On the role of CD4+ T cells and pathogenic mechanisms in HSV-1-induced ocular disease : herpetic stromal keratitis

Experimental infection of mice by scarification of cornea with HSV exhibits ocular disease pattem similar to the histopathological manifestations seen in man. An experiment in athymic mice that did not succumb to herpetic stromal keratitis (HSK) showed that HSK is an immunopathological manifestation of host T cell mediated immune response. It was confirmed later that following infection of corneas in the murine model with RE strain of HSV-1, the immunopathology is manifested by CD4+T lymphocytes that exhibit the Thl cytokine profile. A prominent cell type present in HSK lesions is the neutrophil (PMN), the infiltration of which is a biphasic event following HSV infection of immunocompetent mice, an initial transient infiltration appeared to be triggered by replicating virus and a second more intense invasion along with other inflammatory cells including CD4+ cells. This secondary PMN invasion episode, seemingly orchestrated by CD4+T cells, is called HSK and is not apparently triggered by replicating virus which remains undetectable at this stage. By taking advantage of cell depletion and adoptive transfer studies, the type of cells involved and sequence of cellular events in initiation and effector phase have been addressed. While some of these approaches could explain one facet of HSK immunopathology, identifying the antigen(s) (viral and/or host) to which T cells respond is another interesting field. Some of the recent observations lend credence to the growing notion that HSK represents an autoreactive inflammatory reaction set off by HSV that unmasks sequestered corneal antigens is normally accessible to the immune system. In this study, experiments were designed to address such issues and hence primarily focussed on elucidating the mechanisms which drive the inflammatory responses in HSV-infected murine cornea. Experiments described in this dissertation are aimed at T cell specificity, role of viral antigens in lesion development and mechanisms that drive the inflammatory response in the infected cornea. The series of experiments described here and their results indicate \u27nonspecific activation of CD4+T cells\u27 as a novel mechanism in herpes-induced stromal keratitis. Following an overview of herpes induced ocular disease in part 1, part 2 in addition to supporting HSK as a T cell-mediated immunoinflammatory disease, the results also show the potential for controlling the disease by coreceptor modulation. In part 3, characterization of a unique mouse model with skewed T cell repertoire (Tg- SCID) has been described which is useful for studying antigen specificity in T cell-mediated immunopathological disorders. Results of experiments described in part 4 propose nonspecific activation of T cells and hence the cytokine milieu by virus replication as a novel mechanism by which CD4+ T cells could orchestrate a virus induced ocular immunopathology. Finally, using adoptive transfer protocols, part 5 examines whether lesion development in Tg-SCID mice is virus replication, persistence and CD4+ T cell dependent. Additionally, in part 5, the differential role of HSV-specific antiserum and CD8+ T cells on virus dissemination was analyzed in the infected cornea

DET1 and COP1 Modulate the Coordination of Growth and Immunity in Response to Key Seasonal Signals in Arabidopsis

This is the final version. Available from Elsevier via the DOI in this record.Plant growth and development and outcomes of plant-microbe interactions are defined by coordinated responses to seasonal signals. The mechanisms that control the coordinated regulation of growth and immunity are not well understood. Here, we show that a common signaling module integrates environmental signals, such as photoperiod and temperature, to regulate the growth-defense balance. Key light-signaling components De-Etiolated 1 (DET1) and Constitutive Photomorphogenic 1 (COP1) negatively regulate immunity and are essential for immune modulation by photoperiod and temperature. Our results show that this is regulated by the transcription factor Phytochrome Interacting Factor 4 (PIF4), suggesting that the DET1/COP1-PIF4 module acts as a central hub for the control of growth and immunity in response to seasonal signals. These findings provide a regulatory framework for environmental signal integration. In plants, adaptive traits such as growth and immunity are strongly influenced by the environment. How multiple seasonal signals are integrated is not well understood. Gangappa and Kumar show that a common signaling module comprising DET1, COP1, and PIF4 coordinates growth and immunity in response to key seasonal signals

DET1 and HY5 Control PIF4-Mediated Thermosensory Elongation Growth through Distinct Mechanisms

This is the final version. Available from Elsevier via the DOI in this record.Plant growth and development are defined by environmental cues. The transcription factor PHYTOCHROME INTERACTING FACTOR 4 (PIF4) is the central signaling hub that integrates environmental cues, including light and temperature, to regulate growth and development. The thermosensory mechanisms controlling the PIF4-mediated temperature response, and its integration with other environmental responses, remain poorly understood. DE-ETIOLATED 1 (DET1) and CONSTITUTIVE PHOTOMORPHOGENESIS 1 (COP1), key regulators of light signaling, have been proposed to control thermosensory growth by transcriptional regulation of PIF4, through ELONGATED HYPOCOTYL 5 (HY5). Here, we show that DET1/COP1 and HY5 regulate thermosensory elongation through distinct mechanisms. DET1 and COP1 are essential for promoting PIF4 expression and stabilizing PIF4 protein. Furthermore, HY5 inhibits elongation growth through competitive chromatin binding to PIF4 targets, not through transcriptional regulation of PIF4. Our findings reveal a mechanistic framework in which DET1/COP1 and HY5 regulatory modules act independently to regulate growth through the environmental signal integrator PIF4.Biotechnology and Biological Sciences Research Counci

SWR1 Chromatin-Remodeling Complex Subunits and H2A.Z Have Non-overlapping Functions in Immunity and Gene Regulation in Arabidopsis

This is the final version. Available from Elsevier via the DOI in this record.Incorporation of the histone variant H2A.Z into nucleosomes by the SWR1 chromatin remodeling complex is a critical step in eukaryotic gene regulation. In Arabidopsis, SWR1c and H2A.Z have been shown to control gene expression underlying development and environmental responses. Although they have been implicated in defense, the specific roles of the complex subunits and H2A.Z in immunity are not well understood. In this study, we analyzed the roles of the SWR1c subunits, PHOTOPERIOD-INDEPENDENT EARLY FLOWERING1 (PIE1), ACTIN-RELATED PROTEIN6 (ARP6), and SWR1 COMPLEX 6 (SWC6), as well as H2A.Z, in defense and gene regulation. We found that SWR1c components play different roles in resistance to different pathogens. Loss of PIE1 and SWC6 function as well as depletion of H2A.Z led to reduced basal resistance, while loss of ARP6 fucntion resulted in enhanced resistance. We found that mutations in PIE1 and SWC6 resulted in impaired effector-triggered immunity. Mutation in SWR1c components and H2A.Z also resulted in compromised jasmonic acid/ethylene-mediated immunity. Genome-wide expression analyses similarly reveal distinct roles for H2A.Z and SWR1c components in gene regulation, and suggest a potential role for PIE1 in the regulation of the cross talk between defense signaling pathways. Our data show that although they are part of the same complex, Arabidopsis SWR1c components could have non-redundant functions in plant immunity and gene regulation

PIF4 Coordinates Thermosensory Growth and Immunity in Arabidopsis

This is the final version. Available from Elsevier via the DOI in this record.This the final version. Available from Elsevier via the DOI in this record.Temperature is a key seasonal signal that shapes plant growth. Elevated ambient temperature accelerates growth and developmental transitions [1] while compromising plant defenses, leading to increased susceptibility [2, 3]. Suppression of immunity at elevated temperature is at the interface of trade-off between growth and defense [2, 4]. Climate change and the increase in average growth-season temperatures threaten biodiversity and food security [5, 6]. Despite its significance, the molecular mechanisms that link thermosensory growth and defense responses are not known. Here we show that PHYTOCHROME INTERACTING FACTOR 4 (PIF4)-mediated thermosensory growth and architecture adaptations are directly linked to suppression of immunity at elevated temperature. PIF4 positively regulates growth and development and negatively regulates immunity. We also show that natural variation of PIF4-mediated temperature response underlies variation in the balance between growth and defense among Arabidopsis natural strains. Importantly, we find that modulation of PIF4 function alters temperature sensitivity of defense. Perturbation of PIF4-mediated growth has resulted in temperature-resilient disease resistance. This study reveals a molecular link between thermosensory growth and immunity in plants. Elucidation of the molecular mechanisms that define environmental signal integration is key to the development of novel strategies for breeding temperature-resilient disease resistance in crops.Biotechnology and Biological Sciences Research CouncilInstitute Strategic ProgrammeInstitute Strategic ProgrammeEuropean Commissio

Eu ³⁺ Sequestration by Biogenic Nano-Hydroxyapatite Synthesized at Neutral and Alkaline pH

<p>Biogenic hydroxyapatite (bio-HA) has the potential for radionuclide capture and remediation of metal-contaminated environments. Biosynthesis of bio-HA was achieved via the phosphatase activity of a <i>Serratia sp</i>. supplemented with various concentrations of CaCl₂ and glycerol 2-phosphate (G2P) provided at pH 7.0 or 8.6. Presence of hydroxyapatite (HA) was confirmed in the samples by X-ray powder diffraction analysis. When provided with limiting (1 mM) G2P and excess (5 mM) Ca²⁺ at pH 8.6, monohydrocalcite was found. This, and bio-HA with less (1 mM) Ca²⁺ accumulated Eu(III) to ∼31% and 20% of the biomineral mass, respectively, as compared to 50% of the mineral mass accumulated by commercial HA. Optimally, with bio-HA made at initial pH 7.0 from 2 mM Ca²⁺ and 5 mM G2P, Eu(III) accumulated to ∼74% of the weight of bio-HA, which was equal to the mass of the HA mineral component of the biomaterial. The implications with respect to potential bio-HA-barrier development in situ or as a remediation strategy are discussed.</p

Molecular interactions of BBX24 and BBX25 with HYH, HY5 HOMOLOG, to modulate Arabidopsis seedling development

BBX24 and BBX25 are two important transcriptional regulators, which regulate seedling photomorphogenesis in Arabidopsis. Very recently, we have shown that BBX24 and BBX25 negatively regulate the expression of BBX22, reducing the function of HY5, by physically interacting with its bZIP domain.1 Furthermore, HY5 HOMOLOG, HYH, has been reported to heterodimerize with HY5 and enhances its photomorphogenic function in seedling de-etiolation by serving as coactivator.8 Here, we further report that BBX24 and BBX25 physically interact with HYH. The physical interactions of BBX24 and BBX25 with HYH could lead to depletion of HYH molecules from the active pool and, thus indirectly, reduce the function of HY5 in promoting photomorphogenesis. Hence, our results suggest another mode of regulation by which BBX24 and BBX25 exert their negative effects on HY5 indirectly through HYH for the fine-tuning of seedling photomorphogenesis.Fil: Gangappa, Sreeramaiah N.. Gothenburg University. Department of Biological and Environmental Sciences; SueciaFil: Holm, Magnus. Gothenburg University. Department of Biological and Environmental Sciences; SueciaFil: Botto, Javier Francisco. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura; Argentin

Detection of epistasis through triple test cross (TTC) analysis in maize (Zea mays L.)

The present study was carried out to detect the epistasis present in two cross of maize through triple test cross (TTC) analysis. The mean squares due to total epistasis was highly significant at P?0.01 for all the characters in both C-I and C-II, except for ear length in C-I. The i type of epistasis was highly significant for the traits such as days totasseling, days to silking, earlength, ear circumference, kernels row-1,100 grain weight and shelling percentage in C-I and in C-II, ‘i’ type was non-significant for ASI, ear length, kernels row-1and grain yield plot-1. Both j type and l type of epistasis were significant for all characters in both C-I and C-II, except for ear length in C-I and days to silking in C-II. The estimate of additive genetic component (D) was highly significant for all characters in both C-I and C-II. Epistasis played a significant role in the inheritance of all the characters in both C-I and C-II except for ear length in C-I. Both additive and dominance components of genetic variance with a predominance of dominance genetic variance played an important role in the inheritance of all the quantitative traits except ear length in C-I and kernel rows ear-1 in C-II

Arabidopsis CAM7 and HY5 physically interact and directly bind to the HY5 promoter to regulate its expression and thereby promote photomorphogenesis

Arabidopsis thaliana CALMODULIN7 (CAM7), a unique member of the calmodulin gene family, plays a crucial role as a transcriptional regulator in seedling development. The elongated HYPOCOTYL5 (HY5) bZIP protein, an integrator of multiple signaling pathways, also plays an important role in photomorphogenic growth and light-regulated gene expression. CAM7 acts synergistically with HY5 to promote photomorphogenesis at various wavelengths of light. Although the genetic relationships between CAM7 and HY5 in light-mediated seedling development have been demonstrated, the molecular connectivity between CAM7 and HY5 is unknown. Furthermore, whereas HY5-mediated gene regulation has been fairly well investigated, the transcriptional regulation of HY5 is largely unknown. Here, we report that HY5 expression is regulated by HY5 and CAM7 at various wavelengths of light and also at various stages of development. In vitro and in vivo DNA–protein interaction studies suggest that HY5 and CAM7 bind to closely located T/G- and E-box cis-acting elements present in the HY5 promoter, respectively. Furthermore, CAM7 and HY5 physically interact and regulate the expression of HY5 in a concerted manner. Taken together, these results demonstrate that CAM7 and HY5 directly interact with the HY5 promoter to mediate the transcriptional activity of HY5 during Arabidopsis seedling development