Intracellular Anions as the Voltage Sensor of Prestin, the Outer Hair Cell Motor Protein

Oliver, Dominik; He, David Z.Z.; Klöcker, Nikolaj; Ludwig, Jost; Schulte, Uwe; Waldegger, Siegfried; Ruppersberg, J. P.; Dallos, Peter; Fakler, Bernd

doi:10.1126/science.1060939

Cited by 412 publications

(564 citation statements)

References 26 publications

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“…Two membrane proteins, i.e., a fructose transporter, GLUT-5 (Géléoc et al 1999), and an anion/ sulfate transporter, prestin (Zheng et al 2000), were proposed to be this motor. Although recent studies have ascertained that the motor protein is prestin, not GLUT-5 (Belyantseva et al 2000;Oliver et al 2001;Liberman et al 2002), constituents of the 10-nm particles observed in the OHC lateral membrane have remained unclear; in other words, it is unknown whether these 10-nm particles are indeed exclusively constituted of prestin molecules or rather a complex of GLUT-5 and prestin molecules. In the present study, we found particle-like structures with a diameter of 8-12 nm in the prestin-transfected CHO plasma membrane.…”

Section: Imaging Of Membrane Protein Prestinmentioning

confidence: 99%

“…In an early study, this motor protein was proposed to be GLUT-5 (Géléoc et al 1999). Although GLUT-5 has been ruled out as being this motor protein (Belyantseva et al 2000;Oliver et al 2001;Liberman et al 2002), it is unclear whether prestin and GLUT-5 form a complex in the plasma membrane. Therefore, it still remains unknown whether the 10-nm particles seen in EM/AFM images are only prestin or rather a complex of GLUT-5 and prestin molecules.…”

Section: Introductionmentioning

confidence: 99%

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Imaging by Atomic Force Microscopy of the Plasma Membrane of Prestin-Transfected Chinese Hamster Ovary Cells

Murakoshi

Gomi

Iida

et al. 2006

JARO

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The high sensitivity of mammalian hearing is achieved by amplification of the motion of the cochlear partition. This cochlear amplification is thought to be generated by the elongation and contraction of outer hair cells (OHCs) in response to acoustical stimulation. This motility is made possible by a membrane protein embedded in the lateral membrane of OHCs. Although a fructose transporter, GLUT-5, was initially proposed to be this protein, a later study identified the gene of the motor protein distributed throughout the OHC plasma membrane. This protein has been named Bprestin.^However, although previous morphological studies by electron microscopy and atomic force microscopy (AFM) found the lateral wall of OHCs to be covered with 10-nm particles, believed to be motor proteins, it is unknown whether such particles consist only of prestin or are a complex of GLUT-5 and prestin molecules. To determine if the 10-nm particles are indeed constituted only of prestin, plasma membranes of prestin-transfected and untransfected Chinese hamster ovary (CHO) cells, which do not express GLUT-5, were observed by AFM. First, the cells attached to a substrate were sonicated so that only the plasma membrane remained on the substrate. The cytoplasmic face of the cell was observed by the tapping mode of the AFM in liquid. As a result, particle-like structures were recognized on the plasma membranes of both the prestintransfected and untransfected CHO cells. Comparison of the difference in the frequency distribution of these structures between those two cells showed approximately 75% of the particle-like structures with a diameter of 8-12 nm in the prestin-transfected CHO cells to be possibly constituted only by prestin molecules. Our data suggest that the densely packed 10-nm particles observed on the OHC lateral wall are likely to be constituted only of prestin molecules.

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Section: Imaging Of Membrane Protein Prestinmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Imaging by Atomic Force Microscopy of the Plasma Membrane of Prestin-Transfected Chinese Hamster Ovary Cells

Murakoshi

Gomi

Iida

et al. 2006

JARO

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“…The lateral region lacks voltage-gated channels but holds thousands of integral membrane proteins ( 7000/lm 2 ) (Gulley and Reese 1977;Forge 1991;Kalinec et al 1992;Huang and Santos±Sacchi 1993;Kalinec and Kachar 1995;Souter et al 1995;Holley 1996;Santos±Sacchi et al 1997). These integral membrane proteins are currently associated with the motor molecules responsible for the reversible and very fast changes in cell length known as OHC electromotility (Geleoc et al 1999;Zheng et al 2000;Belyantseva et al 2000;Oliver et al 2001). The basal region, in contrast, displays a signi®cant number of voltage-gated channels but has few motor molecules Santos±Sacchi et al 1997).…”

Section: Introductionmentioning

confidence: 99%

Structural Microdomains in the Lateral Plasma Membrane of Cochlear Outer Hair Cells

Zhang

Kalinec

2002

JARO

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The basal and lateral regions of the plasma membrane of cochlear outer hair cells are structurally and functionally distinct. The lateral region contains thousands of motor proteins but few voltage-gated channels. The basal region, conversely, contains a high number of voltage-gated channels but is devoid of motor proteins. It has been suggested that the cortical cytoskeleton is responsible for maintaining this regional distinction. Toward elucidating the structure of the outer hair cell's electromotile mechanism, we investigated the physical organization of the lateral plasma membrane in living guinea pig outer hair cells by analyzing the distribution pattern of the anionic long-chain carbocyanine SP-DiIC 18 (3) within this area, before and after electrical stimulation and with an intact and a disrupted cytoskeleton. We observed punctate, intensely¯uorescent patches as well as areas of weaker¯uorescence, with clear local maxima and minima, upon labeling the cells with this membrane-soluble probe. This discrete distribution of SP-DilC 18 (3) suggests that the lateral plasma membrane of guinea pig outer hair cells may be composed of small structural domains (microdomains). Disrupting the cytoskeleton with either trypsin or toxin B from Clostridium dif®cile did not change this pattern of distribution, thus indicating that this treatment did not facilitate the lateral diffusion of the probes. Electrical stimulation using whole-cell patch-clamp techniques, on the other hand, induced two responses: fast motility and reversible displacement of the¯uo-rescent probes. Both responses were inhibited by internal perfusion with salicylate, while disruption of the cytoskeleton did not inhibit OHC fast motility but affected the electrically induced redistribution of¯uo-rescent probes. Together, these results suggest that the lateral plasma membrane of guinea pig outer hair cells contains structural microdomains and that the cytoskeleton does not appear to be playing a major role in maintaining the lateral separation of these distinct molecular regions.

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“…Sulfate reduces somatic electromotility [6] acting directly on prestin [10]. Replacement of intracellular chloride with sulfate, in our experiment, did not appear to influence the efficacy of the extracellular applied 9AC.…”

Section: Discussionmentioning

confidence: 61%

Dissection of the Mechanical Impedance Components of the Outer Hair Cell Using a Chloride-Channel Blocker

Harasztosi¹,

Gummer²,

Shera³

et al. 2011

AIP Conference Proceedings

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Abstract. The voltage-dependent chloride-channel blocker anthracene-9-carboxylic acid (9AC) has been found to reduce the imaginary but not the real part of the mechanical impedance of the organ of Corti, suggesting that the effective stiffness of outer hair cells (OHCs) is reduced by 9AC. To examine whether 9AC interacts directly with the motor protein prestin to reduce the membrane component of the impedance, the patch-clamp technique in whole-cell configuration was used to measure the nonlinear capacitance (NLC) of isolated OHCs and, as control, prestin-transfected human embryonic kidney 293 (HEK293) cells. Extracellular application of 9AC significantly reduced the NLC of both OHCs and HEK293 cells. Intracellular 9AC did not influence the blocking effect of the extracellular applied drug. These results suggest that 9AC interacts directly with prestin, reducing the effective stiffness of the motor, and that the interaction is extracellular.

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Intracellular Anions as the Voltage Sensor of Prestin, the Outer Hair Cell Motor Protein

Cited by 412 publications

References 26 publications

Imaging by Atomic Force Microscopy of the Plasma Membrane of Prestin-Transfected Chinese Hamster Ovary Cells

Imaging by Atomic Force Microscopy of the Plasma Membrane of Prestin-Transfected Chinese Hamster Ovary Cells

Structural Microdomains in the Lateral Plasma Membrane of Cochlear Outer Hair Cells

Dissection of the Mechanical Impedance Components of the Outer Hair Cell Using a Chloride-Channel Blocker

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