QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

Vol. 16, Issue 2, 997-1010, February 2005

This Article Full Text Full Text (PDF) Supplemental Material All Versions of this Article: E04-10-0895v1 16/2/997    most recent Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Coene, E. D. Articles by Vaux, D. J. Search for Related Content PubMed PubMed Citation Articles by Coene, E. D. Articles by Vaux, D. J.

Phosphorylated BRCA1 Is Predominantly Located in the Nucleus and Mitochondria

Elisabeth D. Coene ^*, Michael S. Hollinshead  , Anouk A.T. Waeytens ^*, Vera R.J. Schelfhout ^*, Willy P. Eechaute , Michael K. Shaw , Patrick M.V. Van Oostveldt ^||, and David J. Vaux  ^¶

^* N. Goormaghtigh Institute of Pathology, University Hospital, B-9000 Gent, Belgium; Sir William Dunn School of Pathology, University of Oxford, Oxford OX1 3RE, United Kingdom; ^|| Laboratory of Biochemistry and Molecular Cytology, B-9000 Gent, Belgium; and Department of Physiology, University Hospital, B-9000 Gent, Belgium

Submitted October 14, 2004; Accepted November 24, 2004
Monitoring Editor: Thomas Fox

Multiple copies of the mitochondrial genome in eukaryotic cells are organized into protein–DNA complexes called nucleoids. Mitochondrial genome repair mechanisms have been reported, but they are less well characterized than their nuclear counterparts. To expand our knowledge of mitochondrial genome maintenance, we have studied the localization of the BRCA1 protein, known to be involved in nuclear repair pathways. Our confocal and immunoelectron microscopy results show that BRCA1 is present in mitochondria of several human cancer cell lines and in primary breast and nasal epithelial cells. BRCA1 localization in mitochondria frequently overlapped that of nucleoids. Small interfering RNA-mediated knockdown of BRCA1 in human cancer cells (confirmed by Western blot) results in decreased nuclear, cytoplasmic, and mitochondrial staining after immunofluorescence microscopy, establishing the specificity of the BRCA1 immunolabeling. Furthermore, using cell fractionation, dephosphorylation, and enzyme protection experiments, we show that a 220-kDa phosphorylated isoform of BRCA1 is enriched in mitochondrial and nuclear fractions but reduced in cytoplasmic subcellular fractions. Submitochondrial fractionation confirmed the presence of BRCA1 protein in isolated mitoplasts. Because phosphorylation of BRCA1 and subsequent changes in subcellular localization are known to follow DNA damage, our data support a universal role for BRCA1 in the maintenance of genome integrity in both mitochondria and nucleus.

Abbreviations used: CCO, cytochrome c oxidase; CF, cytoplasmic fraction; CKBS, cytokeratin broad spectrum; DAPI, 4,6-diamidino-2-phenylindole; EM, electron microscopy; HRP, horseradish peroxidase; MitF, mitochondrial fraction; MLDH, malate dehydrogenase; MP, mitoplasts; mtDNA, mitochondrial DNA; NF, nuclear fraction; PFA, paraformaldehyde; POD, peroxidase; siRNA, small interfering RNA; TX100, Triton X 100; WH, whole homogenate.

The online version of this article contains supplemental material at MBC Online (http://www.molbiolcell.org).

Present address: Henry Wellcome Imaging Centre, Wright-Fleming Institute, Imperial College Faculty of Medicine, Norfolk Place, London W2 1PG, United Kingdom.

^¶ Corresponding author. E-mail address: david.vaux{at}path.ox.ac.uk.

This article has been cited by other articles:

				V. Tembe and B. R. Henderson BARD1 Translocation to Mitochondria Correlates with Bax Oligomerization, Loss of Mitochondrial Membrane Potential, and Apoptosis J. Biol. Chem., July 13, 2007; 282(28): 20513 - 20522. [Abstract] [Full Text] [PDF]