Abstract
An initial and crucial step in the establishment of many microbial infections is the attachment of the pathogen to the host cells. Thus, adherence of Pneumocystis carinii (Pc) to type I pneumocytes is believed to be important in the induction of Pc pneumonia. Little is known about the nature of the attachment of Pc to type I cells, although extracellular matrix (ECM) proteins, such as fibronectin and laminin, have been implicated in the process. We report here the isolation of a Pc gene encoding a receptor protein that binds both fibronectin and laminin in vitro. A cDNA clone encoding the Pc ECM receptor was isolated from a Pc cDNA library and identified on the basis of sequence homology to the human colon carcinoma laminin receptor. Southern blot analysis of Pc genomic DNA confirmed that the cDNA was of Pc origin. Northern blot analysis of Pc total RNA showed a predominant mRNA of approximately 1400 nucleotides that hybridized to the ECM receptor gene. The ECM receptor predicted from the cDNA sequence is 295 amino acid residues long, with a molecular mass of 32.8 kDa. The C-terminal third of the polypeptide is highly negatively charged, whereas the N-terminal two-thirds contains hydrophobic segments that may play a role in membrane association. Sequence analysis and alignment of the N terminus with the laminin receptor cDNA sequence of human colon carcinoma support the conclusion that the Pc ECM receptor cDNA clone is a full-length clone. A Western blot of the overexpressed ECM receptor protein bound both laminin and fibronectin in vitro. Antibodies raised to the overexpressed receptor protein interacted with a 33-kDa protein in total Pc cell lysates. These findings raise the possibility that the Pc ECM receptor protein may mediate the organism's attachment to type I pneumocytes and, thus, may play a crucial role in Pc pathogenesis.
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- Bouchara J. P., Tronchin G., Annaix V., Robert R., Senet J. M. Laminin receptors on Candida albicans germ tubes. Infect Immun. 1990 Jan;58(1):48–54. doi: 10.1128/iai.58.1.48-54.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Edlind T. D., Bartlett M. S., Weinberg G. A., Prah G. N., Smith J. W. The beta-tubulin gene from rat and human isolates of Pneumocystis carinii. Mol Microbiol. 1992 Nov;6(22):3365–3373. doi: 10.1111/j.1365-2958.1992.tb02204.x. [DOI] [PubMed] [Google Scholar]
- Edman J. C., Edman U., Cao M., Lundgren B., Kovacs J. A., Santi D. V. Isolation and expression of the Pneumocystis carinii dihydrofolate reductase gene. Proc Natl Acad Sci U S A. 1989 Nov;86(22):8625–8629. doi: 10.1073/pnas.86.22.8625. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Edman U., Edman J. C., Lundgren B., Santi D. V. Isolation and expression of the Pneumocystis carinii thymidylate synthase gene. Proc Natl Acad Sci U S A. 1989 Sep;86(17):6503–6507. doi: 10.1073/pnas.86.17.6503. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ezekowitz R. A., Williams D. J., Koziel H., Armstrong M. Y., Warner A., Richards F. F., Rose R. M. Uptake of Pneumocystis carinii mediated by the macrophage mannose receptor. Nature. 1991 May 9;351(6322):155–158. doi: 10.1038/351155a0. [DOI] [PubMed] [Google Scholar]
- Farkas I., Hardy T. A., DePaoli-Roach A. A., Roach P. J. Isolation of the GSY1 gene encoding yeast glycogen synthase and evidence for the existence of a second gene. J Biol Chem. 1990 Dec 5;265(34):20879–20886. [PubMed] [Google Scholar]
- Fishman J. A., Samia J. A., Fuglestad J., Rose R. M. The effects of extracellular matrix (ECM) proteins on the attachment of Pneumocystis carinii to lung cell lines in vitro. J Protozool. 1991 Nov-Dec;38(6):34S–37S. [PubMed] [Google Scholar]
- Jönsson K., Signäs C., Müller H. P., Lindberg M. Two different genes encode fibronectin binding proteins in Staphylococcus aureus. The complete nucleotide sequence and characterization of the second gene. Eur J Biochem. 1991 Dec 18;202(3):1041–1048. doi: 10.1111/j.1432-1033.1991.tb16468.x. [DOI] [PubMed] [Google Scholar]
- Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
- Limper A. H., Standing J. E., Hoffman O. A., Castro M., Neese L. W. Vitronectin binds to Pneumocystis carinii and mediates organism attachment to cultured lung epithelial cells. Infect Immun. 1993 Oct;61(10):4302–4309. doi: 10.1128/iai.61.10.4302-4309.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Long E. G., Smith J. S., Meier J. L. Attachment of Pneumocystis carinii to rat pneumocytes. Lab Invest. 1986 Jun;54(6):609–615. [PubMed] [Google Scholar]
- Lopes J. D., dos Reis M., Brentani R. R. Presence of laminin receptors in Staphylococcus aureus. Science. 1985 Jul 19;229(4710):275–277. doi: 10.1126/science.3160113. [DOI] [PubMed] [Google Scholar]
- Peake P., Gooley A., Britton W. J. Mechanism of interaction of the 85B secreted protein of Mycobacterium bovis with fibronectin. Infect Immun. 1993 Nov;61(11):4828–4834. doi: 10.1128/iai.61.11.4828-4834.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pearson W. R., Lipman D. J. Improved tools for biological sequence comparison. Proc Natl Acad Sci U S A. 1988 Apr;85(8):2444–2448. doi: 10.1073/pnas.85.8.2444. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pottratz S. T., Martin W. J., 2nd Role of fibronectin in Pneumocystis carinii attachment to cultured lung cells. J Clin Invest. 1990 Feb;85(2):351–356. doi: 10.1172/JCI114445. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Radding J. A., Armstrong M. Y., Ullu E., Richards F. F. Identification and isolation of a major cell surface glycoprotein of Pneumocystis carinii. Infect Immun. 1989 Jul;57(7):2149–2157. doi: 10.1128/iai.57.7.2149-2157.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rao C. N., Castronovo V., Schmitt M. C., Wewer U. M., Claysmith A. P., Liotta L. A., Sobel M. E. Evidence for a precursor of the high-affinity metastasis-associated murine laminin receptor. Biochemistry. 1989 Sep 5;28(18):7476–7486. doi: 10.1021/bi00444a047. [DOI] [PubMed] [Google Scholar]
- Reim D. F., Speicher D. W. Microsequence analysis of electroblotted proteins. II. Comparison of sequence performance on different types of PVDF membranes. Anal Biochem. 1992 Nov 15;207(1):19–23. doi: 10.1016/0003-2697(92)90493-q. [DOI] [PubMed] [Google Scholar]
- Signäs C., Raucci G., Jönsson K., Lindgren P. E., Anantharamaiah G. M., Hök M., Lindberg M. Nucleotide sequence of the gene for a fibronectin-binding protein from Staphylococcus aureus: use of this peptide sequence in the synthesis of biologically active peptides. Proc Natl Acad Sci U S A. 1989 Jan;86(2):699–703. doi: 10.1073/pnas.86.2.699. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
- Talay S. R., Valentin-Weigand P., Jerlström P. G., Timmis K. N., Chhatwal G. S. Fibronectin-binding protein of Streptococcus pyogenes: sequence of the binding domain involved in adherence of streptococci to epithelial cells. Infect Immun. 1992 Sep;60(9):3837–3844. doi: 10.1128/iai.60.9.3837-3844.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wewer U. M., Liotta L. A., Jaye M., Ricca G. A., Drohan W. N., Claysmith A. P., Rao C. N., Wirth P., Coligan J. E., Albrechtsen R. Altered levels of laminin receptor mRNA in various human carcinoma cells that have different abilities to bind laminin. Proc Natl Acad Sci U S A. 1986 Oct;83(19):7137–7141. doi: 10.1073/pnas.83.19.7137. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Williams D. J., Radding J. A., Dell A., Khoo K. H., Rogers M. E., Richards F. F., Armstrong M. Y. Glucan synthesis in Pneumocystis carinii. J Protozool. 1991 Jul-Aug;38(4):427–437. doi: 10.1111/j.1550-7408.1991.tb01382.x. [DOI] [PubMed] [Google Scholar]
- Yow H. K., Wong J. M., Chen H. S., Lee C. G., Davis S., Steele G. D., Jr, Chen L. B. Increased mRNA expression of a laminin-binding protein in human colon carcinoma: complete sequence of a full-length cDNA encoding the protein. Proc Natl Acad Sci U S A. 1988 Sep;85(17):6394–6398. doi: 10.1073/pnas.85.17.6394. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Zhang J., Stringer J. R. Cloning and characterization of an alpha-tubulin-encoding gene from rat-derived Pneumocystis carinii. Gene. 1993 Jan 15;123(1):137–141. doi: 10.1016/0378-1119(93)90553-f. [DOI] [PubMed] [Google Scholar]
- von der Mark K., Kühl U. Laminin and its receptor. Biochim Biophys Acta. 1985 Dec 17;823(2):147–160. doi: 10.1016/0304-419x(85)90010-1. [DOI] [PubMed] [Google Scholar]