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Characterization of thermosensitive mutants of Physarum polycephalum

Plasmodial screening methods for cell cycle mutants defective in late G2, mitosis or S phase

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Summary

Amoebal thermosensitive mutants of Physarum polycephalum have been isolated after mutagenesis of the amoebal form by nitrosoguanidine treatment. About 70% of the independent thermosensitive amoebal mutants obtained were also thermosensitive in the plasmodial form. Two basic screening methods were applied at the same time to thermosensitive microplasmodia in order to detect strains defective in premitotic events, mitosis or chromosomal DNA synthesis. The first method consists in the determination of increase in protein. RNA and DNA with incubation time at the non-permissive temperature. It allowed the detection of four independent thermosensitive mutant strains, showing an early arrest in DNA synthesis. The second one is the quantification of the variations of the different nuclear types at the restrictive temperature. Two mutant strains presented very large nuclei, uni- or multinucleolate, very similar to those obtained after methyl benzimidazole carbamate treatment, suggesting a defect in one of the mitotic processes. One of these two mutant strains showed an early arrest in DNA synthesis at the restrictive temperature. These two screening procedures were completed by electron microscopic observation. This technique allowed the detection of intra-nuclear macrotubular crystal-like structures in a thermosensitive mutant showing a reduced DNA synthesis at the non-permissive temperature.

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References

  • Aldrich, H.C.: The ultrastructure of mitosis in Myxamoebae and plasmodia of Physarum flavicomum. Amer. J. Bot. 56, 290–299 (1969)

    Google Scholar 

  • Anderson, R.W., Cooke, D.J., Dee, J.: Apogamic development of plasmodia in the Myxomycete Physarum polycephalum: a cinematographic analysis. Protoplasma 89, 29–40 (1976)

    Google Scholar 

  • Birch, B., Turnock, G.: The uptake and metabolism of uridine by the slime mould Physarum polycephalum. Europ. J. Biochem. 69, 257–263 (1976)

    Google Scholar 

  • Bradbury, E.M., Inglis, R.J., Matthews, H.R., Sarner, N.: Phosphorylation of very lysine rich histone in Physarum polycephalum. Correlation with chromosome condensation. Europ. J. Biochem. 33, 131–139 (1973)

    Google Scholar 

  • Braun, R., Mittermayer, C., Rusch, H.P.: Sequential temporal replication of DNA in Physarum polycephalum. Proc. nat. Acad. Sci. (Wash.) 53, 924–931 (1965)

    Google Scholar 

  • Brewer, E.N.: DNA replication by a possible continuous-discontinuous mechanism in homogenates of Physarum polycephalum containing dextran. Biochim. biophys. Acta (Amst.) 402, 363–371 (1975)

    Google Scholar 

  • Brewer, E.N., Rusch, H.P.: DNA synthesis by isolated nuclei of Physarum polycephalum. Biochem. biophys. Res. Commun. 21, 235–241 (1965)

    Google Scholar 

  • Brewer, E.N., Rusch, H.P.: Effect of elevated temperature shocks on mitosis and on the initiation of DNA replication in Physarum polycephalum. Exp. Cell Res. 49, 79–86 (1968)

    Google Scholar 

  • Carlile, M.J.: The lethal interaction following plasmodial fusion between two strains of the Myxomycete Physarum polycephalum. J. gen. Microbiol. 71, 581–590 (1972)

    Google Scholar 

  • Cooke, D.J., Dee, J.: Plasmodium formation without change in nuclear DNA content in Physarum polycephalum. Genet. Res. Camb. 23, 307–317 (1974)

    Google Scholar 

  • Cooke, D.J., Dee, J.: Methods for the isolation and analysis of plasmodial mutants in Physarum polycephalum. Genet. Res. Camb. 24, 175–187 (1975)

    Google Scholar 

  • Daniel, J.W., Baldwin, H.H.: Methods of culture for plasmodial Myxomycetes. In: Methods in cell physiology, (Prescott, D.M., ed.) Vol 1, pp 9–41. New York: Academic Press 1964

    Google Scholar 

  • Daniel, J.W., Rusch, H.P.: The pure culture of Physarum polycephalum on a partially defined soluble medium. J. gen. Microbiol. 25, 47–59 (1961)

    Google Scholar 

  • Dee, J.: A gene unlinked to mating type affecting crossing between strains of Physarum polycephalum. Genet. Res. Camb. 31, 85–92 (1978)

    Google Scholar 

  • Dee, J., Wheals, A.E., Holt, C.E: Inheritance of plasmodial valine requirement in Physarum polycephalum. Genet. Res. Camb. 21, 87–101 (1973)

    Google Scholar 

  • Funderud, S., Haugli, F.: DNA replication in Physarum polycephalum: characterization of replication products made in isolated nuclei. Biochem. biophys. Res. Commun. 74, 941–948 (1977)

    Google Scholar 

  • Giles, K.W., Myers, A.: An improved diphenylamine method for the estimation of deoxyribonucleic acid. Nature (Lond.) 206, 93 (1965)

    Google Scholar 

  • Gingold, E.C., Grant, W.D., Wheals, A.E., Wren, M.: Temperature sensitive mutants of the slime mould Physarum polycephalum. II mutants of the plasmodial phase. Molec. gen. Genet. 149, 115–119 (1976)

    Google Scholar 

  • Goodman, E.M., Ritter, H.: Plasmodial mitosis in Physarum polycephalum. Arch. Protistenk. III, 161–169 (1969)

    Google Scholar 

  • Gorman, J.A., Dove, W.F.: A method of indirect mutant selection in Physarum polycephalum using the antibiotic netropsin. Molec. gen. Genet. 133, 345–351 (1974)

    Google Scholar 

  • Gull, K., Trinci, A.P.J.: Ultrastructural effects of griseofulvin on the Myxomycete Physarum polycephalum. Protoplasma 81, 37–48 (1974)

    Google Scholar 

  • Guttes, E., Guttes, S.: Mitotic synchrony in the plasmodia of Physarum polycephalum and mitotic synchronization by coalescence of microplasmodia. In: Methods in cell physiology (Prescott, D.M., ed.) Vol. 1, pp. 43–54. New York: Academic Press 1964

    Google Scholar 

  • Guttes, S., Guttes, E., Ellis, R.A.: Electron microscope study of mitosis in Physarum polycephalum. J Ultrastruct. Res. 22, 508–529 (1968)

    Google Scholar 

  • Hardie, D.G., Matthews, H.R., Bradbury, E.M.: Cell cycle dependence of two nuclear histone kinase enzyme activities. Europ. J. Biochem. 66, 37–42 (1976)

    Google Scholar 

  • Hartwell, L.H.: Saccharomyces cerevisiae cell cycle. Bact. Rev. 38, 164–198 (1974)

    Google Scholar 

  • Hartwell, L.H., Culotti, J., Pringle, J.R., Reid B.J.: Genetic control of the cell division cycle in Yeast. Science 183, 46–51 (1974)

    Google Scholar 

  • Haugli, F.B., Dove, W.F.: Mutagenesis and mutant selection in Physarum polycephalum. Molec. gen. Genet. 118, 109–124 (1972)

    Google Scholar 

  • Howard, A., Pelc, S.R.: Synthesis of deoxyribonucleic acid in normal and irradiated cells and its relation to chromosome breakage. Heredity, Suppl. 6, 261–273 (1953)

    Google Scholar 

  • Howard, F.L.: Nuclear division in Plasmodia of Physarum. Ann. Bot. 46, 461–477 (1932)

    Google Scholar 

  • Kohiyama, M., Cousin, D., Ryter, A., Jacob, F.: Mutants thermosensibles d'Escherichia coli K12. Isolement et caractérisation rapide. An Inst. Pasteur, 110, 465–486 (1966)

    Google Scholar 

  • Laane, M.M., Haugli, F.B.: Division centers in mitotic nuclei of Physarum polycephalum Plasmodia. Norw. J. Bot. 21, 309–318 (1974)

    Google Scholar 

  • Le Pecq, J.B., Paoletti, C.: A new fluorometric method for RNA and DNA determination. Anal. Bioch. 17, 100–107 (1966)

    Google Scholar 

  • Lovely, J.R., Threlfall, R.J.: Fluctuations in cyclic adenosine 3′:5′-monophosphate and cyclic guanosine 3′:5-monophosphate during the mitotic cycle of the acellular slime mould Physarum polycephalum. Biochem. biophys. Res. Commun. 71, 789–795 (1976)

    Google Scholar 

  • Mejbaum, W.: Über die Bestimmung kleiner Pentosemengen, insbesondere in Derivaten der Adenylsäure. Hoppe-Seylers Z. physiol. Chem. 256, 117–120 (1939)

    Google Scholar 

  • Mohberg, J., Rusch, H.P.: Isolation and DNA content of nucleic of Physarum polycephalum. Exp. Cell Res. 66, 305–316 (1971)

    Google Scholar 

  • Nygaard, O.F., Guttes, S., Rusch, H.P.: Nucleic acid metabolism in a slime mold with synchronous mitosis. Biochim. biophys. Acta (Amst.) 38, 298–306 (1960)

    Google Scholar 

  • Rusch, H.P.: Some biochemical events in the life cycle of Physarum polycephalum. In: Advances in cell biology (D.M. Prescott, L. Goldstein, E. McConkey, eds., Vol. 1, pp. 297–327, New York: Appleton-Century-Crofts 1970

    Google Scholar 

  • Schiebel, W.: The Cell Cycle of Physarum polycephalum. Ber. dtsch. Bot. Ges. 86, S. 11–38 (1973)

    Google Scholar 

  • Waring, M.J.: Complex formation between ethidium bromide and nucleic acids. J. molec. Biol. 13, 269–282 (1965)

    Google Scholar 

  • Wickner, S., Berkower, I., Wright, M., Hurwitz, J.: Studies on in vitro DNA synthesis. IV. Purification of dna C gene product containing dna D activity from Escherichia coli. Proc. nat. Acad. Sci. (Wash.) 70, 2369–2373 (1973)

    Google Scholar 

  • Wickner, S., Wright, M., Hurwitz, J.: Studies on in vitro DNA synthesis. III. Purification of the DNA G gene product from Escherichia coli. Proc. nat. Acad. Sci. (Wash.) 70, 1613–1618 (1973)

    Google Scholar 

  • Wheals, A.E.: A homothallic strain of the Myxomycete Physarum polycephalum. Genetics 66, 623–633 (1970)

    Google Scholar 

  • Wheals, A.E., Grant, W.D., Jockush, B.M.: Temperature sensitive mutants of the slime mould Physarum polycephalum. I. Mutants of the amoebal phase. Molec. gen. Genet. 149, 111–114 (1976)

    Google Scholar 

  • Wright, M., Moisand, A., Tollon, Y., Oustrin, M.L.: Mise en évidence de l'action du méthyl benzimidazole 2 yl carbamate (MBC) et du methyl [5(2 thiényl carbonyl) 1 H benzimidazole 2 yl carbamate] (R17 934) sur le noyau de Physarym polycephalum (Myxomycétes). C.R. Acad. Sci. (Paris) 283, 1361–1364 (1976)

    Google Scholar 

  • Wright M., Tollon, Y.: Heat sensitive factor necessary for mitosis onset in Physarum polycephalum. Molec. gen. Genet. (in press)

  • Wright, M., Wickner, S., Hurwitz, J.: Studies on in vitro DNA synthesis. V. Isolation of DNA B gene product from Escherichia coli. Proc. nat. Acad. Sci. (Wash.) 70, 3120–3124 (1973)

    Google Scholar 

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Communicated by F. Kaudewitz

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Del Castillo, L., Oustrin, M.L. & Wright, M. Characterization of thermosensitive mutants of Physarum polycephalum . Molec. Gen. Genet. 164, 145–154 (1978). https://doi.org/10.1007/BF00267379

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