scholarly journals Role of AmpR in the High Expression of the Plasmid-Encoded AmpC β-Lactamase CFE-1

mSphere ◽  
2017 ◽  
Vol 2 (4) ◽  
Author(s):  
Ryuichi Nakano ◽  
Akiyo Nakano ◽  
Hisakazu Yano ◽  
Ryoichi Okamoto
Keyword(s):  
High Resistance ◽  
High Expression ◽  
Regulator Gene ◽  
High Level Expression ◽  
Expression Levels ◽  
High Level ◽  
Unique Plasmid

ABSTRACT CFE-1 is a unique plasmid-encoded AmpC β-lactamase with the regulator gene ampR. It imparts high resistance to most cephalosporins with constitutive high-level β-lactamase activity. CFE-1 is a unique plasmid-encoded AmpC β-lactamase with the regulator gene ampR. It imparts high resistance to most cephalosporins with constitutive high-level β-lactamase activity. Here, the β-lactamase activities and expression levels of ampC with or without ampR were investigated. Results suggested that the resistance of CFE-1 to cephalosporins is caused by a substitution in AmpR, in which the Asp at position 135 is modified to Ala to allow the constitutive high-level expression (derepression) of ampC.

scholarly journals Structural and functional analysis of a promoter of the human granulin/epithelin gene

1996 ◽  
Vol 319 (2) ◽  
pp. 441-447 ◽  
Author(s):  
Vijay BHANDARI ◽  
Rachael DANIEL ◽  
Pheng Siew LIM ◽  
Andrew BATEMAN
Keyword(s):  
Promoter Activity ◽  
Biologically Active ◽  
Gene Products ◽  
High Level Expression ◽  
Haematopoietic Cells ◽  
Molecular Masses ◽  
Chloramphenicol Acetyltransferase Gene ◽  
High Level

Granulins (grns) or epithelins (epis) are peptides with molecular masses of approx. 6 kDa that modulate the growth of cells. The precursor for the grns/epis, which might itself be biologically active, is a secreted glycoprotein containing multiple repeats of the grn/epi motif. Grn/epi mRNA occurs widely in vivo, particularly in tissues rich in epithelial and haematopoietic cells. To understand better the role of the gene products for grn/epi it is important to determine the patterns of grn/epi gene expression and how this is regulated. To assist in this we have obtained the 5´ sequence of the human grn/epi gene, and using chimaeras of the grn/epi -5´ sequence and the chloramphenicol acetyltransferase gene we have shown a strong promoter activity associated with the 5´ sequence of the human grn/epi gene. We have further delineated regions of the 5´ sequence that confer high-level expression on the chimaeric gene.

scholarly journals Role of high expression levels of STK39 in the growth, migration and invasion of non-small cell type lung cancer cells

Oncotarget ◽  
2016 ◽  
Vol 7 (38) ◽  
pp. 61366-61377 ◽  
Author(s):  
Zhao Li ◽  
Wenzhuo Zhu ◽  
Liwen Xiong ◽  
Xiaobo Yu ◽  
Xi Chen ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Cancer Cells ◽  
Small Cell ◽  
High Expression ◽  
Lung Cancer Cells ◽  
Migration And Invasion ◽  
Cell Type ◽  
Expression Levels

scholarly journals Development of a Novel Chloramphenicol Resistance Expression Plasmid Used for Genetic Complementation of a fliG Deletion Mutant in Treponema denticola

2004 ◽  
Vol 72 (9) ◽  
pp. 5493-5497 ◽  
Author(s):  
Linda L. Slivienski-Gebhardt ◽  
Jacques Izard ◽  
William A. Samsonoff ◽  
Ronald J. Limberger
Keyword(s):  
Treponema Pallidum ◽  
Treponema Denticola ◽  
High Level Expression ◽  
Expression Plasmid ◽  
Chloramphenicol Resistance ◽  
Expression Of Genes ◽  
Colony Spreading ◽  
High Level ◽  
Resistance Expression

ABSTRACT A new expression plasmid containing the fla operon promoter and a staphylococcal chloramphenicol resistance gene, was constructed to help assess the role of fliG in Treponema denticola motility. Deletion of fliG resulted in a nonmotile mutant with a markedly decreased number of flagellar filaments. Wild-type fliG genes from T. denticola and from Treponema pallidum were cloned into this expression plasmid. In both cases, the gene restored the ability of the mutant to gyrate its cell ends and enabled colony spreading in agarose. This shuttle plasmid enables high-level expression of genes in T. denticola and possesses an efficient selectable marker that provides a new tool for treponemal genetics.

scholarly journals Prognosis and functional role of EIF4G1 in lung squamous cell carcinoma

2021 ◽  
Author(s):  
Baoxin Bai ◽  
Lingwei Wang ◽  
Zhiyuan Huang ◽  
Zhiwen Zhang ◽  
Ying Lu ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Squamous Cell Carcinoma ◽  
Cell Carcinoma ◽  
Squamous Cell ◽  
Functional Role ◽  
Lung Squamous Cell Carcinoma ◽  
High Expression ◽  
Expression Levels

Abstract Purpose To study the functional role and prognosis of EIF4G1 in lung squamous cell carcinoma (LSCC). Methods The clinical relevance of EIF4G1 in LSCC was investigated following detection of the expression levels of EIF4G1 by immunohistochemical staining (IHC). The expression levels of EIF4G1, AKT2, p-AKT, mTOR, cyclin D1 and β-actin were detected by western blot analysis. The cell proliferation and colony formation assays were used to detect the cell proliferative ability; flow-cytometry was used to assess the cell cycle; an invasion assay was used to detect cell invasive ability and the real-time quantitative polymerase chain reaction (Q-PCR) assay was used to assess the expression levels of EIF4G1 and β-actin. The role of EIF4G1 was verified by xenograft models. These experimental methods were employed to assess the functional role of EIF4G1 in LSCC pathogenesis. Results EIF4G1 was overexpressed in LSCC tumor tissues (P < 0.05) compared with the corresponding expression noted in paired adjacent tissues and cells. The expression levels of EIF4G1 were dependent on age (P = 0.002) and clinical stage (I vs II vs III + IV) (P < 0.001). High expression of EIF4G1 (P = 0.008, HR = 2.277, 95% CI = 1.250–4.145) could be used to predict the overall survival of LSCC patients as determined by the Cox’s proportional hazard model. High expression of EIF4G1 exhibited a lower survival (LogRank = 7.167, P = 0.007) in LSCC. Downregulation of EIF4G1 significantly inhibited LSCC proliferation, invasion and cell cycle progression. Conclusion EIF4G1 promotes LSCC cell proliferation and may represent an indicator of prognosis in LSCC.

scholarly journals Neuropilin-2 Is Associated With Increased Hepatoblastoma Cell Viability and Motility

2021 ◽  
Vol 9 ◽  
Author(s):  
Katja Eloranta ◽  
Ruth Nousiainen ◽  
Stefano Cairo ◽  
Mikko P. Pakarinen ◽  
David B. Wilson ◽  
...  
Keyword(s):  
Cell Viability ◽  
Cell Lines ◽  
Migration And Invasion ◽  
High Level Expression ◽  
Liver Malignancy ◽  
Rna Analysis ◽  
Cytoskeleton Remodeling ◽  
High Level ◽  
Tumor Types

The neuropilins NRP1 and NRP2 are multifunctional glycoproteins that have been implicated in several cancer-related processes including cell survival, migration, and invasion in various tumor types. Here, we examine the role of neuropilins in hepatoblastoma (HB), the most common pediatric liver malignancy. Using a combination of immunohistochemistry, RNA analysis and western blotting, we observed high level expression of NRP1 and NRP2 in 19 of 20 HB specimens and in a majority of human HB cell lines (HUH6 and five cell lines established from patient-derived xenografts) studied but not in normal hepatocytes. Silencing of NRP2 expression in HUH6 and HB-282 HB cells resulted in decreased cell viability, impaired cytoskeleton remodeling, and reduced cell motility, suggesting that NRP2 contributes to the malignant phenotype. We propose that neuropilins warrant further investigation as biomarkers of HB and potential therapeutic targets.

scholarly journals A Dominant Interfering Bub1 Mutant Is Insufficient To Induce or Alter Thymic Tumorigenesis In Vivo, Even in a Sensitized Genetic Background

2005 ◽  
Vol 25 (17) ◽  
pp. 7796-7802 ◽  
Author(s):  
Dale O. Cowley ◽  
Ginger W. Muse ◽  
Terry Van Dyke
Keyword(s):  
Mitotic Spindle ◽  
Genetic Background ◽  
Cultured Cells ◽  
Spindle Checkpoint ◽  
Transgenic Animals ◽  
Dominant Negative ◽  
High Level Expression ◽  
High Level

ABSTRACT Aneuploidy is a common feature of human tumors, often correlating with poor prognosis. The mitotic spindle checkpoint is thought to play a major role in aneuploidy suppression. To investigate the role of the spindle checkpoint in tumor suppression in vivo, we developed transgenic mice in which thymocytes express a dominant interfering fragment of Bub1, a kinase regulator of the spindle checkpoint. We report that, despite high-level expression of dominant-negative Bub1 (Bub1DN), a protein known to inhibit spindle checkpoint activity in cultured cells, thymocytes show no evidence of spindle checkpoint impairment. Transgenic animals also failed to show an increased predisposition to spontaneous tumors. Moreover, the Bub1DN transgene failed to alter the timing or characteristics of thymic lymphoma development in p53 heterozygous or homozygous null backgrounds, indicating that the lack of tumorigenesis is not due to suppression by p53-dependent checkpoints. These results indicate that overexpression of a Bub1 N-terminal fragment is insufficient to impair the spindle checkpoint in vivo or to drive tumorigenesis in the highly susceptible murine thymocyte system, either alone or in combination with G1 checkpoint disruption.

An evaluation of the role of Src in renal cancer.

2011 ◽  
Vol 29 (7_suppl) ◽  
pp. 358-358
Author(s):  
T. Qayyum ◽  
P. A. McArdle ◽  
M. Seywright ◽  
D. C. Mcmillan ◽  
M. Aitchison ◽  
...  
Keyword(s):  
Multivariate Analysis ◽  
Renal Cancer ◽  
Family Members ◽  
High Expression ◽  
Membrane Expression ◽  
Cancer Specific Survival ◽  
Expression Levels ◽  
Nuclear Expression ◽  
Specific Inhibitors

358 Background: The aim of the current study was to assess the expression levels of c-Src, phosphorylated Src, dephosphorylated c-Src at 530 and the downstream marker Fak 861 in renal cancer. Methods: In all, 60 patients undergoing potentially curative nephrectomy for localised renal cancer were included. Imunohistochemical staining was utilised to assess expression of c-Src, dephosphorlated c-Src at 530, phosphorylated Src at the Y416 site, and Fak 861. Expression was assessed using the weighted histoscore method. Results: High membrane c-Src was associated with increased cancer specific survival (p=0.032). In addition, increased cancer-specific survival was associated with high levels of cytoplasm and membrane expression of c-Src (p=0.039). When assessed individually, membrane, cytoplasm and nuclear expression of phosphorylated Src was not significantly associated with cancer specific survival. However when membrane and cytoplasm expression was combined, high expression was associated with decreased cancer specific survival (p=0.001). Low cytoplasm Fak expression was associated with increased cancer specific survival (p=0.029). When expression of Fak was combined with phosphorylated Src, high expression was associated with decreased cancer specific survival (p=0.003). When taking together high membrane c-Src and phosphorylated Src expression, this was associated with increased cancer specific survival (p=0.029). On multivariate analysis, combined expression of membrane c-Src and phosphorylated Src (HR 0.49, 95% CI 0.24-0.99, p=0.047) and combined Fak and phosphorylated Src expression (HR 1.88, 95% CI 1.06-3.34, p=0.030) were significant independent predictors of cancer specific survival. Conclusions: The results suggest that activated c-Src is associated with improved survival and activated Src family members are associated with decreased survival. This would suggest that another member of the Src family maybe associated with decreased survival. Further work is required to identify which of the Src family members are expressed in renal cancer and would therefore allow specific inhibitors for these Src family members to be developed. No significant financial relationships to disclose.

Expression patterns of auxin-responsive genes during tomato flower pedicel abscission and potential effects of calcium

2012 ◽  
Vol 60 (1) ◽  
pp. 68 ◽  
Author(s):  
Xianhong Zuo ◽  
Tao Xu ◽  
Mingfang Qi ◽  
Shuangshuang Lv ◽  
Jinhong Li ◽  
...  
Keyword(s):  
Expression Patterns ◽  
Negative Regulator ◽  
High Expression ◽  
Rna Expression ◽  
Conserved Domains ◽  
Expression Levels ◽  
Low Levels

This study aimed to determine the expression patterns of auxin (Aux/IAA)-responsive genes (ARG) during tomato flower pedicel abscission and the role of calcium in this auxin-mediated abscission. Most of the 19 proteins encoded by SlIAA genes showed the presence of all four conserved domains (I, II, III and IV). Expressions of some SlIAA genes decreased significantly (SlIAA 1, 3, 5, 8, 9, 10, 16, 17 and 27), while others increased (SlIAA 2, 4, 6, 7, 11, 12, 13, 26 and 29) at 0.5 h after excision. Most SlIAA genes were significantly upregulated at 1 h (except 9 and 27) then decreased to relatively low levels until 4 h after excision (except 4, 5, 8, 12, 14, 26 and 29). The SIAA genes were analysed and screened based on their expression patterns during different abscission phases. SlIAA4, 6, 9, 12 and 27 had relatively high expression levels consistent with the abscission rate, indicating potential roles in mediating abscission. SlIAA2, 3, 4, 5, 7, 9, 12, 13, 14, 16, 17, 26, 27 and 29 may have been important in delaying abscission, while SlIAA1, 9 and 12 may have been required for the completion of ethylene-induced abscission. SlIAA4, 6, 7, 8, 14, 16, 17 and 29 were important in calcium-delayed abscission. Analysis of other ARG revealed that tomato GH3 may have acted as an effective negative regulator in IAA-induced delay in abscission, while small auxin-up RNA expression patterns indicated that it may be a marker of IAA level throughout the abscission process.

scholarly journals IMMU-35. COMPLEMENT SYSTEM AND GLIOBLASTOMA: AN INTRICATE RELATIONSHIP

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi126-vi126
Author(s):  
Sandeep Mittal ◽  
Shayak Sengupta ◽  
Sabbir Khan ◽  
Kain McGee ◽  
Kristin Alfaro-Munoz ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Complement System ◽  
Complement Activation ◽  
Receptor Expression ◽  
High Expression ◽  
Rt Pcr ◽  
Expression Levels ◽  
Human Gbm ◽  
The Complement System

Abstract The complement system is a vital part of the innate immune system which plays a critical role in immune surveillance and inflammatory processes. Malignant and host cells express various complement inhibitory proteins on their surface to protect against complement mediated cytotoxicity. Imbalanced complement activation triggers inflammation and alters the tumor microenvironment. Complement activation has been shown to induce proliferation and migration of breast, ovarian and lung cancer cells. At this time, the expression and functional role of the complement cascade in glioblastoma (GBM) remain elusive. Here, we investigated the role of complement proteins and their receptor expression in human primary glioma stem-like cells (GSCs) and human GBM tissues. Western blot data demonstrated a high level of expression of central complement components and their receptors in GSCs. RT-PCR data further confirmed the high expression of complement genes which was similar or higher to normal human astrocytes (HA), a cell with high baseline expression levels of complement genes. Flow cytometry analysis revealed that almost 95% of GSCs expressed the anaphylatoxin complement receptors C3aR and C5aR on their cell surfaces which was consistent with our immunohistochemistry analysis of freshly resected GBM tissues. Furthermore, anaphylatoxin C5a exposure increased the proliferation of a subgroup of GSCs while reduced in another subset. Interestingly, C5a exposure was found to increase the expression of various pro-inflammatory markers in GSCs with reduced proliferation. Fluorescence-activated cell sorting (FACS) analysis of freshly isolated human GBM tissue revealed predominant expression of C5aR on cancer cells (CD11b-/CD45- cells) rather than on immune cells. RT-PCR analysis also demonstrated high expression levels of complement genes with concomitant decrease in complement inhibitory genes in human GBM tissue. Evaluation of the differential role of the complement system in GSCs along with their role in in vivo glioma models is ongoing.

scholarly journals High-Level Expression of Palmitoylated MPP1 Recombinant Protein in Mammalian Cells

Membranes ◽  
2021 ◽  
Vol 11 (9) ◽  
pp. 715
Author(s):  
Agnieszka Chytła ◽  
Weronika Gajdzik-Nowak ◽  
Agnieszka Biernatowska ◽  
Aleksander F. Sikorski ◽  
Aleksander Czogalla
Keyword(s):  
Recombinant Proteins ◽  
Mammalian Cells ◽  
High Yield ◽  
High Level Expression ◽  
Lateral Membrane ◽  
Protein Membrane ◽  
High Level ◽  
Lateral Organization

Our recent studies have pointed to an important role of the MAGUK family member, MPP1, as a crucial molecule interacting with flotillins and involved in the lateral organization of the erythroid plasma membrane. The palmitoylation of MPP1 seems to be an important element in this process; however, studies on the direct effect of palmitoylation on protein–protein or protein–membrane interactions in vitro are still challenging due to the difficulties in obtaining functional post-translationally modified recombinant proteins and the lack of comprehensive protocols for the purification of palmitoylated proteins. In this work, we present an optimized approach for the high-yield overexpression and purification of palmitoylated recombinant MPP1 protein in mammalian HEK-293F cells. The presented approach facilitates further studies on the molecular mechanism of lateral membrane organization and the functional impact of the palmitoylation of MPP1, which could also be carried out for other palmitoylated proteins.

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