Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.555526
Title: Properties of mammalian P2X₇ receptors
Author: Zheng, Wenxuan
Awarding Body: University of Manchester
Current Institution: University of Manchester
Date of Award: 2012
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Abstract:
To establish comprehensive pharmacology of P2X₇ receptors, membrane current recording, intracellular calcium transient recording and ethidium bromide uptake were carried out to examine several selective (A-740003, A- 438079) and non-selective (suramin) P2X₇ antagonists across mammalian P2X₇ receptors (human, mouse and rat). These P2X₇ receptors demonstrated species-dependent sensitivities to antagonists. In each species, A-740003 revealed variant IC50 values with different assays, indicating the assay- dependent pharmacology of P2X₇ receptors. Conventionally, pharmacology can be used to define a native current but not in the case of the human breast cancer cell line, Hs578T. It is found that P2X₇ was expressed at both mRNA and protein level. The ATP-evoked currents recorded from Hs578T cells were P2X₇-like with distinctive electrophysiological features. But the pharmacology profile of the currents did not fit with P2X₇ receptor. Further experiments are needed to either include or exclude the existence of functional P2X₇ receptors in Hs578T. Transmembrane domain 2 (TM2) is known as the pore-forming region for P2X receptors. TM2 of P2X₇ receptor was investigated with cysteine substitution scanning. The predicted α-helix structure of the TM2 segment was in good agreement with the results from the substituted cysteine accessibility method (SCAM). Thr336, Ser339, Tyr343, Phe344 and Thr348 were found important for both channel dilation and aqueous pore formation. Ser339 was further studied. Various substitutions at Ser339 were explored. The results suggest that the polarity of the side chain at Ser339 is essential for the channel dilation. Furthermore, disulfide bond formation was identified between S339C in the trimeric receptor, implying that the side chains of Ser339 might turn very close to each other during the channel opening and dilation.
Supervisor: Verkhratsky, Alexej. Sponsor: Dorothy Hodgkin Postgraduate Awards ; Wellcome Trust
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.555526  DOI: Not available
Keywords: P2X7 receptor ; pharmacology ; cancer ; amino acid substitution
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