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  Two-color STED microscopy reveals different degrees of colocalization between hexokinase-I and the three human VDAC isoforms.

Neumann, D., Bueckers, J., Kastrup, L., Hell, S. W., & Jakobs, S. (2010). Two-color STED microscopy reveals different degrees of colocalization between hexokinase-I and the three human VDAC isoforms. PMC Biophysics, 3(1): 4. doi:10.1186/1757-5036-3-4.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0010-931C-9 Version Permalink: http://hdl.handle.net/11858/00-001M-0000-0027-F276-F
Genre: Journal Article

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 Creators:
Neumann, D.1, Author              
Bueckers, J.2, Author              
Kastrup, L.2, Author              
Hell, S. W.2, Author              
Jakobs, S.1, Author              
Affiliations:
1Research Group of Mitochondrial Structure and Dynamics, MPI for biophysical chemistry, Max Planck Society, ou_578566              
2Department of NanoBiophotonics, MPI for biophysical chemistry, Max Planck Society, ou_578627              

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 Abstract: The voltage-dependent anion channel (VDAC, also known as mitochondrial porin) is the major transport channel mediating the transport of metabolites, including ATP, across the mitochondrial outer membrane. Biochemical data demonstrate the binding of the cytosolic protein hexokinase-I to VDAC, facilitating the direct access of hexokinase-I to the transported ATP. In human cells, three hVDAC isoforms have been identified. However, little is known on the distribution of these isoforms within the outer membrane of mitochondria and to what extent they colocalize with hexokinase-I. In this study we show that whereas hVDAC1 and hVDAC2 are localized predominantly within the same distinct domains in the outer membrane, hVDAC3 is mostly uniformly distributed over the surface of the mitochondrion. We used two-color stimulated emission depletion (STED) microscopy enabling a lateral resolution of ~40 nm to determine the detailed sub-mitochondrial distribution of the three hVDAC isoforms and hexokinase-I. Individual hVDAC and hexokinase-I clusters could thus be resolved which were concealed in the confocal images. Quantitative colocalization analysis of two-color STED images demonstrates that within the attained resolution, hexokinase-I and hVDAC3 exhibit a higher degree of colocalization than hexokinase-I with either hVDAC1 or hVDAC2. Furthermore, a substantial fraction of the mitochondria-bound hexokinase-I pool does not colocalize with any of the three hVDAC isoforms, suggesting a more complex interplay of these proteins than previously anticipated. This study demonstrates that two-color STED microscopy in conjunction with quantitative colocalization analysis is a powerful tool to study the complex distribution of membrane proteins in organelles such as mitochondria.

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Language(s): eng - English
 Dates: 2010-03-05
 Publication Status: Published online
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 Table of Contents: -
 Rev. Method: Peer
 Identifiers: eDoc: 474769
DOI: 10.1186/1757-5036-3-4
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Title: PMC Biophysics
Source Genre: Journal
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Pages: 15 Volume / Issue: 3 (1) Sequence Number: 4 Start / End Page: - Identifier: -