This Article 
   
 Share 
   
 Bibliographic References 
   
 Add to: 
 
Digg
Furl
Spurl
Blink
Simpy
Google
Del.icio.us
Y!MyWeb
 
 Search 
   
Spatio-Temporal Analysis of Constitutive Exocytosis in Epithelial Cells
January-March 2006 (vol. 3 no. 1)
pp. 17-32
Exocytosis is an essential cellular trafficking process integral to the proper distribution and function of a plethora of molecules, including transporters, receptors, and enzymes. Moreover, incorrect protein targeting can lead to pathological conditions. Recently, the application of evanescent wave microscopy has allowed us to image the final steps of exocytosis. However, spatio-temporal analysis of fusion of constitutive vesicular traffic with the plasma membrane has not been systematically performed. Also, the spatial sites and times of vesicle fusion have not yet been analyzed together. In addition, more formal tests are required in testing biological hypotheses, rather than visual inspection combined with statistical descriptives. Ripley {\cal K}{\hbox{-}}{\rm functions} are used to examine the joint and marginal behavior of locations and fusion times. Semiautomatic detection and mapping of constitutive fusion sites reveals spatial and temporal clustering, but no dependency between the locations and times of fusion events. Our novel approach could be translated to other studies of membrane trafficking in health and diseases such as diabetes.

[1] A. Agresti, Categorical Data Analysis, second ed. Wiley, 2002.
[2] B. Alberts, Molecular Biology of the Cell, third ed. New York and London: Garland Publishing, 1994.
[3] M.W. Allersma, L. Wang, D. Axelrod, and R.W. Holz, “Visualization of Regulated Exocytosis with a Granule-Membrane Probe Using Total Internal Reflection Microscopy,” Molecular Biology of the Cell, vol. 15, no. 10, pp. 4658-4668, 2004.
[4] D. Axelrod, “Total Internal Reflection Fluorescence Microscopy in Cell Biology,” Traffic, vol. 2, pp. 764-774, 2001.
[5] D.R. Cox and V. Isham, Point Processes. Londong: Chapman and Hall, 1980.
[6] N.A.C. Cressie, Statistics for Spatial Data, revised ed. New York: John Wiley and Sons, 1993.
[7] D. Dernick, G. Alvarez de Toledo, and M. Lindau, “Exocytosis of Single Chromaffin Granules in Cell-Free Inside-Out Membrane Patches,” Nature Cell Biology, vol. 5, pp. 358-362, 2003.
[8] P.J. Diggle, Statistical Analysis of Spatial Point Patterns, second ed. London: Ar nold, 2003.
[9] P.J. Diggle, A.G. Chetwynd, R. Haggkvist, and S.E. Morris, “Second-Order Analysis of Space-Time Clustering,” Statistical Methods in Medical Research, vol. 4, pp. 124-136, 1995.
[10] M. Ehrlich, W. Boll, A. Van Oijen, K. Hariharan, R. Chandran, M.L. Nibert, and T. Kirchhausen, “Endocytosis by Random Initiation and Stabilization of Clathrin-Coated Pits,” Cell, vol. 118, pp. 591-605, 2004.
[11] A.C. Gatrell, T.C. Bailey, P.J. Diggle, and B.S. Rowlingson, “Spatial Point Pattern Analysis and Its Application in Geographical Epidemiology,” Trans.— Inst. of British Geographers, vol. 21, pp. 256-274, 1996.
[12] R.C. Gonzalez and R. Woods, Digital Image Processing. New York: Addison-Wesley, 1992.
[13] J.P. Henry, F. Darchen, and S. Cribier, “Physical Techniques for the Study of Exocytosis in Isolated Cells,” Biochimie, vol. 80, pp. 371-377, 1998.
[14] K. Hirschberg, C.M. Miller, J. Ellenberg, J.F. Presley, E.D. Siggia, R.D. Phair, and J. Lippincott-Schwartz, “Kinetic Analysis of Secretory Protein Traffic and Characterization of Golgi to Plasma Membrane Transport Intermediates in Living Cells,” J. Cell Biology, vol. 143, pp. 1485-1503, 1998.
[15] P. Keller, D. Toomre, E. Diaz, J. White, and K. Simons, “Multicolor Imaging of Post-Golgi Sorting and Trafficking in Live Cells,” Nature Cell Biology, vol. 3, pp. 140-149, 2001.
[16] G. Kreitzer, “Three Dimensional Analysis of Post-Golgi Carrier Exocytosis in Epithelial Cells,” Nature Cell Biology, vol. 5, pp. 126-139, 2003.
[17] Y. Mimori-Kiyosue and S. Tsukita, “`Search-and-Capture' of Microtubules through Plus-End-Binding Proteins (+Tips),” J. Biochemistry (Tokyo), vol. 134, pp. 321-326, 2003.
[18] D.J. O'Brien, J.B. Kaneene, A. Getis, J.W. Lloyd, M.R. Rip, and R.W. Leader, “Spatial and Temporal Distribution of Selected Canine Cancers in Michigan,” Preventive Veterinary Medicine, vol. 42, pp. 1-15, 1999.
[19] G.E. Palade, “Intracellular Aspects of the Process of Protein Secretion,” Science, vol. 189, pp. 347-1358, 1975.
[20] S. Pfeiffer, S.D. Fuller, and K. Simons, “Intracellular Sorting and Basolateral Appearance of the g Protein of Vesicular Stomatitis Virus in Madin-Darby Canine Kidney Cells,” J. Cell Biology, vol. 101, pp. 470-476, 1975.
[21] A.A. Philimonenko and P. Janaček, and J. Hozák, “Statistical Evaluation of Colocalization Patterns in Immunogold Labeling Experiments,” J. Structural Biology, vol. 132, pp. 201-210, 2000.
[22] I.A. Prior, C. Muncke, R.G. Parton, and J.F. Hancock, “Direct Visualization of Ras Proteins in Spatially Distinct Cell Surface Microdomains,” J. Cell Biology, vol. 160, pp. 165-170, Apr. 2004.
[23] R Development Core Team, R: A Language and Environment for Statistical Computing. R Foundation for Statistical Computing, Vienna, Austria, 2004.
[24] S. Reader, “Using Survival Analysis to Study Spatial Point Patterns in Geographical Epidemiology,” Social Science & Medicine, vol. 50, pp. 985-1000, 2000.
[25] P. Salembier and J. Serra, “Flat Zones Filtering, Connected Operators, and Filters by Reconstruction,” IEEE Trans. Image Processing, vol. 4, no. 8, pp. 1153-1160, Aug. 1995.
[26] J. Schmoranzer, M. Goulian, D. Axelrod, and S.M. Simon, “Imaging Constitutive Exocytosis with Total Internal Reflection Fluorescence Microscopy,” J. Cell Biology, vol. 149, pp. 23-31, Apr. 2000.
[27] J. Schmoranzer and S.M. Simon, “Role of Microtubules in Fusion of Post-Golgi Vesicles to the Plasma Membrane,” Molecular Biology of the Cell, vol. 14, pp. 1558-1569, Apr. 2003.
[28] J. Serra, Image Analysis and Mathematical Morphology, vol. 1, fourth ed. San Diego, Calif., and London: Academic Press, 1993.
[29] B.M. Slepchencko, J.C. Schaff, J.H Carson, and L.M. Loew, “Computational Cell Biology: Spatiotemporal Simulation of Cellular Events,” Ann. Rev. Biophysical Biomorphology Structures, vol. 31, pp. 423-41, 2002.
[30] P. Soille, Morphological Image Analysis: Principles and Applications. Berlin: Springer-Verlag, 1999.
[31] D. Toomre, P. Keller, J. White, J.C. Olivo, and K. Simons, “Dual-Color Visualization of Trans-Golgi Network to Plasma Membrane Traffic Alone Microtubules in Living Cells,” J. Cell Science, vol. 112, pp. 21-33, 1999.
[32] D. Toomre and D.J. Manstein, “Lighting Up the Cell Surface with Evanescent Wave Microscopy,” Trends Cell Biology, vol. 11, pp. 298-303, 2001.
[33] D. Toomre, J.A. Steyer, P. Keller, W. Almers, and K. Simons, “Fusion of Constitutive Membrane Traffic with the Cell Surface Observed by Evanescent Wave Microscopy,” J. Cell Biology, vol. 149, pp. 33-40, 2000.
[34] D. Zenisek, J.A. Steyer, M.E. Feldman, and W. Almers, “A Membrane Marker Leaves Synaptic Vesicles in Milliseconds after Exocytosis in Retinal Bipolar Cells,” Neuron, vol. 35, pp. 1085-1097, 2002.

Index Terms:
Spatio-temporal clustering, exocytosis, epithelial cells, total internal reflection fluorescent microscopy (TIRFM).
Citation:
Rafael Sebastian, Mar?a-Elena Diaz, Guillermo Ayala, Kresimir Letinic, Jose Moncho-Bogani, Derek Toomre, "Spatio-Temporal Analysis of Constitutive Exocytosis in Epithelial Cells," IEEE/ACM Transactions on Computational Biology and Bioinformatics, vol. 3, no. 1, pp. 17-32, Jan.-March 2006, doi:10.1109/TCBB.2006.11
Usage of this product signifies your acceptance of the Terms of Use.