ABC  Vol.7 No.1 , February 2017
Lipid-Protein Microinclusions in the Morphological Structures of Organelle Membranes Studied by Fluorescent Confocal Microscopy
Abstract: Peculiar properties of morphological structures of organelle membranes were studied by fluorescent confocal microscopy. The list of objects in our experiments was represented by mitochondria, chloroplasts and vacuoles. During this study, identification of lipid microinclusions having the form of such lipid-protein structural microformations as lipid-protein microdomains, vesicles and membrane tubular structures (cytoplasmic transvacuolar strands and nanotubes) located in organelle membranes or bound up with them was conducted. Such membrane probes as laurdan, DPH, ANS and bis-ANS were used. Comparison of fluorescence intensity of these membrane probes was conducted. This investigation of the morphological properties of lipid-protein structural microformations was accompanied with analysis of 1) the phase state and 2) dynamics of microviscosity variations in the membrane elements of isolated plant cell organelles. Distributions of laurdan fluorescence generalized polarization (GP) values for the membrane on the whole and for the intensively fluorescing membrane segments were obtained. It was discovered that the microviscosity of intensively fluorescing membrane segments essentially differed from the microviscosity of the rest part of the membrane. In conclusion, some results of the study of peculiar properties of lipid-protein structural microformations related to the structure of organelle membranes and the discoveries made in this investigation are discussed.
Cite this paper: Chernyshov, M. , Nurminsky, V. and Ozolina, N. (2017) Lipid-Protein Microinclusions in the Morphological Structures of Organelle Membranes Studied by Fluorescent Confocal Microscopy. Advances in Biological Chemistry, 7, 42-59. doi: 10.4236/abc.2017.71003.

[1]   Sakuma, Y., Taniguchi, T., Kawakatsu, T. and Imai, M. (2013) Tubular Membrane Formation of Binary Giant Unilamellar Vesicles Composed of Cylinder and Inverse-Cone-Shaped Lipids. Biophysical Journal, 105, 2074-2081.

[2]   Roux, A. (2013) The Physics of Membrane Tubes: Soft Templates for Studying Cellular Membranes. Soft Matter, 9, 6726-6736.

[3]   Ozolina, N.V., Nesterkina, I.S., Kolesnikova, E.V., Salyaev, R.K., Nurminsky, V.N., Rakevich, A. L., Martynovich, E.F. and Chernyshov, M.Yu. (2013) Tonoplast of Beta vulgaris L. Contains Detergent-Resistant Membrane Microdomains. Planta, 237, 859-871.

[4]   Etxeberria, E., Gonzalez, P. and Pozueta-Romero, J. (2013) Architectural Remodeling of the Tonoplast during Fluid-Phase Endocytosis. Plant Signaling & Behavior, 8, e24793.

[5]   Yoshida, K., Ohnishi, M., Fukao, Y., Okazaki, Y., Fujiwara, M., Song, C., Nakanishi, Y., Saito, K., Shimmen, T., Suzaki, T., Yayashi, F., Fukaki, H., Maeshima, M. and Mimura, T. (2013) Studies on Vacuolar Membrane Microdomains Isolated from Arabidopsis Suspension-Cultured Cells: Local Distribution of Vacuolar Membrane Proteins. Plant and Cell Physiology, 54, 1571-1584.

[6]   Lingwood, D., Kaiser, H.J., Levental, I. and Simons, K. (2009) Lipid Rafts as Functional Heterogeneity in Cell Membranes. Biochemical Society Transactions, 37, 955-960.

[7]   Lingwood, D. and Simons, K. (2010) Lipid Rafts as a Membrane-Organizing Principle. Science, 327, 46-50.

[8]   Salyer, S.A., Olberding, J.R., Distler, A.A., Lederer, E.D., Clark, B.J., Delamere, N.A. and Khunfmiri, S.J. (2013) Vacuolar APTase Driven Potassium Transport in Highly Metastatic Breast Cancer Cells. Biochimica et Biophysica Acta, 1832, 1734-1743.

[9]   Queirós, F., Fontes, N., Silva, P., Almeida, D., Maeshima, M., Gerós, H. and Fidalgo, F. (2009) Activity of Tonoplast Proton Pumps and Na+/H+ Exchange in Potato Cell Cultures Is Modulated by Salt. Journal of Experimental Botany, 60, 1363-1374.

[10]   Janicka-Russak, M., Kabala, K., Mlodzinska, E. and Klobus, G. (2010) The Role of Polyamines in the Regulation of the Plasma Membrane and the Tonoplast Proton Pumps Under Salt Stress. The Journal of Plant Physiology, 167, 261-269.

[11]   Wang, L., He, X., Zhao, Y., Shen, Y. and Huang, Z. (2011) Wheat Vacuolar H+-ATPase Subunit B Cloning and Its Involvement in Salt Tolerance. Planta, 234, 1-7.

[12]   Zhang, M., Fang, Y., Liang, Z. and Huang, L. (2012) Enhanced Expression of Vacuolar H+-ATPase Subunit E in the Roots Is Associated with the Adaptation of Broussonetia papyrifera to Salt Stress. PLoS ONE, 7, e48183.

[13]   Andreyev, I.M. (2001) Vacuole Functions in Higher Plant Cells. Fiziologiya Rastenii, 48, 777-787.

[14]   Andreyev, I.M. (2012) The Role of Vacuole in Redox Homeostasis of Plant Cells. Fiziologiya Rastenii, 59, 660-667.

[15]   Sheahan, M.B., Rose, R.J. and McCurdy, D.W. (2007) Actin-Filament-Dependent Remodeling of the Vacuole in Cultured Mesophyll Protoplasts. Protoplasma, 230, 141-152.

[16]   Assani, A., Moundanga, S., Beney, L. and Gervais, P. (2009) Vesicle Formation in the Membrane of Onion Cells (Allium cepa) during Rapid Osmotic Dehydration. Annals of Botany, 104, 1389-1395.

[17]   Nurmimsky, V.N., Ozolina, N.V., Nesterkina, I.S., Kolesnikova, E.V., Salyaev, R.K., Rakevich, A.L., Martynovich, E.F., Pilipchenko, A.A. and Chernyshov, M.Y. (2015) Peculiar Properties of Some Components in a Plant Cell Vacuole Morphological Structure Revealed by Confocal Microscopy. Cell and Tissue Biology, 9, 406-414.

[18]   Loiseau, E., Schneider, J.A., Keber, F.C., Pelzl, C., Massiera, G., Salbreux, G. and Bausch, A.R. (2016) Shape Remodeling and Blebbing of Active Cytoskeletal Vesicles. Science Advances, 2, e1500465.

[19]   Nurminsky, V.N., Chernyshov, M.Y., Ozolina, N.V., Nesterkina, I.S., Kolesnikova, E.V., Rakevich, A.L., Martynovich, E.F. and Salyaev, R.K. (2012) Detergent-Resistant Microdomains (Rafts) in Tonoplast. In: Rubin, A.B., Ed., Proceedings of 4th Symposium of Russian Biophysicists Session 4. New Tendencies and Methods in Biophysics, University Publishing, Nizhnii Novgorod, 74.

[20]   Gao, X.Q., Wang, X.L., Ren, F., Chen, J. and Wang, X.C. (2009) Dynamics of Vacuoles and Actin Filaments in Guard Cells and Their Roles in Stomatal Movement. Plant, Cell & Environment, 32, 1108-1116.

[21]   Neuhaus, H.E. and Trentmann, O. (2014) Regulation of Transport Processes across the Tonoplast. Frontiers in Plant Science, 5, 460.

[22]   Salyaev, R.K., Kuzevanov, V.Y., Khaptagaev, V.Y. and Kopytchuk, V.N. (1981) Isolation and Purification of Vacuoles and Vacuolar Membranes from Plant Cells. Fiziologiya Rastenii, 28, 1295-1305.

[23]   Gaus, K., Zech, T. and Harder, T. (2006) Visualizing Membrane Microdomains by Laurdan 2-Photon Microscopy. Molecular Membrane Biology, 23, 41-48.

[24]   Baranov, S.I., Nurminsky, H.N. and Nurminsky, V.N. (2014) Automation of Fluorescent Microscopy Data Processing in Course of Measurement of Membrane Microviscosity. Proceedings of 27th International Science Conference on Mathematical Methods in Engineering and Technology, Tambov, 175-178.

[25]   Dabora, S.L. and Sheetz, M.P. (1988) The Microtubule-Dependent Formation of a Tubulovesicular Network with Characteristics of the ER from Cultured Cell Extracts. Cell, 54, 27-35.

[26]   Reisen, D., Marty, F. and Leborgne-Castel, N. (2005) New Insights into the Tonoplast Architecture of Plant Vacuoles and Vacuolar Dynamics during Osmotic Stress. BMC Plant Biology, 5, 13.

[27]   Lollike, K. and Lindau, M. (1999) Membrane Capacitance Techniques to Monitor Granule Exocytosis in Neutrophils. Journal of Immunological Methods, 232, 111-120.

[28]   Farsad, K. and De Camilli, P. (2003) Mechanisms of Membrane Deformation. Current Opinion in Cell Biology, 15, 372-381.

[29]   Dautry-Vasart, A. and Luini, A. (2003) Membranes and Organelles. Current Opinion in Cell Biology, 15, 369-371.

[30]   Rustom, A., Saffrich, R., Markovic, I., Walther, P. and Gerdes, H.H. (2004) Nano-Tubular Highways for Intercellular Organelle Transport. Science, 303, 1007-1010.

[31]   Koster, G., VanDuijn, M., Hofs, B. and Dogterom, M. (2003) Membrane Tube Formation from Giant Vesicles by Dynamic Association of Motor Proteins. PNAS, 100, 15583-15588.

[32]   Liu, A.P. and Fletcher, D.A. (2006) Actin Polymerization Serves as a Membrane Domain Switch in Model Lipid Bilayers. Biophysical Journal, 91, 4064-4070.

[33]   Neumann, A.K., Itano, M.S. and Jacobson, K. (2010) Understanding Lipid Rafts and Other Related Membrane Domains. F1000 Biology Reports, 2, 31.

[34]   Oda, Y., Higaki, T., Hasezawa, S. and Kutsuna, N. (2009) New Insights into Plant Vacuolar Structure and Dynamics. International Review of Cell and Molecular Biology, 277, 103-135.

[35]   Duman, J.G., Pathak, N.J., Ladinsky, M.S., Mc Donald, K.L. and Forte, J.G. (2002) Three-Dimensional Reconstruction of Cytoplasmic Membrane Networks in Parietal Cells. Journal of Cell Science, 115, 1251-1258.

[36]   Schneckenburger, H., Wagner, M., Kretzschmar, M., Strauss, W.S.L. and Sailer, R. (2004) Laser-Assisted Fluorescence Microscopy for Measuring Cell Membrane Dynamics. Photochemical & Photobiological Sciences, 3, 817-822.

[37]   Antonov, V.F., Chernysh, A.M., Pasechnik, V.I., Voznesensky, S.A. and Kozlova, E.K. (1999) Biophysics. Vlados Publising, Moscow, 288 p.

[38]   Guillier, C., Cacas, J.-L., Recorbet, G., Depretre, N., Mounier, A., Mongrand, S., Simon-Plas, F., Wipf, D. and Dumas-Gaudot, E. (2014) Direct Purification of Detergent-Insoluble Membranes from Medicago truncatula Root Microsomes: Comparison between Flotation and Sedimentation. BMC Plant Biology, 14, 255.

[39]   Bagnat, M., Keranen, S., Shevchenko, A., Shevchenko, A. and Simons, K. (2000) Lipid Rafts Function in Biosynthetic Delivery of Proteins to the Cell Surface in Yeast. Proceedings of the National Academy of Sciences, 97, 3254-3259.

[40]   Brown, D. (2002) Structure and Function of Membrane Rafts. International Journal of Medical Microbiology, 291, 433-437.

[41]   Vladimirov, Y.A., Roshupkin, D.I., Potapenko, A.Y. and Deyev, A.I. (1983) Biophysics. Medicine Publishing, Moscow, 272 p.

[42]   Boldyrev, A.A., Kyaivyaryainen, E.I. and Ilyuha, B.A. (2006) Biomembranology. Karelian Scientific Center of RAS Publishing, Petrozavodsk, 226 p.

[43]   Los, D.A. and Murata, N. (2004) Membrane Fluidity and Its Role in the Perception of Environmental Signals. Biochimica et Biophysica Acta, 1666, 142-157.