| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
* Image Analysis Laboratory, National Cancer Institute, Frederick, Maryland; National Cancer Institute/Science Applications International Corporation, Frederick, Maryland; and Human Retrovirus Section, National Cancer Institute, Frederick, Maryland
Correspondence: Address reprint requests to Sylvain V. Costes, Tel.: 510-486-6988; Fax: 510-486-6746; E-mail: SVCostes{at}lbl.gov.
We introduce a novel statistical approach that quantifies, for the first time, the amount of colocalization of two fluorescent-labeled proteins in an image automatically, removing the bias of visual interpretation. This is done by estimating simultaneously the maximum threshold of intensity for each color below which pixels do not show any statistical correlation. The sensitivity of the method was illustrated on simulated data by statistically confirming the existence of true colocalization in images with as little as 3% colocalization. This method was then tested on a large three-dimensional set of fixed cells cotransfected with CFP/YFP pairs of proteins that either co-compartmentalized, interacted, or were just randomly localized in the nucleolus. In this test, the algorithm successfully distinguished random color overlap from colocalization due to either co-compartmentalization or interaction, and results were verified by fluorescence resonance energy transfer. The accuracy and consistency of our algorithm was further illustrated by measuring, for the first time in live cells, the dissociation rate (kd) of the HIV-1 Rev/CRM1 export complex induced by the cytotoxin leptomycin B. Rev/CRM1 colocalization in nucleoli dropped exponentially after addition of leptomycin B at a rate of 1.25 x 10–3 s–1. More generally, this algorithm can be used to answer a variety of biological questions involving protein-protein interactions or co-compartmentalization and can be generalized to colocalization of more than two colors.
This article has been cited by other articles:
 |
|
 |
|
  S. Bultel, L. Helin, V. Clavey, G. Chinetti-Gbaguidi, E. Rigamonti, M. Colin, J.-C. Fruchart, B. Staels, and S. Lestavel Liver X Receptor Activation Induces the Uptake of Cholesteryl Esters From High Density Lipoproteins in Primary Human Macrophages Arterioscler. Thromb. Vasc. Biol., December 1, 2008; 28(12): 2288 - 2295. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M.-H. Blanchet, J. A. Le Good, V. Oorschot, S. Baflast, G. Minchiotti, J. Klumperman, and D. B. Constam Cripto Localizes Nodal at the Limiting Membrane of Early Endosomes Sci. Signal., November 11, 2008; 1(45): ra13 - ra13. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. Pellistri, M. Bucciantini, A. Relini, D. Nosi, A. Gliozzi, M. Robello, and M. Stefani Nonspecific Interaction of Prefibrillar Amyloid Aggregates with Glutamatergic Receptors Results in Ca2+ Increase in Primary Neuronal Cells J. Biol. Chem., October 31, 2008; 283(44): 29950 - 29960. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Burdak-Rothkamm, K. Rothkamm, and K. M. Prise ATM Acts Downstream of ATR in the DNA Damage Response Signaling of Bystander Cells Cancer Res., September 1, 2008; 68(17): 7059 - 7065. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. J. Mulholland, E. P. Carpenter-Hyland, M. C. Hearing, H. C. Becker, J. J. Woodward, and L. J. Chandler Glutamate Transporters Regulate Extrasynaptic NMDA Receptor Modulation of Kv2.1 Potassium Channels J. Neurosci., August 27, 2008; 28(35): 8801 - 8809. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Carrel, J. Masson, S. Al Awabdh, C. B. Capra, Z. Lenkei, M. Hamon, M. B. Emerit, and M. Darmon Targeting of the 5-HT1A Serotonin Receptor to Neuronal Dendrites Is Mediated by Yif1B J. Neurosci., August 6, 2008; 28(32): 8063 - 8073. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. S. Nicolas, K.-H. Park, A. El Harchi, J. Camonis, R. S. Kass, D. Escande, J. Merot, G. Loussouarn, F. Le Bouffant, and I. Baro IKs response to protein kinase A-dependent KCNQ1 phosphorylation requires direct interaction with microtubules Cardiovasc Res, August 1, 2008; 79(3): 427 - 435. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Karjalainen, E. Kakkonen, P. Upla, H. Paloranta, P. Kankaanpaa, P. Liberali, G. H. Renkema, T. Hyypia, J. Heino, and V. Marjomaki A Raft-derived, Pak1-regulated Entry Participates in {alpha}2{beta}1 Integrin-dependent Sorting to Caveosomes Mol. Biol. Cell, July 1, 2008; 19(7): 2857 - 2869. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. E. Nowak, B. Tian, M. Jamaluddin, I. Boldogh, L. A. Vergara, S. Choudhary, and A. R. Brasier RelA Ser276 Phosphorylation Is Required for Activation of a Subset of NF-{kappa}B-Dependent Genes by Recruiting Cyclin-Dependent Kinase 9/Cyclin T1 Complexes Mol. Cell. Biol., June 1, 2008; 28(11): 3623 - 3638. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Tabone-Eglinger, F. Subra, H. El Sayadi, L. Alberti, E. Tabone, J.-P. Michot, N. Theou-Anton, A. Lemoine, J.-Y. Blay, and J.-F. Emile KIT Mutations Induce Intracellular Retention and Activation of an Immature Form of the KIT Protein in Gastrointestinal Stromal Tumors Clin. Cancer Res., April 15, 2008; 14(8): 2285 - 2294. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Michalczyk, E. Bastow, M. Greenough, J. Camakaris, D. Freestone, P. Taylor, M. Linder, J. Mercer, and M. L. Ackland ATP7B Expression in Human Breast Epithelial Cells Is Mediated by Lactational Hormones J. Histochem. Cytochem., April 1, 2008; 56(4): 389 - 399. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Lin, L. Landmann, M. A. Ruegg, and H. R. Brenner The Role of Nerve- versus Muscle-Derived Factors in Mammalian Neuromuscular Junction Formation J. Neurosci., March 26, 2008; 28(13): 3333 - 3340. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Conus, R. Perozzo, T. Reinheckel, C. Peters, L. Scapozza, S. Yousefi, and H.-U. Simon Caspase-8 is activated by cathepsin D initiating neutrophil apoptosis during the resolution of inflammation J. Exp. Med., March 17, 2008; 205(3): 685 - 698. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Piazzon, F. Rage, F. Schlotter, H. Moine, C. Branlant, and S. Massenet In Vitro and in Cellulo Evidences for Association of the Survival of Motor Neuron Complex with the Fragile X Mental Retardation Protein J. Biol. Chem., February 29, 2008; 283(9): 5598 - 5610. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. Krzewski, X. Chen, and J. L. Strominger WIP is essential for lytic granule polarization and NK cell cytotoxicity PNAS, February 19, 2008; 105(7): 2568 - 2573. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Sanchez, M. Escobar, Z. Pedrozo, P. Macho, R. Domenech, S. Hartel, C. Hidalgo, and P. Donoso Exercise and tachycardia increase NADPH oxidase and ryanodine receptor-2 activity: possible role in cardioprotection Cardiovasc Res, January 15, 2008; 77(2): 380 - 386. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Singh, M. Plassmeyer, D. Gaur, and L. H. Miller Mononeme: A new secretory organelle in Plasmodium falciparum merozoites identified by localization of rhomboid-1 protease PNAS, December 11, 2007; 104(50): 20043 - 20048. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. W. D. Comeau, S. Costantino, and P. W. Wiseman A Guide to Accurate Fluorescence Microscopy Colocalization Measurements Biophys. J., December 15, 2006; 91(12): 4611 - 4622. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. Cella, A. Contreras, K. Latha, J. M. Rosen, and M. Zhang Maspin is physically associated with {beta}1 integrin regulating cell adhesion in mammary epithelial cells FASEB J, July 1, 2006; 20(9): 1510 - 1512. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Shah, K. Patel, S. Mukhopadhyay, F. Xu, G. Guo, and P. B. Sehgal Membrane-associated STAT3 and PY-STAT3 in the Cytoplasm J. Biol. Chem., March 17, 2006; 281(11): 7302 - 7308. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L.-S. Song, E. A. Sobie, S. McCulle, W. J. Lederer, C. W. Balke, and H. Cheng Orphaned ryanodine receptors in the failing heart. PNAS, March 14, 2006; 103(11): 4305 - 4310. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. J. North Seeing is believing? A beginners' guide to practical pitfalls in image acquisition J. Cell Biol., January 3, 2006; 172(1): 9 - 18. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. Jia, L. Pignataro, C. M. Schofield, M. Yue, N. L. Harrison, and P. A. Goldstein An Extrasynaptic GABAA Receptor Mediates Tonic Inhibition in Thalamic VB Neurons J Neurophysiol, December 1, 2005; 94(6): 4491 - 4501. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Sampieri, M. Diaz-Munoz, A. Antaramian, and L. Vaca The Foot Structure from the Type 1 Ryanodine Receptor Is Required for Functional Coupling to Store-operated Channels J. Biol. Chem., July 1, 2005; 280(26): 24804 - 24815. [Abstract] [Full Text] [PDF]
|
|
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
