scholarly journals The rice annexin gene OsAnn5 is a positive regulator of cold stress tolerance at the seedling stage

2020 ◽  
Author(s):  
chunxiu shen ◽  
Zhiqun Que ◽  
Qineng Lu ◽  
Tao Liu ◽  
Shengqiang Li ◽  
...  
Keyword(s):  
Cold Stress ◽  
Stress Tolerance ◽  
Stress Responses ◽  
Fluorescent Protein ◽  
Functional Characterization ◽  
Survival Rates ◽  
Seedling Stage ◽  
Positive Regulator ◽  
Annexin Gene ◽  
Green Fluorescent

Abstract Annexins exist widely in plants as multigene families and play critical roles in stress responses and a range of cellular processes. In this study, we report on the cloning and functional characterization of the rice annexin gene OsAnn5. We found that the expression of OsAnn5 was induced by cold stress treatment at the seedling stage of rice. GUS staining assay indicated that the expression of OsAnn5 was non tissue-specific and was detected in almost all rice tissues. Subcellular localization indicated that OsAnn5-GFP (green fluorescent protein) signals were found in the endoplasmic reticulum apparatus. Compared with wild type rice, overexpression of OsAnn5 significantly increased survival rates at the seedling stage under cold stress, while knocking out OsAnn5 using the CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR associated proteins) mediated genome editing resulted in sensitivity to cold treatments. These results indicate that OsAnn5 is a positive regulator of cold stress tolerance at the seedling stage.

scholarly journals RNAi-Mediated Knockdown of Imaginal Disc Growth Factors (IDGFs) Genes Causes Developmental Malformation and Mortality in Melon Fly, Zeugodacus cucurbitae

2021 ◽  
Vol 12 ◽  
Author(s):  
Shakil Ahmad ◽  
Momana Jamil ◽  
Muhammad Fahim ◽  
Shujing Zhang ◽  
Farman Ullah ◽  
...  
Keyword(s):  
Growth Factors ◽  
Imaginal Disc ◽  
Fluorescent Protein ◽  
Developmental Stages ◽  
Small Body ◽  
Functional Characterization ◽  
Oral Feeding ◽  
Developmental Defects ◽  
Melon Fly ◽  
Green Fluorescent

This study reports the first successful use of oral feeding dsRNA technique for functional characterization of imaginal disc growth factors (IDGFs) genes (IDGF1, IDGF3_1, IDGF4_0, IDGF4_1, and IDGF6) in melon fly Zeugodacus cucurbitae. Phylogenetic and domain analysis indicates that these genes had high similarity with other Tephritidae fruit flies homolog and contain only one conserved domain among these five genes, which is glyco-18 domain (glyco-hydro-18 domain). Gene expression analysis at different developmental stages revealed that these genes were expressed at larval, pupal, and adult stages. To understand their role in different developmental stages, larvae were fed dsRNA-corresponding to each of the five IDGFs, in an artificial diet. RNAi-mediated knockdown of IDGF1 shows no phenotypic effects but caused mortality (10.4%), while IDGF4_0 caused malformed pharate at the adult stage where insects failed to shed their old cuticle and remained attached with their body, highest mortality (49.2%) was recorded compared to dsRNA-green fluorescent protein (GFP) or DEPC. Silencing of IDGF3_1 and IDGF4_1 cause lethal phenotype in larvae, (17.2%) and (40%) mortality was indexed in Z. cucurbitae. IDGF6 was mainly expressed in pupae and adult stages, and its silencing caused a malformation in adult wings. The developmental defects such as malformation in wings, larval–larval lethality, pupal–adult malformation, and small body size show that IDGFs are key developmental genes in the melon fly. Our results provide a baseline for the melon fly management and understanding of IDGFs specific functions in Z. cucurbitae.

scholarly journals Functional Characterization of a Drought-Responsive Invertase Inhibitor from Maize (Zea mays L.)

2019 ◽  
Vol 20 (17) ◽  
pp. 4081 ◽  
Author(s):  
Lin Chen ◽  
Xiaohong Liu ◽  
Xiaojia Huang ◽  
Wei Luo ◽  
Yuming Long ◽  
...  
Keyword(s):  
Cell Wall ◽  
Fluorescent Protein ◽  
Functional Characterization ◽  
Protein Fusion ◽  
Drought Treatment ◽  
Vacuolar Invertase ◽  
Maize Leaves ◽  
Green Fluorescent ◽  
Proteinaceous Inhibitor

Invertases (INVs) play essential roles in plant growth in response to environmental cues. Previous work showed that plant invertases can be post-translationally regulated by small protein inhibitors (INVINHs). Here, this study characterizes a proteinaceous inhibitor of INVs in maize (Zm-INVINH4). A functional analysis of the recombinant Zm-INVINH4 protein revealed that it inhibited both cell wall and vacuolar invertase activities from maize leaves. A Zm-INVINH4::green fluorescent protein fusion experiment indicated that this protein localized in the apoplast. Transcript analysis showed that Zm-INVINH4 is specifically expressed in maize sink tissues, such as the base part of the leaves and young kernels. Moreover, drought stress perturbation significantly induced Zm-INVINH4 expression, which was accompanied with a decrease of cell wall invertase (CWI) activities and an increase of sucrose accumulation in both base parts of the leaves 2 to 7 days after pollinated kernels. In summary, the results support the hypothesis that INV-related sink growth in response to drought treatment is (partially) caused by a silencing of INV activity via drought-induced induction of Zm-INVINH4 protein.

Induced Expression of the Heat Shock Protein Genes uspA and grpE during Starvation at Low Temperatures and Their Influence on Thermal Resistance of Escherichia coli O157:H7

2003 ◽  
Vol 66 (11) ◽  
pp. 2045-2050 ◽  
Author(s):  
YI ZHANG ◽  
MANSEL W. GRIFFITHS
Keyword(s):  
Heat Shock ◽  
Heat Shock Proteins ◽  
Stress Responses ◽  
Thermal Tolerance ◽  
Molecular Mechanisms ◽  
Fluorescent Protein ◽  
Bacterial Cells ◽  
Induced Expression ◽  
Green Fluorescent ◽  
Shock Proteins

Heat shock proteins play an important role in protecting bacterial cells against several stresses, including starvation. In this study, the promoters for two genes encoding heat shock proteins involved in many stress responses, UspA and GrpE, were fused with the green fluorescent protein (gfp) gene. Thus, the expression of the two genes could be quantified by measuring the fluorescence emitted by the cells under different environmental conditions. The heat resistance levels of starved and nonstarved cells during storage at 5, 10, and 37°C were compared with the levels of expression of the uspA and grpE genes. D52-values (times required for decimal reductions in count at 52°C) increased by 11.5, 14.6, and 18.5 min when cells were starved for 3 h at 37°C, for 24 h at 10°C, and for 2 days at 5°C, respectively. In all cases, these increases were significant (P < 0.01), indicating that the stress imposed by starvation altered the ability of E. coli O157:H7 to survive subsequent heat treatments. Thermal tolerance was correlative with the induction of UspA and GrpE. At 5°C, the change in the thermal tolerance of the pathogen was positively linked to the induced expression of the grpE gene but negatively related to the expression of the uspA gene. The results obtained in this study indicate that UspA plays an important role in starvation-induced thermal tolerance at 37°C but that GrpE may be more involved in regulating this response at lower temperatures. An improvement in our understanding of the molecular mechanisms involved in these cross-protection responses may make it possible to devise strategies to limit their effects.

The Wheat Aquaporin Gene TaAQP7 Confers Tolerance to Cold Stress in Transgenic Tobacco

2014 ◽  
Vol 69 (3-4) ◽  
pp. 142-148 ◽  
Author(s):  
Chao Huang ◽  
Shiyi Zhou ◽  
Wei Hu ◽  
Xiaomin Deng ◽  
Shuya Wei ◽  
...  
Keyword(s):  
Cold Stress ◽  
Stress Tolerance ◽  
Transgenic Tobacco ◽  
Stress Responses ◽  
Cold Treatment ◽  
Membrane Damage ◽  
Pcr Analysis ◽  
Ion Leakage ◽  
Chlorophyll Contents ◽  
Aquaporin Gene

Aquaporin proteins (AQPs) have been shown to be involved in abiotic stress responses. However, the precise role of AQPs, especially in response to cold stress, is not understood in wheat (Triticum aestivum). In the present study, quantitative real time polymerase chain reaction (qRT-PCR) analysis revealed that TaAQP7 expression increased in leaves, but decreased in roots after cold treatment. Expression of TaAQP7 in tobacco plants resulted in increased root elongation and better growth compared with wild-type (WT) plants under cold stress. Moreover, after cold treatment, the transgenic tobacco lines exhibited higher chlorophyll contents, lower levels of malondialdehyde (MDA), and less ion leakage (IL) than WT plants. Thus, expression of TaAQP7 enhanced cold stress tolerance in transgenic tobacco. Taken together, our results suggest that TaAQP7 confers cold stress tolerance by relieving membrane damage in the transgenic plants.

scholarly journals Phagocytosed Bordetella pertussis Fails To Survive in Human Neutrophils

2000 ◽  
Vol 68 (2) ◽  
pp. 956-959 ◽  
Author(s):  
Derrick H. Lenz ◽  
Christine L. Weingart ◽  
Alison A. Weiss
Keyword(s):  
Green Fluorescent Protein ◽  
Fluorescence Microscopy ◽  
Bordetella Pertussis ◽  
Fluorescent Protein ◽  
Survival Rates ◽  
Polymyxin B ◽  
Immune Serum ◽  
Human Neutrophils ◽  
Green Fluorescent

ABSTRACT Previous studies have reported that phagocytosed Bordetella pertussis survives in human neutrophils. This issue has been reexamined. Opsonized or unopsonized bacteria expressing green fluorescent protein (GFP) were incubated with adherent human neutrophils. Phagocytosis was quantified by fluorescence microscopy, and the viability of phagocytosed bacteria was determined by colony counts following treatment with polymyxin B to kill extracellular bacteria. Only 1 to 2% of the phagocytosed bacteria remained viable. Opsonization with heat-inactivated immune serum reduced the amount of attachment and phagocytosis of the bacteria but did not alter survival rates. In contrast to previous reports, these data suggest that phagocytosed B. pertussis bacteria are killed by human neutrophils.

scholarly journals A Mutation in a Novel Yeast Proteasomal Gene,RPN11/MPR1, Produces a Cell Cycle Arrest, Overreplication of Nuclear and Mitochondrial DNA, and an Altered Mitochondrial Morphology

1998 ◽  
Vol 9 (10) ◽  
pp. 2917-2931 ◽  
Author(s):  
Teresa Rinaldi ◽  
Carlo Ricci ◽  
Danilo Porro ◽  
Monique Bolotin-Fukuhara ◽  
Laura Frontali
Keyword(s):  
Mitochondrial Dna ◽  
Fluorescent Protein ◽  
Functional Characterization ◽  
Mitochondrial Morphology ◽  
Nonpermissive Temperature ◽  
Protein Fusion ◽  
In The Wild ◽  
A Cell ◽  
Green Fluorescent ◽  
Cytoplasmic Structures

We report here the functional characterization of an essentialSaccharomyces cerevisiae gene, MPR1, coding for a regulatory proteasomal subunit for which the name Rpn11p has been proposed. For this study we made use of thempr1-1 mutation that causes the following pleiotropic defects. At 24°C growth is delayed on glucose and impaired on glycerol, whereas no growth is seen at 36°C on either carbon source. Microscopic observation of cells growing on glucose at 24°C shows that most of them bear a large bud, whereas mitochondrial morphology is profoundly altered. A shift to the nonpermissive temperature produces aberrant elongated cell morphologies, whereas the nucleus fails to divide. Flow cytometry profiles after the shift to the nonpermissive temperature indicate overreplication of both nuclear and mitochondrial DNA. Consistently with the identification of Mpr1p with a proteasomal subunit, the mutation is complemented by the human POH1proteasomal gene. Moreover, the mpr1-1 mutant grown to stationary phase accumulates ubiquitinated proteins. Localization of the Rpn11p/Mpr1p protein has been studied by green fluorescent protein fusion, and the fusion protein has been found to be mainly associated to cytoplasmic structures. For the first time, a proteasomal mutation has also revealed an associated mitochondrial phenotype. We actually showed, by the use of [rho°] cells derived from the mutant, that the increase in DNA content per cell is due in part to an increase in the amount of mitochondrial DNA. Moreover, microscopy of mpr1-1 cells grown on glucose showed that multiple punctate mitochondrial structures were present in place of the tubular network found in the wild-type strain. These data strongly suggest that mpr1-1 is a valuable tool with which to study the possible roles of proteasomal function in mitochondrial biogenesis.

scholarly journals Enhanced Survival of Salmonella enterica in Vesicles Released by a Soilborne Tetrahymena Species

2005 ◽  
Vol 71 (3) ◽  
pp. 1562-1569 ◽  
Author(s):  
M. T. Brandl ◽  
B. M. Rosenthal ◽  
A. F. Haxo ◽  
S. G. Berk
Keyword(s):  
Public Health ◽  
Salmonella Enterica ◽  
Fluorescent Protein ◽  
Survival Rates ◽  
Human Pathogen ◽  
Initial Ratio ◽  
Calcium Hypochlorite ◽  
Human Macrophages ◽  
Low Concentrations ◽  
Green Fluorescent

ABSTRACT Nondestructive ingestion by soilborne protozoa may enhance the environmental resiliency of important bacterial pathogens and may model how such bacteria evade destruction in human macrophages. Here, the interaction of Salmonella enterica serovar Thompson with a soilborne Tetrahymena sp. isolate was examined using serovar Thompson cells labeled with the green fluorescent protein. The bacteria were mixed in solution with cells of Tetrahymena at several ratios. During incubation with serovar Thompson, Tetrahymena cells released a large number of vesicles containing green fluorescent serovar Thompson cells. In comparison, grazing on Listeria monocytogenes cells resulted in their digestion and thus the infrequent release of this pathogen in vesicles. The number of serovar Thompson cells per vesicle increased significantly as the initial ratio of serovar Thompson to Tetrahymena cells increased from 500:1 to 5,000:1. The density of serovar Thompson was as high as 50 cells per vesicle. Staining with propidium iodide revealed that a significantly higher proportion of serovar Thompson cells remained viable when enclosed in vesicles than when free in solution. Enhanced survival rates were observed in vesicles that were secreted by both starved (F = 28.3, P < 0.001) and unstarved (F = 14.09, P < 0.005) Tetrahymena cells. Sequestration in vesicles also provided greater protection from low concentrations of calcium hypochlorite. Thus, the release of this human pathogen from Tetrahymena cells in high-density clusters enclosed in a membrane may have important implications for public health.

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