ABC  Vol.3 No.3 , June 2013
Homeostatic cell cycle and the origin of autophagosomal vesicles
ABSTRACT
The autophagosomes were identified in the viable cycloheximide (CHX)-treated cells which had an incapacitated translational process and thus disabled synthesis of endoplasmic reticulum (ER)-derived vesicular transporters. They were found devoid of the proteins transported from ER to cell organelles, were unable to fuse with ER, Golgi or mitochondria, and displayed affinity with lysosomes. The analysis of autophagosomes, derived from the CHX cell organelles, revealed that their lipid composition resemble that of the maternal organelle. Thus, the ER-derived autophagosomes were marked with the presence of phosphatidylinositol (PI), Golgi-derived vesicles contained sphingomyelin (SM) and glycosphingolipids (GLL), and the mitochondria-derived autophagosomes contained phosphatidylglycerol (PG) and cardiolipin (CL). The incubation of the vesicles with intact lysosomes afforded their and the lysosome membrane lipids degradation. The analysis of the products derived from incubation of lysosomes and autophagosomes with radiolabeled SM, in the presence and the absence of TritonX100, allowed us to conclude that during autophagosome degradation the lysosomal enzymes are not released to cytosol, and that only lysosomes contain the enzymes degrading membrane lipids. In summary, our findings allowed us to authenticate the vesicles generated in the CHX-treated cells as organelle-specific autophagosomes and to determine that complete cycle of cell restitution and debridement includes intralysosomal degradation of the lysosomal membrane engulfing the autophagosomes vesicles.

Cite this paper
Slomiany, A. and Slomiany, B. (2013) Homeostatic cell cycle and the origin of autophagosomal vesicles. Advances in Biological Chemistry, 3, 275-287. doi: 10.4236/abc.2013.33031.
References
[1]   Toulmay, A. and Prinz, W.A. (2011) Lipid transfer and signaling at organelle contact sites: The tip of the iceberg. Current Opinions in Cell Biology, 23, 458-463. doi:10.1016/j.ceb.2011.04.006

[2]   Orsi, A., Polson, H.E. and Tooze, S.A. (2010) Membrane trafficking events that partake in autophagy. Current Opinions in Cell Biology, 22, 150-156. doi:10.1016/j.ceb.2009.11.013

[3]   Saftig, P. and Klumperman, J. (2009) Lysosome biogenesis and lysosomal proteins: Trafficking meets function. National Review of the Molecular Cell Biology, 10, 623635. doi:10.1038/nrm2745

[4]   Mellman, I. and Nelson, W.J. (2008) Coordinated protein sorting, targeting and distribution in polarized cells. Nature Reviews/Molecular Cell Biology, 9, 833-855.

[5]   Walter, P. and Ron, D. (2011) The unfolded protein response: From stress pathway to homeostatic regulation. Science, 334, 1081-1086. doi:10.1126/science.1209038

[6]   Smith, M.H., Ploegh, H.L. and Weissman, J.S. (2011) Road to ruin: Targeting proteins for degradation in the endoplasmic reticulum. Science, 334, 1086-1074. doi:10.1126/science.1209235

[7]   Keiser, M., Alfalah, M., Propsting, M.J., Castelletti, D. and Naim, H.Y. (2006) Altered folding, turnover, and polarized sorting act in concert to define a novel pathomechanism of congenital sucrose-isomaltase deficiency. Journal of Biological Chemistry, 281, 14393-14399. doi:10.1074/jbc.M513631200

[8]   Bilder, D., Schober, M. and Perrimon, N. (2003) Integrated activity of PDZ complexes regulates epithelial polarity. Nature Cell Biology, 5, 53-58. doi:10.1038/ncb897

[9]   Sharma, N., Low, S.H., Misra, S., Pallavi, B. and Weimbs, T. (2006) Apical targeting of synthaxin 3 is essential for epithelial cell polarity. Journal of Cell Biology, 173, 937948. doi:10.1083/jcb.200603132

[10]   Lukacs, G.L. and Verkman, A.S. (2012) CFTR; folding, misfolding and correcting the ΔF508 conformational defect. Trends in Molecular Medicine, 18, 81-91. doi:10.1016/j.molmed.2011.10.003

[11]   Ron, E., Shenkman, M., Groisman, B., Izenshtein, Y.B., Leitman, J. and Lederkremer, G.Z. (2011) Bypyss of glycan-dependent glycoprotein delivery to ERAD by upregulated EDEM 1. Molecular Biology of the Cell, 22, 3945-3954. doi:10.1091/mbc.E10-12-0944

[12]   Rodriguez-Boulan, E. and Musch, A. (2005) Protein sorting in Golgi complex: Shifting paradigms. Biochimica Biophysica Acta, 1744, 455-464. doi:10.1016/j.bbamcr.2005.04.007

[13]   De Matteis, M.A. and Luini, A. (2008) Exiting the Golgi complex. Nature Reviews of Molecular Cell Biology, 9, 273-284. doi:10.1038/nrm2378

[14]   Folch, H. (2008) Regulation of membrane trafficking in polarized epithelial cells. Current Opinions in Cell Biology, 20, 208-213.

[15]   Slomiany, A. and Slomiany, B.L. (2010) Cell membranes composition is defined in ER and their restitution proceeds by en bloc fusion of ER generated transport vesicles. Health, 2, 1444-1454. doi:10.4236/health.2010.212214

[16]   Slomiany, A. and Slomiany, B.L. (2003) Lipidomic processes in homeostatic and LPS-modified cell renewal cycle. Role of phosphatidylinositol 3-kinase pathway in biomembrane synthesis and restitution of apical epithelial membrane. Journal Physiology and Pharmacology, 54, 533-551.

[17]   Slomiany, A., Grabska, M., Piotrowski, E. and Slomiany, B.L. (1994) Intracellular processes associated with vesicular transport from endoplasmic reticulum to Golgi and exocytosis. Archives of Biochemistry and Biophysics, 310, 247-255. doi:10.1006/abbi.1994.1164

[18]   Slomiany, A., Grzelinska, E., Grabska, M. and Slomiany, B.L. (1992) Intracelular processes associated with glycolprotein transport and processing. Archives of Biochemistry and Biophysics, 298, 167-175. doi:10.1016/0003-9861(92)90108-9

[19]   Slomiany, A., Grzelinska, E., Yamaki, K. and Slomiany, B.L. (1992) Function of intracellular phospholipase A2 in vectorial transport of apoproteins from ER to Golgi. International Journal of Biochemistry, 24, 1397-1406.

[20]   Slomiany, A., Grzelinska, E., Kasinathan, C., et al. (1992) Biogenesis of endoplasmic reticulum transport vesicles transferring gastric apomucin from ER to Golgi. Experimental Cell Research, 201, 1669-1682. doi:10.1016/0014-4827(92)90280-L

[21]   Slomiany, A. and Slomiany, B.L. (2011) Transformations of phosphatidylinositol phosphates in the outer and inner nuclear membrane are linked to synthesis and restitution of cellular membranes. Health, 3, 187-199. doi:10.4236/health.2011.34035

[22]   Slomiany, A., Grabska, M. and Slomiany, B.L. (2006) Homeostatic restitution of cell membranes. Nuclear membrane lipid biogenesis and transport of protein from cytosol to intranuclear spaces. International Journal of Biological Sciences, 2, 216-226. doi:10.7150/ijbs.2.216

[23]   Slomiany, A., Nowak, P., Piotrowski, E. and Slomiany, B.L. (1998) Effect of ethanol on intracellular vesicular transport from Golgi to the apical membrane. Role of phosphatidylinositol 3-kinase and phospholipase A2 in Golgi transport vesicles association and fusion with the apical membrane. Alcohol, Clinical and Experimental Research, 22, 167-175. doi:10.1111/j.1530-0277.1998.tb03634.x

[24]   Slomiany, A., Grabska, M., Slomiany, B.A., et al. (1993) Intracellular transport, organelle biogenesis and establishment of Golgi identity: Impact of Brefeldin A on the activity of lipid synthesizing enzymes. International Journal of Biochemistry, 25, 891-901. doi:10.1016/0020-711X(93)90245-A

[25]   Slomiany, A. and Slomiany, B.L. (2012) Phosphatidylglycerol-containing ERtransport vesicles built and restore outer mitochondrial membrane and deliver nuclear DNA translation products to generate cardiolipin in the inner mitochondrial membrane. Advances in Biological Chemistry, 2, 132-145.

[26]   Slomiany, A., Sano, S., Grabska, M., Yamaki, K. and Slomiany, B.L. (2004) Gastric mucosal cell homeostatic physiome. Critical role of ER-initiated membrane restitution in the fidelity of cell function renewal. Journal of Physiology and Pharmacology, 55, 837-860.

[27]   Mari, M., Tooze, S.E. and Reggiori, F. (2011) The puzzling origin of the autophagosomal membrane. F1000 Biology Reports, 3, 25-34.

[28]   Maiuri, M.C., Zalckvar, E., Kimchi, A. and Kroemer, G. (2007) Self-eating and self-killing: Cross talk between autophagy and apoptosis. National Reviews in Molecular Cell Biology, 8, 741-752. doi:10.1038/nrm2239

[29]   Yoshimori, T. and Noda, T. (2008) Toward unraveling membrane biogenesis in mammalian autophagy. Current Opinions in Cell Biology, 20, 401-407. doi:10.1016/j.ceb.2008.03.010

[30]   Tooze, S.E. and Yoshimuri, T. (2010) The origin of the autophagosomal membrane. National Cell Biology, 12, 831-835. doi:10.1038/ncb0910-831

[31]   Brodsky, J.L. (2012) Cleaning up: ER-associated degradation to the rescue. Cell, 151, 1163-1167. doi:10.1016/j.cell.2012.11.012

[32]   Ragusa, M.J., Stanley, R.E. and Hurley, J.H. (2012) Architecture of the Atg17 complex as a scaffold for autophagosome biogenesis. Cell, 151, 1501-1512. doi:10.1016/j.cell.2012.11.028

[33]   Wong, E. and Cuervo, A.M. (2010) Autophagy gone awry in neurodegenerative diseases. National Neurosciences, 13, 805-811.

[34]   Croze, E.M. and Morre, D.J. (1984) Isolation of plasma membrane, Golgi apparatus, and endoplasmic reticulum from single homogenates of mouse liver. Journal of Cellular Physiology, 119, 46-57. doi:10.1002/jcp.1041190109

[35]   Parsons, D.F., Willims, G.R. and Chance, B. (1966) Characteristics of isolated and purified preparations of the outer and inner membranes of mitochondria. Annals New York Academy of Sciences, 137, 643-666. doi:10.1111/j.1749-6632.1966.tb50188.x

[36]   Tjelle, T.E., Brech, A., Juvet, L.K., Griffiths, G. and Berg, T. (1996) Isolation and characterization of early endosomes, late endosomes, late endosomes and terminal lysosomes: Their role in protein degradation. Journal of Cell Sciences, 109, 2905-2914.

[37]   Rome, L.H. and Crain, L.R. (1981) Degradation of mucopolysaccharide in intact isolated lysosomes. Journal of Biological Chemistry, 256, 10763-10768.

[38]   Kawashima, A., Sato, A., Kawashima, M., Nitta, K.,Yamura, W., Sugino, N., Nihei, H. and Natori, Y. (1998) A simple procedure for the isolation of rat kidney lysosomes. Kidney International, 54, 275-278. doi:10.1046/j.1523-1755.1998.00958.x

[39]   Schneider-Poetsch, T., Ju, J., Eyler, D.E., Dang, Y., Bhat, S., Merrick, W.C., Green, R., Shen, B. and Liu, J.O. (2010) Inhibition of eukaryotic translation elongation by cycloheximide and lactimidomycin. Nature Chemistry and Biology, 6, 209-217.

[40]   Obrig, T.G., Culp, W.J., McKeehan, W.L. and Hardesty, B. (1971) The mechanism by which cycloheximide and related glutarimide antibiotics inhibit peptide synthesis on reticulocyte ribosomes. Journal of Biological Chemistry, 246, 174-181.

[41]   Kapp, L.D. and Lorsch, J.R. (2004) The molecular mechanics of eukaryotic translation. Annual Reviews of Biochemistry, 73, 657-704. doi:10.1146/annurev.biochem.73.030403.080419

[42]   Lee, M.C., Miller, E.A., Goldberg, J., Orci, L. and Schekman, R. (2004) Bi-directional protein transport between the ER and Golgi. Annual Reviews of Cell Development and Biology, 20, 87-123. doi:10.1146/annurev.cellbio.20.010403.105307

[43]   Rothman, J.E. (1994) Mechanisms of intracellular protein transport. Nature, 372, 55-63. doi:10.1038/372055a0

[44]   Beck, R., Prinz, S., Distellkoter-Bechart, P., et al. (2011) Coatomer and dimeric ADP ribosylation factor 1 promote distinct steps in membrane scission. Journal of Cell Biology, 194, 765-777.

[45]   Idone, V., Tam, C. and Andrews, N.W. (2008) Two-way traffic on the road to plasma membrane repair. Trends in Cellular Biology, 18, 552-559. doi:10.1016/j.tcb.2008.09.001

[46]   Sarrl, E., Sicart, A., Lazaro-Dieguez, F. and Egea, G. (2011) Phospholipid synthesis participates in the regulation of diacylglycerol required for membrane trafficking at Golgi complex. The Journal of Biological Chemistry, 286, 28632-28643. doi:10.1074/jbc.M111.267534

[47]   Sudhof, T.C. and Rothman, J.E. (2009) Membrane fusion: Grappling with SNARE and SM proteins. Science, 323, 474-477. doi:10.1126/science.1161748

[48]   Glick, B.S. and Nakano, A. (2009) Membrane traffic within the Golgi apparatus. Annual Review of Cell Development Biology, 25, 113-132. doi:10.1146/annurev.cellbio.24.110707.175421

[49]   Kawano, M., Kumagai, K., Nishijima, M. and Handa, K. (2006) Efficient trafficking of ceramide from endoplasmic reticulum to the Golgi apparatus requires a VAMPassociated protein interacting FFAT motif of CERT. The Journal of Biological Chemistry, 281, 30279-30288. doi:10.1074/jbc.M605032200

[50]   van Meer, G., Halter, D., Sprong, H., Somerharju, P. and Egmond, M.R. (2006) ABC lipid transporters: Extruders, flippases, or flopless activators? FEBS Letters, 580, 11711177. doi:10.1016/j.febslet.2005.12.019

[51]   Mellman, I. and Nelson, W.J. (2009) Coordinated protein sorting, targeting and distribution in polarized cells. National Review of the Molecular Cell Biology, 9, 833-845.

[52]   Neupert, H. and Herrmann, J.M. (2007) Translocation of proteins into mitochondria. Annual Review of Biochemistry, 76, 723-749. doi:10.1146/annurev.biochem.76.052705.163409

[53]   Lemmon, M.A. (2008) Membrane recognition by phospholipids binding proteins. National Review of the Molecular Cell Biology, 9, 99-111.

[54]   Reggiori, V. and Tooze, S.H. (2012) Autophagy regulation through Atg9 traffic. Journal of Cell Biology, 198, 151-153. doi:10.1083/jcb.201206119

[55]   Whitelegge, J. (2011) Up close with membrane lipidprotein complexes. Science, 334, 320-321. doi:10.1126/science.1214084

[56]   Hayashi-Nishino, M., Fujita, M., Noda, T., Yamaguchi, A., Yoshimori, T. and Yamamoto, A. (2009) A subdomain of the endoplasmic reticulum forms a cradle for autophagosome formation. National Cell Biology, 5, 1180-1185.

[57]   Hailey, D.W., Rambold, A.S., Satpute-Krishnan, P., Mitra, K., Sougrat, R., Kim, P.K. and Lippincott-Schwartz, J. (2010) Mitochondria supply membranes for autophagosomal biogenesis during starvation. Cell, 141, 656-667. doi:10.1016/j.cell.2010.04.009

[58]   Ravikumar, B., Moreau, K., Jahreiss, L., Puri, C. and Rubinsztein, D.C. (2010) Plasma membrane contributes to the formation of pre-autophagosomal structures. National Cell Biology, 12, 747-757.

[59]   van der Vaart, A. and Reggiori, F., (2010) The Golgi complex as a source for yeast autophagosomal membranes. Autophagy, 6, 800-801. doi:10.4161/auto.6.6.12575

[60]   Marion, G., Madeo, F. and Kroemer, G. (2011) Autophagy for tissue homeostasis and neuroprotection. Current Opinions in Cell Biology, 23, 198-206. doi:10.1016/j.ceb.2010.10.001

[61]   Cay, Q., Lu, L., Tian, J.H., Zhu, Y.B., Qiao, H. and Sheng, Z.H. (2010) Snapin regulated late endosomal transport is critical for efficient autophagy-lysosomal function in neurons. Neuron, 68, 73-86.

[62]   Platt, F.M., Boland, B. and van der Spoel, A.C. (2012) Lysosomal storage disorders: The cellular impact of lysosomal dysfunction. Journal of Cell Biology, 199, 723734. doi:10.1083/jcb.201208152

[63]   Kale, J., Liu, Q., Leber, B. and Andrews, D.W. (2012) Shedding light on apoptosis at subcellular membranes. Cell, 151, 1179-1184. doi:10.1016/j.cell.2012.11.013

[64]   Osman, C., Voelker, D.R. and Langer, T. (2011) Making heads or tails of phospholipids in mitochondria. Journal of Cell Biology, 192, 7-16.

[65]   Gohil, V.M. and Greenberg, M.L. (2009) Mitochondrial membrane biogenesis: Phospholipids and proteins go hand in hand. Journal of Cell Biology, 184, 469-472.

[66]   Klausner, R.D., Donaldson, J.G. and Lippincott-Schwartz, J. (1992) Brefeldin A: Insights into the control of membrane traffic and organelle structure. Journal of Cell Biology, 116, 1071-1080. doi:10.1083/jcb.116.5.1071

 
 
Top