Abstract
Infection of genital epithelial cells by the closely related sexually transmitted pathogens Chlamydia trachomatis serovars E and L2 results in different clinical disease manifestations. Following entry into target host cells, individual vesicles containing chlamydiae fuse with one another to form one large inclusion. At the cellular level, the only obvious difference between these serovars is the time until inclusion maturation, which is 48 h for the invasive serovar L2 and 72 h for serovar E. To begin to define the intracellular events of these pathogens, the effect of cytoskeletal disruption on early endosome fusion and inclusion development in epithelial (HEC-1B) and fibroblast (McCoy) cells was analyzed by fluorescence microscopy. Disruption of microfilaments with cytochalasin D markedly reduced serovar E, but not serovar L2, infection of both cell lines. Conversely, microfilament as well as microtubule disruption, with colchicine or nocodazole, had no effect on serovar E inclusion development but resulted in the formation of multiple serovar L2 inclusions per cell during early and mid-development. Later in serovar L2 inclusion development (> 36 h postinfection), vesicles containing chlamydiae fused to form one large inclusion in the absence of an intact cytoskeleton. These results imply that (i) C. trachomatis serovar E may utilize a different pathway for uptake and development from serovar L2; (ii) these differences are consistent in both epithelial cells and fibroblasts; and (iii) the cytoskeleton plays a unique role in the infection of host cells by these two genital pathogens.
Full Text
The Full Text of this article is available as a PDF (4.2 MB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Aniento F., Emans N., Griffiths G., Gruenberg J. Cytoplasmic dynein-dependent vesicular transport from early to late endosomes. J Cell Biol. 1993 Dec;123(6 Pt 1):1373–1387. doi: 10.1083/jcb.123.6.1373. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Banerjee D., Manning C. P., Redman C. M. The in vivo effect of colchicine on the addition of galactose and sialic acid to rat hepatic serum glycoproteins. J Biol Chem. 1976 Jul 10;251(13):3887–3892. [PubMed] [Google Scholar]
- Blok J., Ginsel L. A., Mulder-Stapel A. A., Onderwater J. J., Daems W. T. The effect of colchicine on the intracellular transport of 3H-fucose-labelled glycoproteins in the absorptive cells of cultured human small-intestinal tissue. An autoradiographical and biochemical study. Cell Tissue Res. 1981;215(1):1–12. doi: 10.1007/BF00236244. [DOI] [PubMed] [Google Scholar]
- Blyth W. A., Taverne J. Some consequences of the multiple infection of cell cultures by TRIC organisms. J Hyg (Lond) 1972 Mar;70(1):33–37. doi: 10.1017/s0022172400022063. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Boll W., Partin J. S., Katz A. I., Caplan M. J., Jamieson J. D. Distinct pathways for basolateral targeting of membrane and secretory proteins in polarized epithelial cells. Proc Natl Acad Sci U S A. 1991 Oct 1;88(19):8592–8596. doi: 10.1073/pnas.88.19.8592. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bomsel M., Parton R., Kuznetsov S. A., Schroer T. A., Gruenberg J. Microtubule- and motor-dependent fusion in vitro between apical and basolateral endocytic vesicles from MDCK cells. Cell. 1990 Aug 24;62(4):719–731. doi: 10.1016/0092-8674(90)90117-w. [DOI] [PubMed] [Google Scholar]
- Bowser S. S., Travis J. L., Rieder C. L. Microtubules associate with actin-containing filaments at discrete sites along the ventral surface of Allogromia reticulopods. J Cell Sci. 1988 Mar;89(Pt 3):297–307. doi: 10.1242/jcs.89.3.297. [DOI] [PubMed] [Google Scholar]
- Campbell S., Richmond S. J., Yates P. The development of Chlamydia trachomatis inclusions within the host eukaryotic cell during interphase and mitosis. J Gen Microbiol. 1989 May;135(5):1153–1165. doi: 10.1099/00221287-135-5-1153. [DOI] [PubMed] [Google Scholar]
- Campbell S., Yates P. S., Richmond S. J. Chlamydia trachomatis infection of cultured motile cells after uptake of chlamydiae from the substratum. J Gen Microbiol. 1993 Sep;139(9):2151–2158. doi: 10.1099/00221287-139-9-2151. [DOI] [PubMed] [Google Scholar]
- Edds K. T. Effects of cytochalasin and colcemid on cortical flow in coelomocytes. Cell Motil Cytoskeleton. 1993;26(3):262–273. doi: 10.1002/cm.970260309. [DOI] [PubMed] [Google Scholar]
- Eilers U., Klumperman J., Hauri H. P. Nocodazole, a microtubule-active drug, interferes with apical protein delivery in cultured intestinal epithelial cells (Caco-2). J Cell Biol. 1989 Jan;108(1):13–22. doi: 10.1083/jcb.108.1.13. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Eissenberg L. G., Wyrick P. B., Davis C. H., Rumpp J. W. Chlamydia psittaci elementary body envelopes: ingestion and inhibition of phagolysosome fusion. Infect Immun. 1983 May;40(2):741–751. doi: 10.1128/iai.40.2.741-751.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Eissenberg L. G., Wyrick P. B. Inhibition of phagolysosome fusion is localized to Chlamydia psittaci-laden vacuoles. Infect Immun. 1981 May;32(2):889–896. doi: 10.1128/iai.32.2.889-896.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Friis R. R. Interaction of L cells and Chlamydia psittaci: entry of the parasite and host responses to its development. J Bacteriol. 1972 May;110(2):706–721. doi: 10.1128/jb.110.2.706-721.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Genty N., Bussereau F. Is cytoskeleton involved in vesicular stomatitis virus reproduction? J Virol. 1980 Jun;34(3):777–781. doi: 10.1128/jvi.34.3.777-781.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gilbert T., Le Bivic A., Quaroni A., Rodriguez-Boulan E. Microtubular organization and its involvement in the biogenetic pathways of plasma membrane proteins in Caco-2 intestinal epithelial cells. J Cell Biol. 1991 Apr;113(2):275–288. doi: 10.1083/jcb.113.2.275. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gottlieb T. A., Ivanov I. E., Adesnik M., Sabatini D. D. Actin microfilaments play a critical role in endocytosis at the apical but not the basolateral surface of polarized epithelial cells. J Cell Biol. 1993 Feb;120(3):695–710. doi: 10.1083/jcb.120.3.695. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gruenberg J., Howell K. E. Membrane traffic in endocytosis: insights from cell-free assays. Annu Rev Cell Biol. 1989;5:453–481. doi: 10.1146/annurev.cb.05.110189.002321. [DOI] [PubMed] [Google Scholar]
- Hansen S. H., Sandvig K., van Deurs B. Molecules internalized by clathrin-independent endocytosis are delivered to endosomes containing transferrin receptors. J Cell Biol. 1993 Oct;123(1):89–97. doi: 10.1083/jcb.123.1.89. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Herman B., Albertini D. F. A time-lapse video image intensification analysis of cytoplasmic organelle movements during endosome translocation. J Cell Biol. 1984 Feb;98(2):565–576. doi: 10.1083/jcb.98.2.565. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hodinka R. L., Davis C. H., Choong J., Wyrick P. B. Ultrastructural study of endocytosis of Chlamydia trachomatis by McCoy cells. Infect Immun. 1988 Jun;56(6):1456–1463. doi: 10.1128/iai.56.6.1456-1463.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hodinka R. L., Wyrick P. B. Ultrastructural study of mode of entry of Chlamydia psittaci into L-929 cells. Infect Immun. 1986 Dec;54(3):855–863. doi: 10.1128/iai.54.3.855-863.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hunziker W., Mâle P., Mellman I. Differential microtubule requirements for transcytosis in MDCK cells. EMBO J. 1990 Nov;9(11):3515–3525. doi: 10.1002/j.1460-2075.1990.tb07560.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Jin M., Snider M. D. Role of microtubules in transferrin receptor transport from the cell surface to endosomes and the Golgi complex. J Biol Chem. 1993 Aug 25;268(24):18390–18397. [PubMed] [Google Scholar]
- Kelly R. B. Microtubules, membrane traffic, and cell organization. Cell. 1990 Apr 6;61(1):5–7. doi: 10.1016/0092-8674(90)90206-t. [DOI] [PubMed] [Google Scholar]
- Kohtz D. S., Hanson V., Puszkin S. Novel proteins mediate an interaction between clathrin-coated vesicles and polymerizing actin filaments. Eur J Biochem. 1990 Sep 11;192(2):291–298. doi: 10.1111/j.1432-1033.1990.tb19226.x. [DOI] [PubMed] [Google Scholar]
- Kuznetsov S. A., Langford G. M., Weiss D. G. Actin-dependent organelle movement in squid axoplasm. Nature. 1992 Apr 23;356(6371):722–725. doi: 10.1038/356722a0. [DOI] [PubMed] [Google Scholar]
- Kübler E., Riezman H. Actin and fimbrin are required for the internalization step of endocytosis in yeast. EMBO J. 1993 Jul;12(7):2855–2862. doi: 10.1002/j.1460-2075.1993.tb05947.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Leopold P. L., McDowall A. W., Pfister K. K., Bloom G. S., Brady S. T. Association of kinesin with characterized membrane-bounded organelles. Cell Motil Cytoskeleton. 1992;23(1):19–33. doi: 10.1002/cm.970230104. [DOI] [PubMed] [Google Scholar]
- Liu S. M., Magnusson K. E., Sundqvist T. Microtubules are involved in transport of macromolecules by vesicles in cultured bovine aortic endothelial cells. J Cell Physiol. 1993 Aug;156(2):311–316. doi: 10.1002/jcp.1041560213. [DOI] [PubMed] [Google Scholar]
- Majeed M., Ernst J. D., Magnusson K. E., Kihlström E., Stendahl O. Selective translocation of annexins during intracellular redistribution of Chlamydia trachomatis in HeLa and McCoy cells. Infect Immun. 1994 Jan;62(1):126–134. doi: 10.1128/iai.62.1.126-134.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Majeed M., Gustafsson M., Kihlström E., Stendahl O. Roles of Ca2+ and F-actin in intracellular aggregation of Chlamydia trachomatis in eucaryotic cells. Infect Immun. 1993 Apr;61(4):1406–1414. doi: 10.1128/iai.61.4.1406-1414.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Majeed M., Kihlström E. Mobilization of F-actin and clathrin during redistribution of Chlamydia trachomatis to an intracellular site in eucaryotic cells. Infect Immun. 1991 Dec;59(12):4465–4472. doi: 10.1128/iai.59.12.4465-4472.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Margolis R. L., Wilson L. Addition of colchicine--tubulin complex to microtubule ends: the mechanism of substoichiometric colchicine poisoning. Proc Natl Acad Sci U S A. 1977 Aug;74(8):3466–3470. doi: 10.1073/pnas.74.8.3466. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Matteoni R., Kreis T. E. Translocation and clustering of endosomes and lysosomes depends on microtubules. J Cell Biol. 1987 Sep;105(3):1253–1265. doi: 10.1083/jcb.105.3.1253. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Oelschlaeger T. A., Guerry P., Kopecko D. J. Unusual microtubule-dependent endocytosis mechanisms triggered by Campylobacter jejuni and Citrobacter freundii. Proc Natl Acad Sci U S A. 1993 Jul 15;90(14):6884–6888. doi: 10.1073/pnas.90.14.6884. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Oka J. A., Weigel P. H. Microtubule-depolymerizing agents inhibit asialo-orosomucoid delivery to lysosomes but not its endocytosis or degradation in isolated rat hepatocytes. Biochim Biophys Acta. 1983 Dec 19;763(4):368–376. doi: 10.1016/0167-4889(83)90098-8. [DOI] [PubMed] [Google Scholar]
- Olmsted J. B., Borisy G. G. Microtubules. Annu Rev Biochem. 1973;42:507–540. doi: 10.1146/annurev.bi.42.070173.002451. [DOI] [PubMed] [Google Scholar]
- Osborn M., Weber K. Dimethylsulfoxide and the ionophore A23187 affect the arrangement of actin and induce nuclear actin paracrystals in PtK2 cells. Exp Cell Res. 1980 Sep;129(1):103–114. doi: 10.1016/0014-4827(80)90335-3. [DOI] [PubMed] [Google Scholar]
- Pagano R. E., Sepanski M. A., Martin O. C. Molecular trapping of a fluorescent ceramide analogue at the Golgi apparatus of fixed cells: interaction with endogenous lipids provides a trans-Golgi marker for both light and electron microscopy. J Cell Biol. 1989 Nov;109(5):2067–2079. doi: 10.1083/jcb.109.5.2067. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Parczyk K., Haase W., Kondor-Koch C. Microtubules are involved in the secretion of proteins at the apical cell surface of the polarized epithelial cell, Madin-Darby canine kidney. J Biol Chem. 1989 Oct 5;264(28):16837–16846. [PubMed] [Google Scholar]
- Prain C. J., Pearce J. H. Ultrastructural studies on the intracellular fate of Chlamydia psittaci (strain guinea pig inclusion conjunctivitis) and Chlamydia trachomatis (strain lymphogranuloma venereum 434): modulation of intracellular events and relationship with endocytic mechanism. J Gen Microbiol. 1989 Jul;135(7):2107–2123. doi: 10.1099/00221287-135-7-2107. [DOI] [PubMed] [Google Scholar]
- Quaroni A., Kirsch K., Weiser M. M. Synthesis of membrane glycoproteins in rat small-intestinal villus cells. Effect of colchicine on the redistribution of L-[1,5,6-3H]fucose-labelled membrane glycoproteins among Golgi, lateral basal and microvillus membranes. Biochem J. 1979 Jul 15;182(1):213–221. doi: 10.1042/bj1820213. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Reynolds D. J., Pearce J. H. Endocytic mechanisms utilized by chlamydiae and their influence on induction of productive infection. Infect Immun. 1991 Sep;59(9):3033–3039. doi: 10.1128/iai.59.9.3033-3039.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ridderhof J. C., Barnes R. C. Fusion of inclusions following superinfection of HeLa cells by two serovars of Chlamydia trachomatis. Infect Immun. 1989 Oct;57(10):3189–3193. doi: 10.1128/iai.57.10.3189-3193.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rogalski A. A., Bergmann J. E., Singer S. J. Effect of microtubule assembly status on the intracellular processing and surface expression of an integral protein of the plasma membrane. J Cell Biol. 1984 Sep;99(3):1101–1109. doi: 10.1083/jcb.99.3.1101. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rubino S., Fighetti M., Unger E., Cappuccinelli P. Location of actin, myosin, and microtubular structures during directed locomotion of Dictyostelium amebae. J Cell Biol. 1984 Feb;98(2):382–390. doi: 10.1083/jcb.98.2.382. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schachter J. The intracellular life of Chlamydia. Curr Top Microbiol Immunol. 1988;138:109–139. [PubMed] [Google Scholar]
- Schroer T. A., Sheetz M. P. Functions of microtubule-based motors. Annu Rev Physiol. 1991;53:629–652. doi: 10.1146/annurev.ph.53.030191.003213. [DOI] [PubMed] [Google Scholar]
- Shelden E., Wadsworth P. Observation and quantification of individual microtubule behavior in vivo: microtubule dynamics are cell-type specific. J Cell Biol. 1993 Feb;120(4):935–945. doi: 10.1083/jcb.120.4.935. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stults N. L., Fechheimer M., Cummings R. D. Relationship between Golgi architecture and glycoprotein biosynthesis and transport in Chinese hamster ovary cells. J Biol Chem. 1989 Nov 25;264(33):19956–19966. [PubMed] [Google Scholar]
- Söderlund G., Kihlström E. Effect of methylamine and monodansylcadaverine on the susceptibility of McCoy cells to Chlamydia trachomatis infection. Infect Immun. 1983 May;40(2):534–541. doi: 10.1128/iai.40.2.534-541.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tam J. E., Knight S. T., Davis C. H., Wyrick P. B. Eukaryotic cells grown on microcarrier beads offer a cost-efficient way to propagate Chlamydia trachomatis. Biotechniques. 1992 Sep;13(3):374–378. [PubMed] [Google Scholar]
- Thyberg J., Moskalewski S. Microtubules and the organization of the Golgi complex. Exp Cell Res. 1985 Jul;159(1):1–16. doi: 10.1016/s0014-4827(85)80032-x. [DOI] [PubMed] [Google Scholar]
- Van der Sluijs P., Bennett M. K., Antony C., Simons K., Kreis T. E. Binding of exocytic vesicles from MDCK cells to microtubules in vitro. J Cell Sci. 1990 Apr;95(Pt 4):545–553. doi: 10.1242/jcs.95.4.545. [DOI] [PubMed] [Google Scholar]
- Walker R. A., Sheetz M. P. Cytoplasmic microtubule-associated motors. Annu Rev Biochem. 1993;62:429–451. doi: 10.1146/annurev.bi.62.070193.002241. [DOI] [PubMed] [Google Scholar]
- Ward M. E., Murray A. Control mechanisms governing the infectivity of Chlamydia trachomatis for HeLa cells: mechanisms of endocytosis. J Gen Microbiol. 1984 Jul;130(7):1765–1780. doi: 10.1099/00221287-130-7-1765. [DOI] [PubMed] [Google Scholar]
- Ward M. E., Salari H. Control mechanisms governing the infectivity of Chlamydia trachomatis for hela cells: modulation by cyclic nucleotides, prostaglandins and calcium. J Gen Microbiol. 1982 Mar;128(3):639–650. doi: 10.1099/00221287-128-3-639. [DOI] [PubMed] [Google Scholar]
- Weiner O. H., Murphy J., Griffiths G., Schleicher M., Noegel A. A. The actin-binding protein comitin (p24) is a component of the Golgi apparatus. J Cell Biol. 1993 Oct;123(1):23–34. doi: 10.1083/jcb.123.1.23. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wyrick P. B., Choong J., Davis C. H., Knight S. T., Royal M. O., Maslow A. S., Bagnell C. R. Entry of genital Chlamydia trachomatis into polarized human epithelial cells. Infect Immun. 1989 Aug;57(8):2378–2389. doi: 10.1128/iai.57.8.2378-2389.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wyrick P. B., Davis C. H., Knight S. T., Choong J., Raulston J. E., Schramm N. An in vitro human epithelial cell culture system for studying the pathogenesis of Chlamydia trachomatis. Sex Transm Dis. 1993 Sep-Oct;20(5):248–256. doi: 10.1097/00007435-199309000-00002. [DOI] [PubMed] [Google Scholar]
- Zeichner S. L. Isolation and characterization of macrophage phagosomes containing infectious and heat-inactivated Chlamydia psittaci: two phagosomes with different intracellular behaviors. Infect Immun. 1983 Jun;40(3):956–966. doi: 10.1128/iai.40.3.956-966.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]