stress at the vegetative stage. The gudk mutants interestingly showed significant reduction in grain yield, both under normal well-watered conditions and under drought stress at the reproductive stage. Phosphoproteome profiling of the mutant followed by in vitro assays identified the AP2/ERF transcription factor OsAP37 as a phosphorylation target of GUDK. The involvement of OsAP37 in regulating grain yield under drought through activation of several stress genes has been shown previously (Oh et al., 2009). Our transactivation assays confirmed that GUDK is required for activation of stress genes by OsAP37. We propose that GUDK mediates drought stress signaling through phosphorylation and activation of OsAP37, resulting in transcriptional activation of stress regulated genes, which impart tolerance and improve yield under drought. Our study reveals new insights around drought stress signaling mediated by RLCKs, and also identifies a primary regulator of grain yield in rice that offers the opportunity to improve and stabilize rice grain yield under normal and drought stress conditions.
- Deep brain stimulation of the basolateral amygdala for treatment-refractory combat post-traumatic stress disorder (PTSD): study protocol for a pilot randomized controlled trial with blinded, staggered onset of stimulation. [JOURNAL ARTICLE]
- Trials 2014 Sep 10; 15(1):356.
Combat post-traumatic stress disorder (PTSD) involves significant suffering, impairments in social and occupational functioning, substance use and medical comorbidity, and increased mortality from suicide and other causes. Many veterans continue to suffer despite current treatments. Deep brain stimulation (DBS) has shown promise in refractory movement disorders, depression and obsessive-compulsive disorder, with deep brain targets chosen by integration of clinical and neuroimaging literature. The basolateral amygdala (BLn) is an optimal target for high-frequency DBS in PTSD based on neurocircuitry findings from a variety of perspectives. DBS of the BLn was validated in a rat model of PTSD by our group, and limited data from humans support the potential safety and effectiveness of BLn DBS.We describe the protocol design for a first-ever Phase I pilot study of bilateral BLn high-frequency DBS for six severely ill, functionally impaired combat veterans with PTSD refractory to conventional treatments. After implantation, patients are monitored for a month with stimulators off. An electroencephalographic (EEG) telemetry session will test safety of stimulation before randomization to staggered-onset, double-blind sham versus active stimulation for two months. Thereafter, patients will undergo an open-label stimulation for a total of 24 months. Primary efficacy outcome is a 30% decrease in the Clinician Administered PTSD Scale (CAPS) total score. Safety outcomes include extensive assessments of psychiatric and neurologic symptoms, psychosocial function, amygdala-specific and general neuropsychological functions, and EEG changes. The protocol requires the veteran to have a cohabiting significant other who is willing to assist in monitoring safety and effect on social functioning. At baseline and after approximately one year of stimulation, trauma script-provoked 18FDG PET metabolic changes in limbic circuitry will also be evaluated.While the rationale for studying DBS for PTSD is ethically and scientifically justified, the importance of the amygdaloid complex and its connections for a myriad of emotional, perceptual, behavioral, and vegetative functions requires a complex trial design in terms of outcome measures. Knowledge generated from this pilot trial can be used to design future studies to determine the potential of DBS to benefit both veterans and nonveterans suffering from treatment-refractory PTSD.Trial registration: PCC121657, 19 March 2014.
- Biomechanical adaptation to hindlimb suspension: it involves not only transcriptional mechansims but also postranscriptional modification of the molecular motor, myosin. [EDITORIAL]
- Acta Physiol (Oxf) 2014 Sep 9.
Muscle disuse leads to atrophy, a decrease in force and power output, a slow-to-fast shift of the fiber and myosin heavy chain distribution and an increased shortening velocity, although the latter observations are not equivocal. This editorial comments on the recent paper of Maffei et al. (2014), in which evidence was presented, that the shortening velocity of fast type 2B fibers is in fact reduced and not increase. By investigating the motor function of pure myosin 2B in the in vitro motility assay, they found that actin sliding velocity was decreased and they could ascribed this to an increased phosphorylation of the regulatory light chains of myosin. Thus, the adaptations of muscle function to disuse involves not only transcriptional mechanisms but also posttranscriptional regulation of the motor protein. This article is protected by copyright. All rights reserved.
- Cognitive profiles in patients with multi-infarct dementia: an omani study. [Journal Article]
- Dement Geriatr Cogn Dis Extra 2014 May; 4(2):271-82.
Studies on neurocognitive impairment among patients presenting with multi-infarct dementia (MID) have received little attention from non-Western societies, and the Arab world is no exception. To our knowledge, this is the first study to characterize neurocognitive, affective and vegetative functioning in patients with MID in Oman.In this study, we recruited 20 Omani patients presenting with MID and age- and gender-matched controls at the outpatient clinic of the Department of Behavioral Medicine, Sultan Qaboos University Hospital, Sultan Qaboos University, Muscat, Oman. In addition to the collection of clinical and demographic information, various cognitive batteries were administered to the consenting participants, including those indexing nonverbal reasoning abilities, working memory (attention, concentration and recall) and executive functioning. Questionnaires that elicit the affective range and the quality of sleep were also administered.Compared with the matched healthy subjects, the patients diagnosed with MID significantly differed in the presently operationalized indices of visuospatial function, semantic memory and affective and vegetative functioning. In contrast, episodic memory and some attentional capacities were not significantly different compared with the control subjects.The present study was explorative and clinically designed to describe neurocognitive functioning in patients with MID seeking consultation at a tertiary care center in Oman. Our data are necessary for planning and setting up community services and health care programs for demented patients in a society where dementia is a growing silent epidemic.
- A CRISPR with roles in Myxococcus xanthus development and exopolysaccharide production. [JOURNAL ARTICLE]
- J Bacteriol 2014 Sep 8.
The Gram-negative soil bacterium Myxococcus xanthus utilizes its social (S) gliding motility to move on surfaces during its vegetative and developmental cycles. It is known that S motility requires the type IV pilus (T4P) and the exopolysaccharide (EPS) to function. T4P is the S motility motor and it powers cell movement by retraction. As the key regulator of the S motor, EPS is proposed to be the anchor and trigger for T4P retraction. The production of EPS is regulated in turn by T4P in M. xanthus and T4P(-) mutants are S(-) and EPS(-). In this study, a ΔpilA strain (T4P(-) and EPS(-)) was mutagenized by a transposon and screened for EPS(+) mutants. A pilA suppressor isolated as such harbored an insertion in the 3rd Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR3) in M. xanthus. Evidence indicates that this transposon insertion, designated as CRISPR3*, is a gain-of-function (GOF) mutation. Moreover, CRISPR3* eliminated developmental aggregation in both the wild-type and the pilA mutant backgrounds. Upstream of CRISPR3 are genes coding the Repeat Associated Mysterious Proteins (RAMPs). These RAMP genes are indispensable for CRISPR3* to affect development and EPS in M. xanthus. Analysis by RT-PCR suggested that CRISPR3* led to an increase in the processing of the RNA transcribed from CRISPR3. We propose that certain CRISPR3 transcripts, once expressed and processed, target genes critical for M. xanthus fruiting body development and EPS production in a RAMP-dependent manner.
- Accumulation of astaxanthin by a new Haematococcus pluvialis strain BM1 from the white sea coastal rocks (Russia). [Journal Article]
- Mar Drugs 2014 Aug; 12(8):4504-20.
We report on a novel arctic strain BM1 of a carotenogenic chlorophyte from a coastal habitat with harsh environmental conditions (wide variations in solar irradiance, temperature, salinity and nutrient availability) identified as Haematococcus pluvialis Flotow. Increased (25‰) salinity exerted no adverse effect on the growth of the green BM1 cells. Under stressful conditions (high light, nitrogen and phosphorus deprivation), green vegetative cells of H. pluvialis BM1 grown in BG11 medium formed non-motile palmelloid cells and, eventually, hematocysts capable of a massive accumulation of the keto-carotenoid astaxanthin with a high nutraceutical and therapeutic potential. Routinely, astaxanthin was accumulated at the level of 4% of the cell dry weight (DW), reaching, under prolonged stress, 5.5% DW. Astaxanthin was predominantly accumulated in the form of mono- and diesters of fatty acids from C16 and C18 families. The palmelloids and hematocysts were characterized by the formation of red-colored cytoplasmic lipid droplets, increasingly large in size and number. The lipid droplets tended to merge and occupied almost the entire volume of the cell at the advanced stages of stress-induced carotenogenesis. The potential application of the new strain for the production of astaxanthin is discussed in comparison with the H. pluvialis strains currently employed in microalgal biotechnology.
- PRO40 Is a Scaffold Protein of the Cell Wall Integrity Pathway, Linking the MAP Kinase Module to the Upstream Activator Protein Kinase C. [Journal Article]
- PLoS Genet 2014 Sep; 10(9):e1004582.
Mitogen-activated protein kinase (MAPK) pathways are crucial signaling instruments in eukaryotes. Most ascomycetes possess three MAPK modules that are involved in key developmental processes like sexual propagation or pathogenesis. However, the regulation of these modules by adapters or scaffolds is largely unknown. Here, we studied the function of the cell wall integrity (CWI) MAPK module in the model fungus Sordaria macrospora. Using a forward genetic approach, we found that sterile mutant pro30 has a mutated mik1 gene that encodes the MAPK kinase kinase (MAPKKK) of the proposed CWI pathway. We generated single deletion mutants lacking MAPKKK MIK1, MAPK kinase (MAPKK) MEK1, or MAPK MAK1 and found them all to be sterile, cell fusion-deficient and highly impaired in vegetative growth and cell wall stress response. By searching for MEK1 interaction partners via tandem affinity purification and mass spectrometry, we identified previously characterized developmental protein PRO40 as a MEK1 interaction partner. Although fungal PRO40 homologs have been implicated in diverse developmental processes, their molecular function is currently unknown. Extensive affinity purification, mass spectrometry, and yeast two-hybrid experiments showed that PRO40 is able to bind MIK1, MEK1, and the upstream activator protein kinase C (PKC1). We further found that the PRO40 N-terminal disordered region and the central region encompassing a WW interaction domain are sufficient to govern interaction with MEK1. Most importantly, time- and stress-dependent phosphorylation studies showed that PRO40 is required for MAK1 activity. The sum of our results implies that PRO40 is a scaffold protein for the CWI pathway, linking the MAPK module to the upstream activator PKC1. Our data provide important insights into the mechanistic role of a protein that has been implicated in sexual and asexual development, cell fusion, symbiosis, and pathogenicity in different fungal systems.