ABSTRACT
BACKGROUND: Herbo-mineral-metallic formulations are an inseparable part of the Ayurveda system of traditional medicine. Hridayarnava Rasa (HR) is a preparation containing metals like copper, sulphur, and mercury in processed forms and other herbs that do not produce toxic effects and adverse drug reactions when taken in appropriate dosage. Ayurveda practitioners use it in treating cardiac diseases like hypertension, cardiotoxicity and many more. The rasa-aushadhis possess characteristics such as rapid efficacy, little dosage required, and extensive therapeutic applicability. Hridayarnava Rasa [AFI Part-1, 20:55] has been employed for the treatment of various diseases from ancient times. A systematic study of these formulations manufacturing is required to maintain their quality, safety, and efficacy is a need of time to protect the immense faith of patients in Ayurveda. OBJECTIVES: The present study aimed to prepare HR as per standard operating procedures mentioned in the classical text and to characterize it physio-chemically using advanced analytical techniques. MATERIALS AND METHODS: HR was prepared and physicochemical analyses and assay of elements by ICP-AES were carried out as per Ayurvedic Pharmacopoeia of India (API). Powder X-ray diffraction (XRD), Field emission gun scanning electron microscopy with energy dispersive spectroscopy (FEG SEM, EDAX), CHNS-O analysis, Fourier Transform Infrared Spectroscopy (FTIR), Thermo-gravimetric analysis (TGA), Particle size distribution analysis (PSD) was carried out. RESULTS: The XRD analysis of HR showed the presence of unreacted sulphur and sulfides of copper and mercury. FEG SEM revealed the particles in the form of aggregates as nanocrystallites in the range of 100-1000 nm. Elemental analysis showed the presence of copper, sulphur, and mercury in major, along with traces of iron, calcium, sodium, potassium, and magnesium. In FTIR analysis, 18 peaks were observed, which strongly suggests the presence of various organic groups. In the TGA, four peaks were seen, which can be attributed to sulphur volatilization and oxidative changes in mercury. In PSD analysis, 50% of the material was found below 16.40 µm. CONCLUSION: To establish a piece of fundamental knowledge and ensure uniformity of these rasa-aushadhis, it is imperative to conduct an analysis of their characteristics as per classical texts and modern analytical techniques. Additionally, it is crucial to investigate the significance of each procedural step included in the preparation process. The inferences drawn are helpful as an essential aid for quality assurance and standardization of this herbo-mineral-metallic formulation.
ABSTRACT
A 2.45 GHz electron cyclotron resonance (ECR) ion source coupled to a gas-jet skimmer system has been developed for the online production of radioactive ion beams (RIBs). Using radial injection of gas jet in the ion source, RIBs of 11C1+, 11CO21+, and 11CO1+ have been produced online with beam intensity up to about 9 × 103 particles per second for a 1 µA primary proton beam bombarding a nitrogen gas target. The details of the gas jet coupled ECR ion source and the results for stable isotope beams and RIBs are reported.
ABSTRACT
Trophoblast stem (TS) cells have the unique capacity to differentiate into specialized cell types, including extravillous trophoblast (EVT) cells. EVT cells invade into and transform the uterus where they act to remodel the vasculature facilitating the redirection of maternal nutrients to the developing fetus. Disruptions in EVT cell development and function are at the core of pregnancy-related disease. WNT-activated signal transduction is a conserved regulator of morphogenesis of many organ systems, including the placenta. In human TS cells, activation of canonical WNT signaling is critical for maintenance of the TS cell stem state and its downregulation accompanies EVT cell differentiation. We show that aberrant WNT signaling undermines EVT cell differentiation. Notum, palmitoleoyl-protein carboxylesterase (NOTUM), a negative regulator of canonical WNT signaling, was prominently expressed in first trimester EVT cells developing in situ and upregulated in EVT cells derived from human TS cells. Furthermore, NOTUM was required for human TS cell differentiation to EVT cells. Activation of NOTUM in EVT cells is driven, at least in part, by endothelial PAS domain 1 (also called hypoxia-inducible factor 2 alpha). Collectively, our findings indicate that canonical WNT signaling is essential for maintenance of human trophoblast cell stemness and prevention of human TS cell differentiation. Downregulation of canonical WNT signaling via the actions of NOTUM is required for EVT cell differentiation.
ABSTRACT
Group VII ethylene-responsive factor (ERFVII) transcription factors are crucial for the adaption of plants to conditions that limit oxygen availability. A recent study by Zubrycka et al. reveals new aspects of ERFVII stabilization through the PLANT CYSTEINE OXIDASE (PCO)-N degron pathway and non-autonomous regulation in response to different endogenous and exogenous cues.
Subject(s)
Arabidopsis Proteins , Arabidopsis , Arabidopsis Proteins/metabolism , Arabidopsis/metabolism , Transcription Factors/genetics , Transcription Factors/metabolism , Hypoxia/genetics , Plants/genetics , Plants/metabolism , Oxygen/metabolism , Gene Expression Regulation, Plant/geneticsABSTRACT
N-terminal cysteine oxidases (NCOs) use molecular oxygen to oxidise the amino-terminal cysteine of specific proteins, thereby initiating the proteolytic N-degron pathway. To expand the characterisation of the plant family of NCOs (plant cysteine oxidases [PCOs]), we performed a phylogenetic analysis across different taxa in terms of sequence similarity and transcriptional regulation. Based on this survey, we propose a distinction of PCOs into two main groups. A-type PCOs are conserved across all plant species and are generally unaffected at the messenger RNA level by oxygen availability. Instead, B-type PCOs appeared in spermatophytes to acquire transcriptional regulation in response to hypoxia. The inactivation of two A-type PCOs in Arabidopsis thaliana, PCO4 and PCO5, is sufficient to activate the anaerobic response in young seedlings, whereas the additional removal of B-type PCOs leads to a stronger induction of anaerobic genes and impairs plant growth and development. Our results show that both PCO types are required to regulate the anaerobic response in angiosperms. Therefore, while it is possible to distinguish two clades within the PCO family, we conclude that they all contribute to restrain the anaerobic transcriptional programme in normoxic conditions and together generate a molecular switch to toggle the hypoxic response.
Subject(s)
Cysteine Dioxygenase , Oxygen , Cysteine , Phylogeny , HypoxiaABSTRACT
In this experimental work, we report our findings about a transition between states of even parity 5S1/2â5D5/2 of Rb atoms under different conditions of transit time broadening. For this purpose, the blue fluorescence emanating from the associated decay 5D5/2â6P3/2â5S1/2 channel is monitored. Based on the relative magnification of the laser beam, which is used in excitation between levels of even parity, the system exhibits two photon absorption (TPA) through blue fluorescence peaks. The TPA profile appears as a Lorentzian peak on a Doppler background. The change in the laser beam diameter directly influences the relative contributions of Lorentzian and Doppler parts. A direct inference drawn from of this study may be utilized in optimizing the extraction of the first derivative spectrum corresponding to 5Sâ5D transitions.
ABSTRACT
The rat possesses hemochorial placentation with deep intrauterine trophoblast cell invasion and trophoblast-guided uterine spiral artery remodeling, which resembles human placentation. Uterine spiral arteries are extensively remodeled to deliver sufficient supply of maternal blood and nutrients to the developing fetus. Inadequacies in these key processes negatively impact fetal growth and development. Recent innovations in genome editing combined with effective phenotyping strategies have provided new insights into placental development. Application of these research approaches has highlighted both conserved and species-specific features of hemochorial placentation. The review provides foundational information on rat hemochorial placental development and function during physiological and pathological states, especially as related to the invasive trophoblast cell-guided transformation of uterine spiral arteries. Our goal is to showcase the utility of the rat as a model for in vivo mechanistic investigations targeting regulatory events within the uterine-placental interface.
Subject(s)
Placenta , Trophoblasts , Animals , Female , Placenta/blood supply , Placentation/physiology , Pregnancy , Rats , Trophoblasts/physiology , Uterine Artery , Uterus/blood supplyABSTRACT
Introduction: Root and fruits of Bhumi Udumbara (Ficus semicordata Buch.-Ham. ex Sm.) are traditionally used in the treatment of aphthous complaints, leprosy, headache, abdominal diseases, bladder ailments, visceral obstruction, and various disorders. Aim: Present study reports the microscopic including powder microscopy, physiochemical and preliminary phytochemical characters of root and fruit of F. semicordata. Materials and methods: Root and fruit of the plant, after proper authentication, were evaluated following standard pharmacopoeial recommended procedures. Results: Striking characters of the F. semicordata root is the presence of profusely branched root-like structures, nearer to the trunk, and reaches the ground as a supporting root (aerial root or false root). The diagrammatic section of the root, circular in outline, made up of an outer cork, followed by cortex, vascular bundle, and central pseudo pith. Fruit is shortly peduncled, in pairs or clusters, globose in shape, hispid and warted surface, reddish brown in color. Loss on drying at 110°C was found to be 10.54% and 11.73% of root and fruit respectively. High-performance thin-layer chromatography results showed 2 peaks at 254 and 366 nm of root and 3 peaks at 254 and 1 peak at 366 nm of fruit respectively. Conclusion: F. semicordata is a small or medium-sized evergreen tree, bears a supporting root (aerial root or false root) nearer to the trunk region. Root reddish brown in color with smooth surface, prominent nodes, and internodes, presence of fruits over nodes and internodes is the key identifying character. Stone cells found in root and the presence of pollen grains, simple starch grains, and compound starch grains in fruit are the diagnostic characteristics of F. semicordata.
ABSTRACT
The oviduct is a site for early reproductive events including gamete maturation, fertilization, and early embryo development. Secretory cells lining the oviduct lumen synthesize and secrete proteins that interact with gametes and developing embryos. Although previous studies have identified some of the secretory proteins in the oviduct, however, knowledge and their precise specific functions in the oviduct are poorly understood. In this study, by using proteomic approach, we identified a secretory protein, Peroxiredoxin 6 (PRDX6), and evaluated its role in mediating early pregnancy events, fertilization, and embryo development in rabbit oviduct. The expression of PRDX6 was significantly higher in ampulla and isthmus sections of the oviduct in mated animal groups compared to non-mated controls. Furthermore, significant reduction in number of embryos recovered from PRDX6 siRNA-transfected oviductal horn was observed compared to the control contralateral horn. Moreover, in animals receiving PRDX6 siRNA in their oviductal horn, the number of implanted blastocysts was significantly less in the uterus as observed on day 9 post-coital (p.c.). Further, during embryo-rabbit oviduct epithelial cell (ROEC) co-culture, siRNA-mediated PRDX6 silencing attenuated the early embryonic development. Mechanistically, increased levels of ROS and expression of oxidative stress- and inflammation-related proteins were found in PRDX6 siRNA-treated ROEC cells as compared to control cells, implicating that ablation of PRDX6 in the oviduct creates a stress-induced micro-environment detrimental to early embryonic development in oviduct. Taken together, our data suggest that PRDX6 maintains an optimal micro-environment conducive to successful embryo development and can be considered as a candidate to evaluate its therapeutic potential in IVF strategies.
Subject(s)
Embryonic Development , Fertilization , Peroxiredoxin VI , Proteomics , Animals , Fallopian Tubes , Female , Oviducts/metabolism , Peroxiredoxin VI/metabolism , Pregnancy , Proteins/metabolism , RNA, Small Interfering/metabolism , RabbitsABSTRACT
Plant cells coated with hydrophobic compounds constitute a protective barrier to control movement of materials through plant tissues. In roots, the endodermis develops two barriers: the Casparian strips establish an apoplastic barrier and suberin lamellae prevent diffusion through the plasma membrane. Suberin is a complex biopolymer and its deposition is highly responsive to the environment. While the enzymatic framework involved in suberin biosynthesis is well characterized, subsequent steps in suberin formation and regulation remained elusive. Recent publications, studying suberin from a cell biological perspective, have enriched our knowledge on suberin transport and polymerization in the cell wall. These studies have also elucidated the molecular mechanisms controlling suberin biosynthesis and regulation as well as its physiological role in plant abiotic and biotic interactions.
Subject(s)
Arabidopsis , Arabidopsis/metabolism , Cell Wall/metabolism , Lipids/physiology , Plant Roots/metabolismABSTRACT
Suberin is a hydrophobic biopolymer that can be deposited at the periphery of cells, forming protective barriers against biotic and abiotic stress. In roots, suberin forms lamellae at the periphery of endodermal cells where it plays crucial roles in the control of water and mineral transport. Suberin formation is highly regulated by developmental and environmental cues. However, the mechanisms controlling its spatiotemporal regulation are poorly understood. Here, we show that endodermal suberin is regulated independently by developmental and exogenous signals to fine-tune suberin deposition in roots. We found a set of four MYB transcription factors (MYB41, MYB53, MYB92, and MYB93), each of which is individually regulated by these two signals and is sufficient to promote endodermal suberin. Mutation of these four transcription factors simultaneously through genome editing leads to a dramatic reduction in suberin formation in response to both developmental and environmental signals. Most suberin mutants analyzed at physiological levels are also affected in another endodermal barrier made of lignin (Casparian strips) through a compensatory mechanism. Through the functional analysis of these four MYBs, we generated plants allowing unbiased investigation of endodermal suberin function, without accounting for confounding effects due to Casparian strip defects, and were able to unravel specific roles of suberin in nutrient homeostasis.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/growth & development , Lipids/physiology , Proto-Oncogene Proteins c-myb/metabolism , Transcription Factors/metabolism , Arabidopsis/genetics , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Proto-Oncogene Proteins c-myb/genetics , Transcription Factors/geneticsABSTRACT
Transforming growth factor (TGFß) is known to play a major role in establishment and maintenance of endometriosis as reported by our group earlier, the underlying mechanism remains to be explored. We deciphered the involvement of TAK1 in TGFß1- induced cellular responses and delineated the signaling mechanism in human endometriotic cells. The endometriotic cells showed elevated expression of TGFß1 signaling-effector molecules. TGFß1 exposure to endometriotic cells induced the expression of the downstream target molecules indicating that TGFß1 is implicated in the commencement ofTAK1/NFκB-p65/Smad7 cascade. The silencing of TAK1 in endometriotic cells attenuated the TGFß1 -induced NFκB transcriptional activation and nuclear translocation of NFκB-p65 subunit. The pharmacological inhibition of NFκB by QNZ or knockdown of TAK1 reduced the expression of Smad7 and Cox2. The knockdown of TAK1 in endometriotic cells showed G1 phase cell-cycle arrest and showed low BrdU-incorporation in the presence of TGFß1. The inhibition of TAK1 attenuated the TGFß1 signaling activation indicating that TAK1 is a crucial mediator for TGFß1 action in endometriotic cells. The exposure of endometriotic cells to TAK1 inhibitor, celastrol caused activation of caspase-3 and -9 that led to PARP cleavage and induced apoptosis. Simultaneously, autophagy occurred in celastrol-treated and TAK1-silenced cells as was evidenced by the formation of autophagosome and the increased expression of autophagic markers. Thus, TAK1 activation appears to protect the growth of endometriotic cells by suppressing the cell death process. Overall, our study provided the evidence that of TAK1 significant in the endometriotic cell regulation and mediates a functional cross-talk between TGFß1 and NFκB-p65 that promotes the growth and inflammatory response in endometriotic cells.
Subject(s)
Autophagy , Endometriosis/metabolism , Endometriosis/pathology , MAP Kinase Kinase Kinases/antagonists & inhibitors , NF-kappa B/metabolism , Signal Transduction , Smad7 Protein/metabolism , Transforming Growth Factor beta1/metabolism , Apoptosis/drug effects , Autophagy/drug effects , Cell Cycle Checkpoints/drug effects , Cell Death/drug effects , Cell Line , Cell Proliferation/drug effects , Cells, Cultured , Cyclooxygenase 2/metabolism , Endometrium/pathology , Female , G1 Phase/drug effects , Gene Knockdown Techniques , Humans , Models, Biological , Pentacyclic Triterpenes/pharmacology , Signal Transduction/drug effects , Stromal Cells/drug effects , Stromal Cells/metabolismABSTRACT
Plants, including most crops, are intolerant to waterlogging, a stressful condition that limits the oxygen available for roots, thereby inhibiting their growth and functionality. Whether root growth inhibition represents a preventive measure to save energy or is rather a consequence of reduced metabolic rates has yet to be elucidated. In the present study, we gathered evidence for hypoxic repression of root meristem regulators that leads to root growth inhibition. We also explored the contribution of the hormone jasmonic acid (JA) to this process in Arabidopsis thaliana. Analysis of transcriptomic profiles, visualisation of fluorescent reporters and direct hormone quantification confirmed the activation of JA signalling under hypoxia in the roots. Further, root growth assessment in JA-related mutants in aerobic and anaerobic conditions indicated that JA signalling components contribute to active root inhibition under hypoxia. Finally, we show that the oxygen-sensing transcription factor (TF) RAP2.12 can directly induce Jasmonate Zinc-finger proteins (JAZs), repressors of JA signalling, to establish feedback inhibition. In summary, our study sheds new light on active root growth restriction under hypoxic conditions and on the involvement of the JA hormone in this process and its cross talk with the oxygen sensing machinery of higher plants.
ABSTRACT
Plants need to attune their stress responses to the ongoing developmental programmes to maximize their efficacy. For instance, successful submergence adaptation is often associated with a delicate balance between saving resources and their expenditure to activate measures that allow stress avoidance or attenuation. We observed a significant decrease in submergence tolerance associated with ageing in Arabidopsis thaliana, with a critical step between 2 and 3 weeks of post-germination development. This sensitization to flooding was concomitant with the transition from juvenility to adulthood. Transcriptomic analyses indicated that a group of genes related to abscisic acid and oxidative stress response was more highly expressed in juvenile plants than in adult ones. These genes are induced by the endomembrane tethered transcription factor ANAC017 that was in turn activated by submergence-associated oxidative stress. A combination of molecular, biochemical and genetic analyses showed that these genes are located in genomic regions that move towards a heterochromatic state with adulthood, as marked by lysine 4 trimethylation of histone H3. We concluded that, while the mechanisms of flooding stress perception and signal transduction were unaltered between juvenile and adult phases, the sensitivity that these mechanisms set into action is integrated, via epigenetic regulation, into the developmental programme of the plant.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/genetics , Epigenesis, Genetic , Oxygen/metabolism , Transcription Factors/metabolism , Abscisic Acid/metabolism , Adaptation, Physiological , Arabidopsis/physiology , Arabidopsis Proteins/genetics , Gene Expression Profiling , Germination , Oxidative Stress , Plant Growth Regulators/metabolism , Plants, Genetically Modified , Stress, Physiological , Transcription Factors/geneticsABSTRACT
BACKGROUND: Latex of Euphorbia caducifolia Haines. (Euphorbiaceae), botanical source of Rakta Snuhi, a caustic irritant, is being used in different Ayurvedic therapeutics, after proper processing (Shodhana) in some specific media. Shodhana of Snuhi latex with Chincha-Patra Swarasa (juice of tamarind leaves) using Raudra Yantra (instrument/pot kept under sunlight for drying) has been recommended in Ayurveda texts. Snuhi is one of the Upavisha (sub toxic group of herbal drugs) and a well-known plant in the Indian system of medicine. It is used in pharmaceutical procedures like preparation and processing of drugs. AIM: The aim of the study was to carry out Shodhana of E. caducifolia latex with Chincha-Patra Swarasa. MATERIAL AND METHODS: Fresh latex of Snuhi was collected from the Sapada area of Jamnagar, Gujarat and fresh leaves of Tamarindus indica L. were collected from the herbal garden of the university and Swarasa was extracted by traditional expression technique. Shodhana of Snuhi Kshira was done under sunlight dried and shade dried method by mixing it with leaf juice of Tamarindus indica L. in a glass bowl in different ratio i.e. Kshira: leaf juice was 8:4, 8:2, 8:1 respectively. RESULTS: This study reveals certain changes in physico-chemical parameters (pH) and organoleptic characters of processed E. caducifolia latex by Tamarind leaf juice both in shade-dried and sunlight-dried samples. Under HPTLC, Shodhana of E. caducifolia latex with Tamarind leaf juice alters the number of spots both sunlight-dried and shade-dried samples. In sun-dried sample, the number of spots increased when the concentration of Tamarind leaf juice is decreased in HPTLC study. The study reveals that in case of shade-dried Shodhita (processed) Snuhi latex sample, the concentration of lupeol increases with the increasing quantity of Tamarind leaf juice. CONCLUSION: Shodhana with Tamarind leaf juice changes both qualitative and quantitative property of Snuhi latex.
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INTRODUCTION: Shodhana (purification) is the process by which one can remove the impurity or toxicity of the raw drug and make the drug suitable for therapeutic purpose. Chitraka (Plumbago zeylanica Linn.) is well known drug in Ayurveda and root of this plant is being used for therapeutic purpose and requires purification before used as a medicine. AIMS AND OBJECTIVE: There is no data available for pharmacognostical and analytical profile of processed Chitraka, hence it was planned to develop SOP of processed Chitraka for its identity, purity and strength through pharmacognostical and analytical profile. MATERIALS AND METHODS: Chitraka roots were procured from Pharmacy, Gujarat Ayurved University, Jamnagar. Purification was done in five batches with Churnodaka (lime water). Organoleptic characters, microscopic features, pH, loss on drying, ash value, water soluble extracts, methanol soluble extracts and plumbagin quantification through high-performance thin layer chromatography (HPTLC) were carried out, before and after the purification. RESULTS: Average 98.07% yield of Chitraka was obtained after purification. Differences were found in the processed samples of Chitraka in organoleptic features, pharmacognostical characters and physicochemical parameters, which show the impact of purification procedure on Chitraka. In HPTLC profile, plumbagin content was 0.29% in unpurified Chitraka powder, where in it was noted 0.98% after purification. CONCLUSION: Increase in plumbagin content through pharmaceutical process of Chitraka purification with lime water indicates that, this operating procedure is simple, convenient and can be considered as standard procedure. The organoleptic features, pharmacognostical characters, values of physicochemical parameters and quantity of plumbagin of purified Chitraka powder may be utilized for quality assurance in future studies.
ABSTRACT
Axillary meristems (AMs) contribute to the growth of a plant, determining adult architecture and reproductive success in response to environmental stimuli. The missing flowers (mf) mutant of sunflower (Helianthus annuus) is defective in AM development. mf lacks shoot branching and ray flowers, occasionally producing few disk flowers. Here we demonstrated that a point mutation in the REGULATOR OF AXILLARY MERISTEM FORMATION-LIKE (Ha-ROXL) gene of mf generates a premature stop codon and therefore a nonfunctional bHLH transcription factor, no longer localized in the nucleus, where it should exert its function. Virus-induced gene silencing of Ha-ROXL also causes defects in disk and ray flower development. Ha-ROXL mRNA accumulates at the adaxial boundaries of leaves and AMs. During inflorescence development, Ha-ROXL is expressed in small arcs of cells before a clear separation between abaxial bracts and disk flower primordia. No Ha-ROXL mRNA accumulates in mf inflorescences. Several genes known to play roles in plant architecture, auxin transport, and flower development are differentially expressed in mf and Ha-ROXL-silenced plants. These results highlight the predominant role of Ha-ROXL in regulating AMs in sunflower. In dicot, mf is the first mutant for which the ROXL gene is also required for initiation of flower meristems.
Subject(s)
Flowers/growth & development , Genes, Plant , Helianthus/genetics , Meristem/growth & development , Plant Proteins/genetics , Transcription Factors/genetics , Helianthus/growth & development , Point MutationABSTRACT
The ability to perceive oxygen levels is crucial to many organisms because it allows discerning environments compatible with aerobic or anaerobic metabolism, as well as enabling rapid switch between these two energy strategies. Organisms from different taxa dedicate distinct mechanisms to associate oxygen fluctuations with biological responses. Following from this observation, we speculated that orthogonal oxygen sensing devices can be created by transfer of essential modules from one species to another in which they are not conserved. We expressed plant cysteine oxidase (PCOs) enzymes in Saccharomyces cerevisiae, to confer oxygen-conditional degradability to a bioluminescent protein tagged with the Cys-exposing N-degron typical of plant ERF-VII factors. Co-translation of a second luciferase protein, not subjected to oxygen-dependent proteolysis, made the resulting Double Luciferase Oxygen Reporter (DLOR) ratiometric. We show that DLOR acts as a proxy for oxygen dynamics in yeast cultures. Moreover, since DLOR activity was enabled by the PCO sensors, we employed this device to disclose some of their properties, such as the dispensability of nitric oxide for N-terminal cysteine oxidation and the individual performance of Arabidopsis PCO isoforms in vivo. In the future, we propose the synthetic DLOR device as a convenient, eukaryotic cell-based tool to easily screen substrates and inhibitors of cysteine oxidase enzymes in vivo. Replacement of the luminescent proteins with fluorescent proteins will further turn our system into a visual reporter for oxygen dynamics in living cells.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/metabolism , Cysteine Dioxygenase/metabolism , Oxygen/metabolism , Saccharomyces cerevisiae/metabolism , Arabidopsis/genetics , Arabidopsis Proteins/genetics , Cysteine Dioxygenase/genetics , Gene Expression , Luminescent Measurements , Luminescent Proteins/genetics , Luminescent Proteins/metabolism , Nitric Oxide/metabolism , Oxidation-Reduction , Proteolysis , Saccharomyces cerevisiae/geneticsABSTRACT
Microtubule (MT) dynamics plays a crucial role in fertilization and early embryonic development; however its involvement in uterus during embryo implantation remains unclear. Herein, we report the effect of microtubule depolymerization during embryo implantation in BALB/c mice. Intrauterine treatment with depolymerizing agent nocodazole at pre-implantation phase (D4, 07:00 h) in mice resulted into mitigation in receptivity markers viz. LIF, HoxA10, Integrin-ß3, IHH, WNT4 and led to pregnancy failure. MT depolymerization in endometrial epithelial cells (EECs) also inhibited the blastocyst attachment and the adhesion. The decreased expression of MT polymerization-related proteins TPPP and α/ß-tubulin in luminal and glandular epithelial cells along with the alteration in morphology of pinopodes in the luminal epithelium was observed in nocodazole receiving uteri. Nocodazole treatment also led to increased intracellular Ca+2 levels in EECs, which indicated that altered Ca+2 homeostasis might be responsible for implantation failure. Microtubule depolymerization inhibited WNT4 and Fz-2 interaction, thereby suppressing the downstream WNT4/CaMKIIα signaling cascades calmodulin and calcineurin which led to attenuation of NF-κB transcriptional promoter activity in EECs. MT depolymerization or CaMKIIα knockdown inhibited the transcription factor NFAT and NF-κB expression along with reduced secretion of prostaglandins PGE2 and PGF2α in mouse EECs. Overall, MT depolymerization impaired the WNT4/CaMKIIα signaling and suppressed the secretion of PGE2 and PGF2α in EECs which may be responsible for implantation failure in mice.
Subject(s)
Calcium-Calmodulin-Dependent Protein Kinase Type 2/metabolism , Embryo Implantation , Embryonic Development , Endometrium/pathology , Microtubules/pathology , Uterus/pathology , Wnt4 Protein/metabolism , Animals , Calcium-Calmodulin-Dependent Protein Kinase Type 2/genetics , Endometrium/drug effects , Endometrium/metabolism , Epithelial Cells/drug effects , Epithelial Cells/metabolism , Epithelial Cells/pathology , Female , Mice , Mice, Inbred BALB C , Microtubules/drug effects , Microtubules/metabolism , NF-kappa B/genetics , NF-kappa B/metabolism , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Nocodazole/pharmacology , Pregnancy , Signal Transduction , Tubulin Modulators/pharmacology , Uterus/drug effects , Uterus/metabolism , Wnt4 Protein/geneticsABSTRACT
Poplar (Populus spp.) is a tree species considered for the remediation of soil contaminated by metals, including zinc (Zn). To improve poplar's capacity for Zn assimilation and compartmentalization, it is necessary to understand the physiological and biochemical mechanisms that enable these features as well as their regulation at the molecular level. We observed that the molecular response of poplar roots to Zn excess overlapped with that activated by hypoxia. Therefore, we tested the effect of Zn excess on hypoxia-sensing components and investigated the consequence of root hypoxia on poplar fitness and Zn accumulation capacity. Our results suggest that high intracellular Zn concentrations mimic iron deficiency and inhibit the activity of the oxygen sensors Plant Cysteine Oxidases, leading to the stabilization and activation of ERF-VII transcription factors, which are key regulators of the molecular response to hypoxia. Remarkably, excess Zn and waterlogging similarly decreased poplar growth and development. Simultaneous excess Zn and waterlogging did not exacerbate these parameters, although Zn uptake was limited. This study unveils the contribution of the oxygen-sensing machinery to the Zn excess response in poplar, which may be exploited to improve Zn tolerance and increase Zn accumulation capacity in plants.