Molecular and functional characterization of Hv1 proton channel in human granulocytes.

Voltage-gated proton current (I(Hv)) has been characterized in several cell types, but the majority of the data was collected in phagocytes, especially in human granulocytes. The prevailing view about the role of I(Hv) in phagocytes is that it is an essential supporter of the intense and sustained a...

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Autores principales:	Gábor L Petheo, Anna Orient, Mónika Baráth, István Kovács, Bence Réthi, Arpád Lányi, Anikó Rajki, Eva Rajnavölgyi, Miklós Geiszt
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2010
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/45a1796c13ca4fecb5e1ff42e1f18daf
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spelling	oai:doaj.org-article:45a1796c13ca4fecb5e1ff42e1f18daf2021-12-02T20:05:49ZMolecular and functional characterization of Hv1 proton channel in human granulocytes.1932-620310.1371/journal.pone.0014081https://doaj.org/article/45a1796c13ca4fecb5e1ff42e1f18daf2010-11-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/21124855/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203Voltage-gated proton current (I(Hv)) has been characterized in several cell types, but the majority of the data was collected in phagocytes, especially in human granulocytes. The prevailing view about the role of I(Hv) in phagocytes is that it is an essential supporter of the intense and sustained activity of Nox2 (the core enzyme of the phagocyte NADPH oxidase complex) during respiratory burst. Recently H(v)1, a voltage-gated proton channel, was cloned, and leukocytes from H(v)1 knockout mice display impaired respiratory burst. On the other hand, hardly anything is known about H(v)1 in human granulocytes. Using qPCR and a self made antibody, we detected a significant amount of H(v)1 in human eosinophil and neutrophil granulocytes and in PLB-985 leukemia cells. Using different crosslinking agents and detergents in reducing and non-reducing PAGE, significant expression of H(v)1 homodimers, but not that of higher-order multimers, could be detected in granulocytes. Results of subcellular fractionation and confocal imaging indicate that H(v)1 is resident in both plasmalemmal and granular membrane compartments of resting neutrophils. Furthermore, it is also demonstrated that H(v)1 accumulates in phagosome wall during zymosan engulfment together with, but independently of Nox2. During granulocytic differentiation early and parallel upregulation of H(v)1 and Nox2 expression was observed in PLB-985 cells. The upregulation of H(v)1 or Nox2 expression did not require the normal expression of the other molecule. Using RNA interference, we obtained strong correlation between H(v)1 expression and I(Hv) density in PLB-985 cells. It is also demonstrated that a massive reduction in H(v)1 expression can limit the Nox2 mediated superoxide production of PLB-985 granulocytes. In summary, beside monomers native H(v)1 forms stable proton channel dimer in resting and activated human granulocytes. The expression pattern of H(v)1 in granulocytes is optimized to support intense NADPH oxidase activity.Gábor L PetheoAnna OrientMónika BaráthIstván KovácsBence RéthiArpád LányiAnikó RajkiEva RajnavölgyiMiklós GeisztPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 5, Iss 11, p e14081 (2010)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Gábor L Petheo Anna Orient Mónika Baráth István Kovács Bence Réthi Arpád Lányi Anikó Rajki Eva Rajnavölgyi Miklós Geiszt Molecular and functional characterization of Hv1 proton channel in human granulocytes.
description	Voltage-gated proton current (I(Hv)) has been characterized in several cell types, but the majority of the data was collected in phagocytes, especially in human granulocytes. The prevailing view about the role of I(Hv) in phagocytes is that it is an essential supporter of the intense and sustained activity of Nox2 (the core enzyme of the phagocyte NADPH oxidase complex) during respiratory burst. Recently H(v)1, a voltage-gated proton channel, was cloned, and leukocytes from H(v)1 knockout mice display impaired respiratory burst. On the other hand, hardly anything is known about H(v)1 in human granulocytes. Using qPCR and a self made antibody, we detected a significant amount of H(v)1 in human eosinophil and neutrophil granulocytes and in PLB-985 leukemia cells. Using different crosslinking agents and detergents in reducing and non-reducing PAGE, significant expression of H(v)1 homodimers, but not that of higher-order multimers, could be detected in granulocytes. Results of subcellular fractionation and confocal imaging indicate that H(v)1 is resident in both plasmalemmal and granular membrane compartments of resting neutrophils. Furthermore, it is also demonstrated that H(v)1 accumulates in phagosome wall during zymosan engulfment together with, but independently of Nox2. During granulocytic differentiation early and parallel upregulation of H(v)1 and Nox2 expression was observed in PLB-985 cells. The upregulation of H(v)1 or Nox2 expression did not require the normal expression of the other molecule. Using RNA interference, we obtained strong correlation between H(v)1 expression and I(Hv) density in PLB-985 cells. It is also demonstrated that a massive reduction in H(v)1 expression can limit the Nox2 mediated superoxide production of PLB-985 granulocytes. In summary, beside monomers native H(v)1 forms stable proton channel dimer in resting and activated human granulocytes. The expression pattern of H(v)1 in granulocytes is optimized to support intense NADPH oxidase activity.
format	article
author	Gábor L Petheo Anna Orient Mónika Baráth István Kovács Bence Réthi Arpád Lányi Anikó Rajki Eva Rajnavölgyi Miklós Geiszt
author_facet	Gábor L Petheo Anna Orient Mónika Baráth István Kovács Bence Réthi Arpád Lányi Anikó Rajki Eva Rajnavölgyi Miklós Geiszt
author_sort	Gábor L Petheo
title	Molecular and functional characterization of Hv1 proton channel in human granulocytes.
title_short	Molecular and functional characterization of Hv1 proton channel in human granulocytes.
title_full	Molecular and functional characterization of Hv1 proton channel in human granulocytes.
title_fullStr	Molecular and functional characterization of Hv1 proton channel in human granulocytes.
title_full_unstemmed	Molecular and functional characterization of Hv1 proton channel in human granulocytes.
title_sort	molecular and functional characterization of hv1 proton channel in human granulocytes.
publisher	Public Library of Science (PLoS)
publishDate	2010
url	https://doaj.org/article/45a1796c13ca4fecb5e1ff42e1f18daf
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Molecular and functional characterization of Hv1 proton channel in human granulocytes.

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