Abstract
The flagellum of the parasitic hemoflagellate Trypanosoma brucei contains two major structures: (a) the microtubule axoneme, and (b) a highly ordered, filamentous array, the paraflagellar rod (PFR). This is a complex, three-dimensional structure, of yet unknown function, that extends along most of the axoneme and is closely linked to it. Its major structural component is a single protein of 600 amino acids. This PFR protein can assume two different conformations, resulting in two distinct bands of apparent molecular masses of 73 and 69 kD in SDS-gel electrophoresis. Secondary structure predictions indicate a very high helix content. Despite its biochemical similarity to the intermediate filament proteins (solubility properties, amino acid composition, and high degree of helicity), the PFR protein does not belong in this class of cytoskeletal proteins. The PFR protein is coded for by two tandemly linked genes of identical nucleotide sequence. Both genes are transcribed into stable mRNAs of very similar length that carry the mini-exon sequence at their 5' termini.
Full Text
The Full Text of this article is available as a PDF (3.1 MB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Bartnik E., Osborn M., Weber K. Intermediate filaments in non-neuronal cells of invertebrates: isolation and biochemical characterization of intermediate filaments from the esophageal epithelium of the mollusc Helix pomatia. J Cell Biol. 1985 Aug;101(2):427–440. doi: 10.1083/jcb.101.2.427. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Borst P., van der Ploeg M., van Hoek J. F., Tas J., James J. On the DNA content and ploidy of trypanosomes. Mol Biochem Parasitol. 1982 Jul;6(1):13–23. doi: 10.1016/0166-6851(82)90049-4. [DOI] [PubMed] [Google Scholar]
- Brun R., Schönenberger Cultivation and in vitro cloning or procyclic culture forms of Trypanosoma brucei in a semi-defined medium. Short communication. Acta Trop. 1979 Sep;36(3):289–292. [PubMed] [Google Scholar]
- Clayton C. E. Structure and regulated expression of genes encoding fructose biphosphate aldolase in Trypanosoma brucei. EMBO J. 1985 Nov;4(11):2997–3003. doi: 10.1002/j.1460-2075.1985.tb04035.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cunha N. L., De Souza W., Hassón-Voloch A. Isolation of the flagellum and characterization of the paraxial structure of Herpetomonas megaseliae. J Submicrosc Cytol. 1984 Oct;16(4):705–713. [PubMed] [Google Scholar]
- Dente L., Cesareni G., Cortese R. pEMBL: a new family of single stranded plasmids. Nucleic Acids Res. 1983 Mar 25;11(6):1645–1655. doi: 10.1093/nar/11.6.1645. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Erickson-Viitanen S., DeGrado W. F. Recognition and characterization of calmodulin-binding sequences in peptides and proteins. Methods Enzymol. 1987;139:455–478. doi: 10.1016/0076-6879(87)39106-2. [DOI] [PubMed] [Google Scholar]
- Evans D. A., Ellis D. S., Stamford S. Ultrastructural studies of certain aspects of the development of Trypanosoma congolense in Glossina morsitans morsitans. J Protozool. 1979 Nov;26(4):557–563. doi: 10.1111/j.1550-7408.1979.tb04195.x. [DOI] [PubMed] [Google Scholar]
- Freymuller E., Camargo E. P. Ultrastructural differences between species of trypanosomatids with and without endosymbionts. J Protozool. 1981 May;28(2):175–182. doi: 10.1111/j.1550-7408.1981.tb02829.x. [DOI] [PubMed] [Google Scholar]
- Fridovich-Keil J. L., Bond J. F., Solomon F. Domains of beta-tubulin essential for conserved functions in vivo. Mol Cell Biol. 1987 Oct;7(10):3792–3798. doi: 10.1128/mcb.7.10.3792. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Frischauf A. M., Lehrach H., Poustka A., Murray N. Lambda replacement vectors carrying polylinker sequences. J Mol Biol. 1983 Nov 15;170(4):827–842. doi: 10.1016/s0022-2836(83)80190-9. [DOI] [PubMed] [Google Scholar]
- Fuge H. Electron microscopic studies on the intra-flagellar structures of trypanosomes. J Protozool. 1969 Aug;16(3):460–466. doi: 10.1111/j.1550-7408.1969.tb02301.x. [DOI] [PubMed] [Google Scholar]
- Gallo J. M., Schrével J. Homologies between paraflagellar rod proteins from trypanosomes and euglenoids revealed by a monoclonal antibody. Eur J Cell Biol. 1985 Mar;36(2):163–168. [PubMed] [Google Scholar]
- Garnier J., Osguthorpe D. J., Robson B. Analysis of the accuracy and implications of simple methods for predicting the secondary structure of globular proteins. J Mol Biol. 1978 Mar 25;120(1):97–120. doi: 10.1016/0022-2836(78)90297-8. [DOI] [PubMed] [Google Scholar]
- Geisler N., Weber K. Comparison of the proteins of two immunologically distinct intermediate-sized filaments by amino acid sequence analysis: desmin and vimentin. Proc Natl Acad Sci U S A. 1981 Jul;78(7):4120–4123. doi: 10.1073/pnas.78.7.4120. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gibson W. C., Osinga K. A., Michels P. A., Borst P. Trypanosomes of subgenus Trypanozoon are diploid for housekeeping genes. Mol Biochem Parasitol. 1985 Sep;16(3):231–242. doi: 10.1016/0166-6851(85)90066-0. [DOI] [PubMed] [Google Scholar]
- Goldstein L. S., Laymon R. A., McIntosh J. R. A microtubule-associated protein in Drosophila melanogaster: identification, characterization, and isolation of coding sequences. J Cell Biol. 1986 Jun;102(6):2076–2087. doi: 10.1083/jcb.102.6.2076. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Goldstein S. F., Holwill M. E., Silvester N. R. The effects of laser microbeam irradiation on the flagellum of Crithidia (Strigomonas) oncopelti. J Exp Biol. 1970 Oct;53(2):401–409. doi: 10.1242/jeb.53.2.401. [DOI] [PubMed] [Google Scholar]
- Hanukoglu I., Fuchs E. The cDNA sequence of a human epidermal keratin: divergence of sequence but conservation of structure among intermediate filament proteins. Cell. 1982 Nov;31(1):243–252. doi: 10.1016/0092-8674(82)90424-x. [DOI] [PubMed] [Google Scholar]
- Hyams J. S. The Euglena paraflagellar rod: structure, relationship to other flagellar components and preliminary biochemical characterization. J Cell Sci. 1982 Jun;55:199–210. doi: 10.1242/jcs.55.1.199. [DOI] [PubMed] [Google Scholar]
- Imboden M. A., Laird P. W., Affolter M., Seebeck T. Transcription of the intergenic regions of the tubulin gene cluster of Trypanosoma brucei: evidence for a polycistronic transcription unit in a eukaryote. Nucleic Acids Res. 1987 Sep 25;15(18):7357–7368. doi: 10.1093/nar/15.18.7357. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Imboden M., Blum B., DeLange T., Braun R., Seebeck T. Tubulin mRNAs of Trypanosoma brucei. J Mol Biol. 1986 Apr 5;188(3):393–402. doi: 10.1016/0022-2836(86)90163-4. [DOI] [PubMed] [Google Scholar]
- Kimmel B. E., Samson S., Wu J., Hirschberg R., Yarbrough L. R. Tubulin genes of the African trypanosome Trypanosoma brucei rhodesiense:nucleotide sequence of a 3.7-kb fragment containing genes for alpha and beta tubulins. Gene. 1985;35(3):237–248. doi: 10.1016/0378-1119(85)90002-2. [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]
- McLachlan A. D. Structural implications of the myosin amino acid sequence. Annu Rev Biophys Bioeng. 1984;13:167–189. doi: 10.1146/annurev.bb.13.060184.001123. [DOI] [PubMed] [Google Scholar]
- O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
- Parnes J. R., Velan B., Felsenfeld A., Ramanathan L., Ferrini U., Appella E., Seidman J. G. Mouse beta 2-microglobulin cDNA clones: a screening procedure for cDNA clones corresponding to rare mRNAs. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2253–2257. doi: 10.1073/pnas.78.4.2253. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Parsons M., Nelson R. G., Watkins K. P., Agabian N. Trypanosome mRNAs share a common 5' spliced leader sequence. Cell. 1984 Aug;38(1):309–316. doi: 10.1016/0092-8674(84)90552-X. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Piccinni E. ATPase activity in flagella from Euglena gracilis. Localization of the enzyme and effects of detergents. J Protozool. 1975 Aug;22(3):331–335. doi: 10.1111/j.1550-7408.1975.tb05182.x. [DOI] [PubMed] [Google Scholar]
- Rudolph J. E., Kimble M., Hoyle H. D., Subler M. A., Raff E. C. Three Drosophila beta-tubulin sequences: a developmentally regulated isoform (beta 3), the testis-specific isoform (beta 2), and an assembly-defective mutation of the testis-specific isoform (B2t8) reveal both an ancient divergence in metazoan isotypes and structural constraints for beta-tubulin function. Mol Cell Biol. 1987 Jun;7(6):2231–2242. doi: 10.1128/mcb.7.6.2231. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Russell D. G., Newsam R. J., Palmer G. C., Gull K. Structural and biochemical characterisation of the paraflagellar rod of Crithidia fasciculata. Eur J Cell Biol. 1983 Mar;30(1):137–143. [PubMed] [Google Scholar]
- Rüegg U. T., Rudinger J. Reductive cleavage of cystine disulfides with tributylphosphine. Methods Enzymol. 1977;47:111–116. doi: 10.1016/0076-6879(77)47012-5. [DOI] [PubMed] [Google Scholar]
- Saborio J. L., Manuel Hernandez J., Narayanswami S., Wrightsman R., Palmer E., Manning J. Isolation and characterization of paraflagellar proteins from Trypanosoma cruzi. J Biol Chem. 1989 Mar 5;264(7):4071–4075. [PubMed] [Google Scholar]
- Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schneider A., Lutz H. U., Marugg R., Gehr P., Seebeck T. Spectrin-like proteins in the paraflagellar rod structure of Trypanosoma brucei. J Cell Sci. 1988 Jun;90(Pt 2):307–315. doi: 10.1242/jcs.90.2.307. [DOI] [PubMed] [Google Scholar]
- Schneider A., Sherwin T., Sasse R., Russell D. G., Gull K., Seebeck T. Subpellicular and flagellar microtubules of Trypanosoma brucei brucei contain the same alpha-tubulin isoforms. J Cell Biol. 1987 Mar;104(3):431–438. doi: 10.1083/jcb.104.3.431. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Seebeck T., Whittaker P. A., Imboden M. A., Hardman N., Braun R. Tubulin genes of Trypanosoma brucei: a tightly clustered family of alternating genes. Proc Natl Acad Sci U S A. 1983 Aug;80(15):4634–4638. doi: 10.1073/pnas.80.15.4634. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Smith G. E., Summers M. D. The bidirectional transfer of DNA and RNA to nitrocellulose or diazobenzyloxymethyl-paper. Anal Biochem. 1980 Nov 15;109(1):123–129. doi: 10.1016/0003-2697(80)90019-6. [DOI] [PubMed] [Google Scholar]
- Sogin M. L., Elwood H. J., Gunderson J. H. Evolutionary diversity of eukaryotic small-subunit rRNA genes. Proc Natl Acad Sci U S A. 1986 Mar;83(5):1383–1387. doi: 10.1073/pnas.83.5.1383. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Souto-Padrón T., de Souza W., Heuser J. E. Quick-freeze, deep-etch rotary replication of Trypanosoma cruzi and Herpetomonas megaseliae. J Cell Sci. 1984 Jul;69:167–178. doi: 10.1242/jcs.69.1.167. [DOI] [PubMed] [Google Scholar]
- Steinert P. M., Steven A. C., Roop D. R. The molecular biology of intermediate filaments. Cell. 1985 Sep;42(2):411–420. doi: 10.1016/0092-8674(85)90098-4. [DOI] [PubMed] [Google Scholar]
- Sundaralingam M., Drendel W., Greaser M. Stabilization of the long central helix of troponin C by intrahelical salt bridges between charged amino acid side chains. Proc Natl Acad Sci U S A. 1985 Dec;82(23):7944–7947. doi: 10.1073/pnas.82.23.7944. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wessel D., Flügge U. I. A method for the quantitative recovery of protein in dilute solution in the presence of detergents and lipids. Anal Biochem. 1984 Apr;138(1):141–143. doi: 10.1016/0003-2697(84)90782-6. [DOI] [PubMed] [Google Scholar]
- Young R. A., Davis R. W. Efficient isolation of genes by using antibody probes. Proc Natl Acad Sci U S A. 1983 Mar;80(5):1194–1198. doi: 10.1073/pnas.80.5.1194. [DOI] [PMC free article] [PubMed] [Google Scholar]
- de Souza W., Souto-Padrón T. The paraxial structure of the flagellum of trypanosomatidae. J Parasitol. 1980 Apr;66(2):229–236. [PubMed] [Google Scholar]