Na+/K+-ATPase α1 identified as an abundant protein in the blood-labyrinth barrier that plays an essential role in the barrier integrity

Yue Yang, Min Dai, Teresa M. Wilson, Irina Omelchenko, John E. Klimek, Phillip Wilmarth, Larry David, Alfred Nuttall, Peter Barr-Gillespie, Xiao Shi

Research output: Contribution to journalArticle

38 Citations (Scopus)

Abstract

Background: The endothelial-blood/tissue barrier is critical for maintaining tissue homeostasis. The ear harbors a unique endothelial-blood/tissue barrier which we term "blood-labyrinth-barrier". This barrier is critical for maintaining inner ear homeostasis. Disruption of the blood-labyrinth-barrier is closely associated with a number of hearing disorders. Many proteins of the blood-brain-barrier and blood-retinal-barrier have been identified, leading to significant advances in understanding their tissue specific functions. In contrast, capillaries in the ear are small in volume and anatomically complex. This presents a challenge for protein analysis studies, which has resulted in limited knowledge of the molecular and functional components of the blood-labyrinth-barrier. In this study, we developed a novel method for isolation of the stria vascularis capillary from CBA/CaJ mouse cochlea and provided the first database of protein components in the bloodlabyrinth barrier as well as evidence that the interaction of Na+/K+-ATPase α1 (ATP1A1) with protein kinase C eta (PKCη) and occludin is one of the mechanisms of loud sound-induced vascular permeability increase. Methodology/Principal Findings: Using a mass-spectrometry, shotgun-proteomics approach combined with a novel "sandwich-dissociation" method, more than 600 proteins from isolated stria vascularis capillaries were identified from adult CBA/CaJ mouse cochlea. The ion transporter ATP1A1 was the most abundant protein in the blood-labyrinth barrier. Pharmacological inhibition of ATP1A1 activity resulted in hyperphosphorylation of tight junction proteins such as occludin which increased the blood-labyrinth-barrier permeability. PKCη directly interacted with ATP1A1 and was an essential mediator of ATP1A1-initiated occludin phosphorylation. Moreover, this identified signaling pathway was involved in the breakdown of the blood-labyrinth-barrier resulting from loud sound trauma. Conclusions/Significance: The results presented here provide a novel method for capillary isolation from the inner ear and the first database on protein components in the blood-labyrinth-barrier. Additionally, we found that ATP1A1 interaction with PKCη and occludin was involved in the integrity of the blood-labyrinth-barrier.

Original languageEnglish (US)
Article numbere16547
JournalPLoS One
Volume6
Issue number1
DOIs
StatePublished - 2011

Fingerprint

sodium-potassium-exchanging ATPase
Inner Ear
blood proteins
Adenosine Triphosphatases
Blood Proteins
Blood
blood
Occludin
Proteins
ears
protein kinase C
Stria Vascularis
Inbred CBA Mouse
Protein Databases
proteins
Cochlea
Tissue
Endothelium
Ear
hearing disorders

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Medicine(all)

Cite this

Na+/K+-ATPase α1 identified as an abundant protein in the blood-labyrinth barrier that plays an essential role in the barrier integrity. / Yang, Yue; Dai, Min; Wilson, Teresa M.; Omelchenko, Irina; Klimek, John E.; Wilmarth, Phillip; David, Larry; Nuttall, Alfred; Barr-Gillespie, Peter; Shi, Xiao.

In: PLoS One, Vol. 6, No. 1, e16547, 2011.

Research output: Contribution to journalArticle

@article{2890ef797bd049c3822a95a8cd9e157e,
title = "Na+/K+-ATPase α1 identified as an abundant protein in the blood-labyrinth barrier that plays an essential role in the barrier integrity",
abstract = "Background: The endothelial-blood/tissue barrier is critical for maintaining tissue homeostasis. The ear harbors a unique endothelial-blood/tissue barrier which we term {"}blood-labyrinth-barrier{"}. This barrier is critical for maintaining inner ear homeostasis. Disruption of the blood-labyrinth-barrier is closely associated with a number of hearing disorders. Many proteins of the blood-brain-barrier and blood-retinal-barrier have been identified, leading to significant advances in understanding their tissue specific functions. In contrast, capillaries in the ear are small in volume and anatomically complex. This presents a challenge for protein analysis studies, which has resulted in limited knowledge of the molecular and functional components of the blood-labyrinth-barrier. In this study, we developed a novel method for isolation of the stria vascularis capillary from CBA/CaJ mouse cochlea and provided the first database of protein components in the bloodlabyrinth barrier as well as evidence that the interaction of Na+/K+-ATPase α1 (ATP1A1) with protein kinase C eta (PKCη) and occludin is one of the mechanisms of loud sound-induced vascular permeability increase. Methodology/Principal Findings: Using a mass-spectrometry, shotgun-proteomics approach combined with a novel {"}sandwich-dissociation{"} method, more than 600 proteins from isolated stria vascularis capillaries were identified from adult CBA/CaJ mouse cochlea. The ion transporter ATP1A1 was the most abundant protein in the blood-labyrinth barrier. Pharmacological inhibition of ATP1A1 activity resulted in hyperphosphorylation of tight junction proteins such as occludin which increased the blood-labyrinth-barrier permeability. PKCη directly interacted with ATP1A1 and was an essential mediator of ATP1A1-initiated occludin phosphorylation. Moreover, this identified signaling pathway was involved in the breakdown of the blood-labyrinth-barrier resulting from loud sound trauma. Conclusions/Significance: The results presented here provide a novel method for capillary isolation from the inner ear and the first database on protein components in the blood-labyrinth-barrier. Additionally, we found that ATP1A1 interaction with PKCη and occludin was involved in the integrity of the blood-labyrinth-barrier.",
author = "Yue Yang and Min Dai and Wilson, {Teresa M.} and Irina Omelchenko and Klimek, {John E.} and Phillip Wilmarth and Larry David and Alfred Nuttall and Peter Barr-Gillespie and Xiao Shi",
year = "2011",
doi = "10.1371/journal.pone.0016547",
language = "English (US)",
volume = "6",
journal = "PLoS One",
issn = "1932-6203",
publisher = "Public Library of Science",
number = "1",

}

TY - JOUR

T1 - Na+/K+-ATPase α1 identified as an abundant protein in the blood-labyrinth barrier that plays an essential role in the barrier integrity

AU - Yang, Yue

AU - Dai, Min

AU - Wilson, Teresa M.

AU - Omelchenko, Irina

AU - Klimek, John E.

AU - Wilmarth, Phillip

AU - David, Larry

AU - Nuttall, Alfred

AU - Barr-Gillespie, Peter

AU - Shi, Xiao

PY - 2011

Y1 - 2011

N2 - Background: The endothelial-blood/tissue barrier is critical for maintaining tissue homeostasis. The ear harbors a unique endothelial-blood/tissue barrier which we term "blood-labyrinth-barrier". This barrier is critical for maintaining inner ear homeostasis. Disruption of the blood-labyrinth-barrier is closely associated with a number of hearing disorders. Many proteins of the blood-brain-barrier and blood-retinal-barrier have been identified, leading to significant advances in understanding their tissue specific functions. In contrast, capillaries in the ear are small in volume and anatomically complex. This presents a challenge for protein analysis studies, which has resulted in limited knowledge of the molecular and functional components of the blood-labyrinth-barrier. In this study, we developed a novel method for isolation of the stria vascularis capillary from CBA/CaJ mouse cochlea and provided the first database of protein components in the bloodlabyrinth barrier as well as evidence that the interaction of Na+/K+-ATPase α1 (ATP1A1) with protein kinase C eta (PKCη) and occludin is one of the mechanisms of loud sound-induced vascular permeability increase. Methodology/Principal Findings: Using a mass-spectrometry, shotgun-proteomics approach combined with a novel "sandwich-dissociation" method, more than 600 proteins from isolated stria vascularis capillaries were identified from adult CBA/CaJ mouse cochlea. The ion transporter ATP1A1 was the most abundant protein in the blood-labyrinth barrier. Pharmacological inhibition of ATP1A1 activity resulted in hyperphosphorylation of tight junction proteins such as occludin which increased the blood-labyrinth-barrier permeability. PKCη directly interacted with ATP1A1 and was an essential mediator of ATP1A1-initiated occludin phosphorylation. Moreover, this identified signaling pathway was involved in the breakdown of the blood-labyrinth-barrier resulting from loud sound trauma. Conclusions/Significance: The results presented here provide a novel method for capillary isolation from the inner ear and the first database on protein components in the blood-labyrinth-barrier. Additionally, we found that ATP1A1 interaction with PKCη and occludin was involved in the integrity of the blood-labyrinth-barrier.

AB - Background: The endothelial-blood/tissue barrier is critical for maintaining tissue homeostasis. The ear harbors a unique endothelial-blood/tissue barrier which we term "blood-labyrinth-barrier". This barrier is critical for maintaining inner ear homeostasis. Disruption of the blood-labyrinth-barrier is closely associated with a number of hearing disorders. Many proteins of the blood-brain-barrier and blood-retinal-barrier have been identified, leading to significant advances in understanding their tissue specific functions. In contrast, capillaries in the ear are small in volume and anatomically complex. This presents a challenge for protein analysis studies, which has resulted in limited knowledge of the molecular and functional components of the blood-labyrinth-barrier. In this study, we developed a novel method for isolation of the stria vascularis capillary from CBA/CaJ mouse cochlea and provided the first database of protein components in the bloodlabyrinth barrier as well as evidence that the interaction of Na+/K+-ATPase α1 (ATP1A1) with protein kinase C eta (PKCη) and occludin is one of the mechanisms of loud sound-induced vascular permeability increase. Methodology/Principal Findings: Using a mass-spectrometry, shotgun-proteomics approach combined with a novel "sandwich-dissociation" method, more than 600 proteins from isolated stria vascularis capillaries were identified from adult CBA/CaJ mouse cochlea. The ion transporter ATP1A1 was the most abundant protein in the blood-labyrinth barrier. Pharmacological inhibition of ATP1A1 activity resulted in hyperphosphorylation of tight junction proteins such as occludin which increased the blood-labyrinth-barrier permeability. PKCη directly interacted with ATP1A1 and was an essential mediator of ATP1A1-initiated occludin phosphorylation. Moreover, this identified signaling pathway was involved in the breakdown of the blood-labyrinth-barrier resulting from loud sound trauma. Conclusions/Significance: The results presented here provide a novel method for capillary isolation from the inner ear and the first database on protein components in the blood-labyrinth-barrier. Additionally, we found that ATP1A1 interaction with PKCη and occludin was involved in the integrity of the blood-labyrinth-barrier.

UR - http://www.scopus.com/inward/record.url?scp=79551626442&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=79551626442&partnerID=8YFLogxK

U2 - 10.1371/journal.pone.0016547

DO - 10.1371/journal.pone.0016547

M3 - Article

C2 - 21304972

AN - SCOPUS:79551626442

VL - 6

JO - PLoS One

JF - PLoS One

SN - 1932-6203

IS - 1

M1 - e16547

ER -