Transcriptomic analysis of the ion channelome of human platelets and megakaryocytic cell lines

Joy R. Wright; Stefan Amisten; Alison H. Goodall; Martyn P. Mahaut-Smith

doi:10.1160/TH15-11-0891

Transcriptomic analysis of the ion channelome of human platelets and megakaryocytic cell lines

Joy R. Wright, Stefan Amisten, Alison H. Goodall, Martyn P. Mahaut-Smith^*

^*Corresponding author for this work

Diabetes

University of Leicester

Research output: Contribution to journal › Article › peer-review

30 Citations (Scopus)

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Abstract

Ion channels have crucial roles in all cell types and represent important therapeutic targets. Approximately 20 ion channels have been reported in human platelets; however, no systematic study has been undertaken to define the platelet channelome. These membrane proteins need only be expressed at low copy number to influence function and may not be detected using proteomic or transcriptomic microar-ray approaches. In our recent work, quantitative real-time PCR (qPCR) provided key evidence that Kv1.3 is responsible for the voltage-dependent K⁺ conductance of platelets and megakaryocytes. The present study has expanded this approach to assess relative expression of 402 ion channels and channel regulatory genes in human platelets and three megakaryoblastic/erythroleukaemic cell lines. mRNA levels in platelets are low compared to other blood cells, therefore an improved method of isolating platelets was developed. This used a cocktail of inhibitors to prevent formation of leukocyte-platelet aggregates, and a combination of positive and negative immunomagnetic cell separation, followed by rapid extraction of mRNA. Expression of 34 channel-related transcripts was quantified in platelets, including 24 with unknown roles in platelet function, but that were detected at levels comparable to ion channels with established roles in haemostasis or thrombosis. Trace expression of a further 50 ion channel genes was also detected. More extensive channelomes were detected in MEG-01, CHRF-288-11 and HEL cells (195, 185 and 197 transcripts, respectively), but lacked several channels observed in the platelet. These “channelome” datasets provide an important resource for further studies of ion channel function in the platelet and megakaryo-cyte.

Original language	English
Pages (from-to)	272-284
Number of pages	13
Journal	Thrombosis and Haemostasis
Volume	116
Issue number	2
Early online date	9 Jun 2016
DOIs	https://doi.org/10.1160/TH15-11-0891
Publication status	Published - 1 Aug 2016

Keywords

Megakaryocytes
Molecular biology methods
Platelet physiology
Receptors

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1160/TH15-11-0891

Transcriptomic analysis of the ion_WRIGHT_Accepted30April2016_GOLD VoRFinal published version, 474 KB

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abstract = "Ion channels have crucial roles in all cell types and represent important therapeutic targets. Approximately 20 ion channels have been reported in human platelets; however, no systematic study has been undertaken to define the platelet channelome. These membrane proteins need only be expressed at low copy number to influence function and may not be detected using proteomic or transcriptomic microar-ray approaches. In our recent work, quantitative real-time PCR (qPCR) provided key evidence that Kv1.3 is responsible for the voltage-dependent K+ conductance of platelets and megakaryocytes. The present study has expanded this approach to assess relative expression of 402 ion channels and channel regulatory genes in human platelets and three megakaryoblastic/erythroleukaemic cell lines. mRNA levels in platelets are low compared to other blood cells, therefore an improved method of isolating platelets was developed. This used a cocktail of inhibitors to prevent formation of leukocyte-platelet aggregates, and a combination of positive and negative immunomagnetic cell separation, followed by rapid extraction of mRNA. Expression of 34 channel-related transcripts was quantified in platelets, including 24 with unknown roles in platelet function, but that were detected at levels comparable to ion channels with established roles in haemostasis or thrombosis. Trace expression of a further 50 ion channel genes was also detected. More extensive channelomes were detected in MEG-01, CHRF-288-11 and HEL cells (195, 185 and 197 transcripts, respectively), but lacked several channels observed in the platelet. These “channelome” datasets provide an important resource for further studies of ion channel function in the platelet and megakaryo-cyte.",

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Transcriptomic analysis of the ion channelome of human platelets and megakaryocytic cell lines

Abstract

Keywords

UN SDGs

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