Ethylene-mediated regulation of A2-type CYCLINs modulates hyponastic growth in arabidopsis

Upward leaf movement (hyponastic growth) is frequently observed in response to changing environmental conditions and can be induced by the phytohormone ethylene. Hyponasty results from differential growth (i.e. enhanced cell elongation at the proximal abaxial side of the petiole relative to the adaxial side). Here, we characterize Enhanced Hyponasty-d, an activation-tagged Arabidopsis (Arabidopsis thaliana) line with exaggerated hyponasty. This phenotype is associated with overexpression of the mitotic cyclin CYCLINA2;1 (CYCA2;1), which hints at a role for cell divisions in regulating hyponasty. Indeed, mathematical analysis suggested that the observed changes in abaxial cell elongation rates during ethylene treatment should result in a larger hyponastic amplitude than observed, unless a decrease in cell proliferation rate at the proximal abaxial side of the petiole relative to the adaxial side was implemented. Our model predicts that when this differential proliferation mechanism is disrupted by either ectopic overexpression or mutation of CYCA2;1, the hyponastic growth response becomes exaggerated. This is in accordance with experimental observations on CYCA2;1 overexpression lines and cyca2;1 knockouts. We therefore propose a bipartite mechanism controlling leaf movement: ethylene induces longitudinal cell expansion in the abaxial petiole epidermis to induce hyponasty and simultaneously affects its amplitude by controlling cell proliferation through CYCA2;1. Further corroborating the model, we found that ethylene treatment results in transcriptional down-regulation of A2-type CYCLINs and propose that this, and possibly other regulatory mechanisms affecting CYCA2;1, may contribute to this attenuation of hyponastic growth. Plants have acquired mechanisms to adjust growth and secure reproduction under unfavorable environmental conditions. Among the strategies to avoid adverse conditions is upward leaf movement, called hyponastic growth. This leaf reorientation is driven by unequal growth rates between adaxial and abaxial sides of the petiole (Cox et al., 2004; Polko et al., 2012b). Arabidopsis (Arabidopsis thaliana) exhibits hyponasty upon several environmental signals (e.g. submergence, waterlogging, proximity of neighboring vegetation, low red:far-red light ratios, reduced blue light fluence rates, low light intensities, and high temperatures; Millenaar et al., 2005, 2009; Mullen et al., 2006; Koini et al., 2009; Moreno et al., 2009; Van Zanten et al., 2009; Keuskamp et al., 2010; Keller et al., 2011; Vasseur et al., 2011; De Wit et al., 2012; Rauf et al., 2013; Dornbusch et al., 2014). Hyponasty alleviates the impact of environmental stresses (Van Zanten et al., 2010b). During submergence, it allows reestablishment of gas exchange with the atmosphere (e.g. Cox et al., 2003); at high plant densities, it positions the leaves in better lit layers of the canopy to improve light interception (e.g. De Wit et al., 2012); and at high temperatures, it improves the cooling capacity of the leaves (Crawford et al., 2012; Bridge et al., 2013). The cellular basis of hyponastic growth in Rumex palustris (Cox et al., 2004) and Arabidopsis (Polko et al., 2012b; Rauf et al., 2013) has been characterized. Ethylene causes reorientation of cortical microtubules (CMTs) in the petiole, which leads to longitudinal cell expansion in an approximately 2-mm-long epidermal cell zone at the proximal part of the abaxial side of the organ (Polko et al., 2012b). The interactions between several hormones (e.g. ethylene, abscisic acid, GAs, and auxin) in controlling hyponasty under various conditions have been studied (Mullen et al., 2006; Benschop et al., 2007; Millenaar et al., 2009; Van Zanten et al., 2009, 2010b; Peña-Castro et al., 2011). The volatile phytohormone ethylene is a key component in the complex regulatory network of hyponastic growth. Ethylene is the trigger and a positive regulator of hyponastic growth in submerged and waterlogged Arabidopsis (Millenaar et al., 2005, 2009; Van Zanten et al., 2010b; Rauf et al., 2013) and a negative regulator of high temperature-induced hyponasty (Van Zanten et al., 2009), but is not involved in low light-induced hyponastic growth in this species (Millenaar et al., 2009). Abscisic acid antagonizes ethylene-induced hyponasty (Benschop et al., 2007) and is a positive regulator of high temperature-induced hyponastic growth (Van Zanten et al., 2009). The growth-promoting GAs positively regulate hyponastic response to all three environmental signals (Peña-Castro et al., 2011), whereas auxins promote low light and high temperature-induced hyponastic growth (Millenaar et al., 2005; Koini et al., 2009; Van Zanten et al., 2009), as well as low red:far-red- and low blue light-induced hyponasty (Moreno et al., 2009; Keller et al., 2011). Finally, brassinosteroids also positively regulate ethylene-induced hyponasty (Polko et al., 2013). Despite the extensive knowledge on hormonal regulation of hyponasty, little is known about the molecular genetic mechanisms that drive this response. One notable exception is the study by Rauf et al. (2013), who showed that hyponastic growth in Arabidopsis in response to root waterlogging is controlled by the NAC (for No Apical Meristem [NAM], Arabidopsis Transcription Activation Factor) transcription factor SPEEDY HYPONASTIC GROWTH that directly affects expression of the ethylene biosynthesis gene 1-AMINOCYCLOPROPANE-1-CARBOXYLIC ACID (ACC) OXIDASE5. Here, we followed a forward genetic approach to identify unique components that control hyponastic growth in Arabidopsis. From a population of activation-tagged plants (Weigel et al., 2000), we isolated Enhanced Hyponasty-D (EHY-D), which showed exaggerated hyponasty under exogenous ethylene application, low light intensities, and high temperature. We found that ectopic expression of the core cell cycle regulator CYCLINA2;1 (CYCA2;1) caused the exaggerated ethylene-induced leaf movement of EHY-D. Mathematical analyses indicated that, besides promoting cell expansion, ethylene can also attenuate the amplitude of hyponasty by affecting differential cell proliferation in the petiole of wild-type plants. We suggest that this occurs through ethylene-dependent effects on CYCA2;1 levels, activity, or sensitivity in petioles of wild-type plants. The ethylene-mediated transcriptional regulation of CYCA2;1 observed here could contribute to this. In EHY-D, however, ethylene-mediated effects on cell proliferation are overruled by ectopic CYCA2;1 overexpression, which consequently results in enhanced hyponasty, in accordance with the predictions of our model. Correspondingly, cyca2;1 knockout lines where ethylene cannot affect CYCA2;1-mediated cell proliferation also exhibited enhanced hyponasty. Our data therefore describe a mechanism by which hyponastic growth is kept within limits, through a bipartite role for ethylene: within the same organ, ethylene initiates hyponastic growth by promoting cell elongation, while simultaneously attenuating the response by regulation of A2-type CYCLIN-mediated cell proliferation

The history of foot-and-mouth disease virus serotype C: the first known extinct serotype?Abstract

Foot-and-mouth disease (FMD) is a highly contagious animal disease caused by an RNA virus subdivided into seven serotypes that are unevenly distributed in Asia, Africa, and South America. Despite the challenges of controlling FMD, since 1996 there have been only two outbreaks attributed to serotype C, in Brazil and in Kenya, in 2004. This article describes the historical distribution and origins of serotype C and its disappearance. The serotype was first described in Europe in the 1920s, where it mainly affected pigs and cattle but as a less common cause of outbreaks than serotypes O and A. No serotype C outbreaks have been reported in Europe since vaccination stopped in 1990. FMD virus is presumed to have been introduced into South America from Europe in the nineteenth century, although whether serotype C evolved there or in Europe is not known. As in Europe, this serotype was less widely distributed and caused fewer outbreaks than serotypes O and A. Since 1994, serotype C had not been reported from South America until four small outbreaks were detected in the Amazon region in 2004. Elsewhere, serotype C was introduced to Asia, in the 1950s to the 1970s, persisting and evolving for several decades in the Indian subcontinent and for eighteen years in the Philippines. Serotype C virus also circulated in East Africa between 1957 and 2004. Many serotype C viruses from European and Kenyan outbreaks were closely related to vaccine strains, including the most recently recovered Kenyan isolate from 2004. International surveillance has not confirmed any serotype C cases, worldwide, for over 15 years, despite more than 2,000 clinical submissions per year to reference laboratories. Serology provides limited evidence for absence of this serotype, as unequivocal interpretation is hampered by incomplete intra-serotype specificity of immunoassays and the continued use of this serotype in vaccines. It is recommended to continue strengthening surveillance in regions of FMD endemicity, to stop vaccination against serotype C and to reduce working with the virus in laboratories, since inadvertent escape of virus during such activities is now the biggest risk for its reappearance in the&nbsp;field.</p

Determination of common genetic variants within the non-structural proteins of foot-and-mouth disease viruses isolated in sub-Saharan Africa

The non-structural proteins of foot-and-mouth disease virus (FMDV) are responsible for RNA replication, proteolytic processing of the viral polyprotein precursor, folding and assembly of the structural proteins and modification of the cellular translation apparatus. Investigation of the amino acid heterogeneity of the non-structural proteins of seventy- nine FMDV isolates of SAT1, SAT2, SAT3, A and O serotypes revealed between 29 and 62% amino acid variability. The Leader protease (Lpro) and 3A proteins were the most variable whilst the RNA-dependent RNA polymerase (3Dpol) the most conserved. Phylogeny based on the non-structural protein-coding regions showed separate clusters for southern African viruses for both the Lpro and 3C protease (3Cpro) and sequences unique to this group of viruses, e.g. in the 2C and 3Cpro proteins. These groupings were unlike serotype groupings based on structural protein-coding regions. The amino acid substitutions and the nature of the naturally occurring substitutions provide insight into the functional domains and regions of the non-structural proteins that are critical for structure–function. The Lpro of southern African SAT type isolates differed from A, O and SAT isolates in northern Africa, particularly in the auto-processing region. Three-dimensional structures of the 3C protease (3Cpro) and 3Dpol showed that the observed variation does not affect the enzymatic active sites or substrate binding sites. Variation in the 3Cpro cleavage sites demonstrates broad substrate specificity.THRIP of the National Research Foundation of South Africahttp://www.elsevier.com/locate/vetmic2016-05-31hb201