Microscopic pores present in the epidermis of plant aerial organs, called stomata, allow gas exchanges between the inner photosynthetic tissue and the atmosphere. Regulation of stomatal aperture, preventing excess transpirational vapor loss, relies on turgor changes of two highly differentiated epidermal cells surrounding the pore, the guard cells. Increased guard cell turgor due to increased solute accumulation results in stomatal opening, whereas decreased guard cell turgor due to decreased solute accumulation results in stomatal closing. Here we provide direct evidence, based on reverse genetics approaches, that the Arabidopsis GORK Shaker gene encodes the major voltage-gated outwardly rectifying K+ channel of the guard cell membrane. Expression of GORK dominant negative mutant polypeptides in transgenic Arabidopsis was found to strongly reduce outwardly rectifying K+ channel activity in the guard cell membrane, and disruption of the GORK gene (T-DNA insertion knockout mutant) fully suppressed this activity. Bioassays on epidermal peels revealed that disruption of GORK activity resulted in impaired stomatal closure in response to darkness or the stress hormone azobenzenearsonate. Transpiration measurements on excised rosettes and intact plants (grown in hydroponic conditions or submitted to water stress) revealed that absence of GORK activity resulted in increased water consumption. The whole set of data indicates that GORK is likely to play a crucial role in adaptation to drought in fluctuating environments
Sexual reproduction in plants requires elongation of the pollen tube through the transmitting tissues toward the ovary. Tube growth rate is a major determinant of pollen competitive ability. We report that a K + channel of the Shaker family in Arabidopsis, SPIK, plays an important role in pollen tube development. SPIK was found to be specifically expressed in pollen. When SPIK was heterologously expressed in COS cells, its product formed hyperpolarization-activated K + channels. Disruption (T-DNA insertion) of the SPIK coding sequence strongly affected inwardly rectifying K + -channel activity in the pollen-grain plasma membrane. Measurements of membrane potential in growing pollen tubes yielded data compatible with a contribution of SPIK to K + influx. In vitro pollen germination assays were performed, revealing that the disruption results in impaired pollen tube growth. Analysis of the transmission rate of the disrupted allele in the progeny of heterozygous plants revealed a decrease in pollen competitive ability, the probability of fertilization by mutant pollen being ∼1.6 times lower than that by wild-type pollen. The whole set of data supports the hypothesis that functional expression of SPIK plays a role in K + uptake in the growing pollen tube, and thereby in tube development and pollen competitive ability.
The AKT2 K ؉ channel is endowed with unique functional properties, being the only weak inward rectifier characterized to date in Arabidopsis. The gene is expressed widely, mainly in the phloem but also at lower levels in leaf epiderm, mesophyll, and guard cells. The AKT2 mRNA level is upregulated by abscisic acid. By screening a two-hybrid cDNA library, we isolated a protein phosphatase 2C (AtPP2CA) involved in abscisic acid signaling as a putative partner of AKT2. We further confirmed the interaction by in vitro binding studies. The expression of AtPP2CA (  -glucuronidase reporter gene) displayed a pattern largely overlapping that of AKT2 and was upregulated by abscisic acid. Coexpression of AtPP2CA with AKT2 in COS cells and Xenopus laevis oocytes was found to induce both an inhibition of the AKT2 current and an increase of the channel inward rectification. Site-directed mutagenesis and pharmacological analysis revealed that this functional interaction involves AtPP2CA phosphatase activity. Regulation of AKT2 activity by AtPP2CA in planta could allow the control of K ؉ transport and membrane polarization during stress situations. INTRODUCTIONPotassium is the most abundant cation in the cytoplasm of the living cell, where it is involved in the regulation of ionic strength, osmotic potential, and membrane polarization. K ϩ channels of the so-called Shaker family (nine genes in Arabidopsis) have been shown to play a role in K ϩ uptake by the root periphery (AKT1: Lagarde et al., 1996; Hirsch et al., 1998), K ϩ secretion into the root xylem sap (SKOR: Gaymard et al., 1998), K ϩ transport in the phloem tissues (AKT2: Marten et al., 1999;Lacombe et al., 2000), or K ϩ inward (KAT1: Ichida et al., 1997;Szyroki et al., 2001; KAT2: Pilot et al., 2001) and outward (GORK: Ache et al., 2000) fluxes in guard cells, leading to stomatal opening/closing.To adapt to fluctuating K ϩ availability in the environment and to cope with other stresses, plants need to tightly regulate K ϩ transport at both the whole plant and the cell level (Kochian and Lucas, 1988;Schroeder et al., 1994). Studies aimed at revealing the molecular determinants of these regulations have highlighted mechanisms likely to target Shaker K ϩ channels at both the transcriptional and posttranslational levels. Expression studies have revealed that transcript levels of Shaker channels are sensitive to hormones (Gaymard et al., 1998;Philippar et al., 1999;Lacombe et al., 2000), sugar synthesis and accumulation, and environmental signals (Deeken et al., 2000). At the post-translational level, indications have been found for regulation by ATP and cyclic GMP (Hoshi, 1995), phosphorylation events (Li et al., 1994; Armstrong et al., 1995; Tang and Hoshi, 1999), functional interactions with the cytoskeleton (Hwang et al., 1997), 14-3-3 proteins (Saalbach et al., 1997 Booij et al., 1999), sulfonylurea receptors (Leonhardt et al., 1997), syntaxins (Leyman et al., 1999), and G proteins (Wu andWang et al., 2001).Searches for interacting proteins have been focused on t...
Understanding how mosquito vectors and malaria parasites interact is of fundamental interest, and it also offers novel perspectives for disease control. Both the genetic and environmental contexts are known to affect the ability of mosquitoes to support malaria development and transmission, i.e., vector competence. Although the role of environment has long been recognized, much work has focused on host and parasite genetic effects. However, the last few years have seen a surge of studies revealing a great diversity of ways in which non-genetic factors can interfere with mosquito-Plasmodium interactions. Here, we review the current evidence for such environmentally mediated effects, including ambient temperature, mosquito diet, microbial gut flora, and infection history, and we identify additional factors previously overlooked in mosquito-Plasmodium interactions. We also discuss epidemiological implications, and the evolutionary consequences for vector immunity and parasite transmission strategies. Finally, we propose directions for further research and argue that an improved knowledge of non-genetic influences on mosquito-Plasmodium interactions could aid in implementing conventional malaria control measures and contribute to the design of novel strategies.
Aim To investigate the phylogeographical structure of the Guinea multimammate mouse, Mastomys erythroleucus (Temminck, 1853), a widespread murid rodent in sub‐Saharan (Sahel and Sudan) savannas, for a better understanding of the impacts of geographical and historical factors on the evolutionary history of this species, in the context of the growing database of phylogeographical studies of African savanna mammal species. Location Sahel and Sudan savannas, Africa. Methods We sequenced the whole cytochrome b gene in 211 individuals from 59 localities distributed from Senegal to Ethiopia. Sequence data were analysed using both phylogenetic (several rooted tree‐construction methods, median‐joining networks) and population genetic methods (spatial analyses of molecular variance, mismatch distributions). Results Haplotypes were distributed into four major monophyletic groups corresponding to distinct geographical regions across a west–east axis. Diversification events were estimated to have occurred between 1.16 and 0.18 Ma. Main conclusions Vicariance events related to the fragmentation of savanna habitats during the Pleistocene era may explain the phylogeographical patterns observed. Genetic structure was consistent with a role of major Sahelian rivers as significant barriers to west–east dispersal. Recent demographic expansions probably occurred during arid phases of the Holocene with the southward expansion of savannas.
BackgroundThe success of current control tools in combatting malaria vectors is well established. However, sustained residual transmission of Plasmodium parasites persists. Mass drug administration (MDA) to humans of the endectocide ivermectin for vector control is receiving increasing attention. However, vectors feeding upon animals escape this promising approach. Zoophagy of mosquitoes sustains both the vector population and endemic population of vector-borne pathogens. Therefore, only a strategy that will combine ivermectin MDAs targeted at humans and their peridomestic animals could be successful at controlling residual malaria transmission.MethodsBurkinabé cattle have been treated with injectable therapeutic dose of ivermectin (0.2 mg/kg of body weight) to render blood meals toxic to field representative populations of Anopheles coluzzii carrying the kdr mutation. Direct skin-feeding assays were performed from 2 to 28 days after injection (DAI) and mosquitoes were followed for their survival, ability to become gravid and fecundity. Membrane feeding assays were further performed to test if an ivermectin blood meal taken at 28 DAI impacts gametocyte establishment and development in females fed with infectious blood.ResultsThe mosquitocidal effect of ivermectin is complete for 2 weeks after injection, whether 12 days cumulative mortalities were of 75 and 45 % the third and fourth weeks, respectively. The third week, a second ivermectin blood meal at sub-lethal concentrations further increased mortality to 100 %. Sub-lethal concentrations of ivermectin also significantly decreased egg production by surviving females, increasing further the detrimental effect of the drug on vector densities. Although females fitness was impaired by sub-lethal ivermectin blood meals, these did not diminish nor increase their susceptibility to infection.ConclusionThis study demonstrates the potential of integrated MDA of ivermectin to both human and peridomestic cattle to target vector reservoirs of residual malaria transmission. Such integration lies in ‘One-Health’ efforts being implemented around the globe, and would be especially relevant in rural communities in Africa where humans are also at risk of common zoonotic diseases.Electronic supplementary materialThe online version of this article (doi:10.1186/s12936-015-1001-z) contains supplementary material, which is available to authorized users.
The African malaria vector Anopheles gambiae is polymorphic for alternative arrangements on the left arm of chromosome 2 (2La and 2L+(a)) that are non-randomly distributed with respect to degree of aridity. Detailed studies on the ecological role of inversion 2La have been hindered by the technical demands of traditional karyotype analysis and by sex- and stage-specific limitations on the availability of polytene chromosomes favorable for analysis. Recent molecular characterization of both inversion breakpoints presented the opportunity to develop a polymerase chain reaction (PCR)-based method for karyotype analysis. Here we report the development of this molecular diagnostic assay and the results of extensive field validation. When tested on 765 An. gambiae specimens sampled across Africa, the molecular approach compared favorably with traditional cytologic methods, correctly scoring > 94% of these specimens. By providing ready access to the 2La karyotype, this tool lays groundwork for future studies of the ecological genomics of this medically important species.
Potassium is the most abundant cation in the cytosol, where it plays a role in basal functions. Rapid uptake and distribution of K+ is therefore required for plant growth. Three members of the so-called Shaker K+ channel gene family (nine genes identified in Arabidopsis) play a role in these transports: AKT1, SKOR and AKT2. The encoded proteins are involved in K+ uptake by the root, K+ secretion into the xylem sap and K+ transport in the phloem tissues, respectively. Using the GUS reporter strategy, we have found that another Shaker channel gene, AtKC1, is expressed in epidermal and cortical cells in roots (supporting the hypothesis of a role in K+ uptake from the soil, together with AKT1), and in trichomes and hydathodes in leaves. These four genes were selected for expression studies, and two-hybrid experiments were performed for channels displaying overlapping expression patterns. The data support the hypothesis that physical interactions could occur in planta between AtKC1 and AKT1, and between AKT1 and AKT2. Potassium deficiency, salt stress and hormonal treatments (ABA, BA, 2,4-D) were found to differentially affect channel mRNA levels, each channel displaying its own regulation pattern. The most prominent effects were (1) a strong induction of AtKC1 transcript accumulation in leaves (hydathodes, trichomes and leaf epidermis) in response to NaCl treatment, suggesting a key role of the protein in adaptation to saline conditions, and (2) a strong decrease in SKOR transcript levels by hormones, supporting the hypothesis that K+ secretion into the xylem sap is under tight hormonal control.
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