Generation of an ABCG2GFPn-puro transgenic line – A tool to study ABCG2 expression in mice

Orford, Michael; Mean, Richard; Lapathitis, George; Genethliou, Nicholas; Panayiotou, Elena; Panayi, Helen; Malas, Stavros

doi:10.1016/j.bbrc.2009.04.089

Cited by 11 publications

(13 citation statements)

References 30 publications

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“…Very recently, transgenic mice expressing GFPn under the control of the ABCG2 promoter have been used to monitor the transcription pattern of the gene. 39 The authors found that in the early embryonic forebrain, ABCG2 localizes to ventral ventricular zone progenitors, consistent with the fact that this region harbors cells with stem-like properties. Dissociated neural progenitors revealed the expression of both ABCG2 and the stem-cell marker CD133.…”

Section: Cscs In Solid Tumors Location and Cell Or Originmentioning

confidence: 63%

The ABCG2 resistance network of glioblastoma

Bleau¹,

Huse²,

Holland³

2009

Cell Cycle

120

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Section: Cscs In Solid Tumors Location and Cell Or Originmentioning

confidence: 63%

The ABCG2 resistance network of glioblastoma

Bleau¹,

Huse²,

Holland³

2009

Cell Cycle

120

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“…BCRP removes a wide range of substances including chemotherapeutic agents, antiviral drugs, and carcinogens from the brain into the blood [56]. It is expressed on the apical membrane of the cerebral endothelial cells from early developmental ages and in rodents has been shown to be expressed from embryonic day 12.5 -approximately week 22 of human gestation [42,44,57,58]. BCRP expression remains largely unchanged across development, with a moderate increase in mRNA found between postnatal ages and adult in the mouse and rat [33,34].…”

Section: Accepted Manuscriptmentioning

confidence: 99%

Review: The blood-brain barrier; protecting the developing fetal brain

et al. 2017

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While placental function is fundamental to normal fetal development, the blood-brain barrier provides a second checkpoint critical to protecting the fetal brain and ensuring healthy brain development. The placenta is considered the key barrier between the mother and fetus, regulating delivery of essential nutrients, removing waste as well as protecting the fetus from potentially noxious substances. However, disturbances to the maternal environment and subsequent adaptations to placental function may render the placenta ineffective for providing a suitable environment for the developing fetus and to providing sufficient protection from harmful substances. The developing brain is particularly vulnerable to changes in the maternal/fetal environment. Development of the blood-brain barrier and maturation of barrier transporter systems work to protect the fetal brain from exposure to drugs, excluding them from the fetal CNS. This review will focus on the role of the 'other' key barrier during gestation - the blood-brain barrier - which has been shown to be functional as early as 8 weeks' gestation.

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“…24-26 Fluorescent proteins fused with membrane transporters have been used to determine the locations and topologies of ABC transporters. 27-28 Our previous studies have showed that fusion of BmrA (ABC transporter) with EGFP retains its efflux function.…”

Section: Introductionmentioning

confidence: 99%

Design and study of the efflux function of the EGFP fused MexAB-OprM membrane transporter in Pseudomonas aeruginosa using fluorescence spectroscopy

Ding

Lee

Vahedi-Faridi

et al. 2014

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Multidrug membrane transporters (efflux pumps) can selectively extrude a variety of structurally and functionally diverse substrates (e.g., chemotoxics, antibiotics), leading to multidrug resistance (MDR) and ineffective treatment of a wide variety of diseases. In this study, we have designed and constructed fusion gene (egfp-mexB) of N-terminal mexB with C-terminal egfp, inserted it into a plasmid vector (pMMB67EH), and successfully expressed it in ΔMexB (MexB deletion) strain of Pseudomonas aeruginosa to create a new strain that expresses MexA-(EGFP-MexB)-OprM. We characterized the fusion gene using gel electrophoresis and DNA sequencing, and determined their expression in live cells by measuring the fluorescence of EGFP in single live cells using fluorescence microscopy. Efflux function of the new strain was studied by measuring its accumulation kinetics of ethidium bromide (EtBr, a pump substrate) using fluorescence spectroscopy, which was compared with the cells (WT, ΔMexM, ΔABM, and nalB1) with various expression levels of MexAB-OprM. The new strain shows 6-fold lower accumulation rates of EtBr (15 μM) than ΔABM, 4-fold lower than ΔMexB, but only 1.1-fold higher than WT. As EtBr concentration increases to 40 μM, the new strain has nearly the same accumulation rate of EtBr as ΔMexB, but 1.4-fold higher than WT. We observed the nearly same level of inhibitory effect of CCCP (carbonyl cyanide-m-chlorophenylhydrazone) on the efflux of EtBr by the new strain and WT. Antibiotic susceptibility study shows that the minimum inhibitory concentrations (MICs) of aztreonam (AZT) and chloramphenicol (CP) for the new strain are 6-fold or 3-fold lower than WT, respectively, and 2-fold higher than those of ΔMexB. Taken together, the results suggest that the fusion protein partially retains the efflux function of MexAB-OprM. Modeled structure of the fusion protein shows that the position and orientation of the N-terminal fused EGFP domain may either partially block the translocation pore or restrict the movement of the individual pump domains, which leads to partially restrict efflux activity.

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Generation of an ABCG2GFPn-puro transgenic line – A tool to study ABCG2 expression in mice

Cited by 11 publications

References 30 publications

The ABCG2 resistance network of glioblastoma

The ABCG2 resistance network of glioblastoma

Review: The blood-brain barrier; protecting the developing fetal brain

Design and study of the efflux function of the EGFP fused MexAB-OprM membrane transporter in Pseudomonas aeruginosa using fluorescence spectroscopy

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