Abstract
In order to assess the selectivity of the distribution patterns of individual nonhistone chromosomal proteins (NHC proteins), immunofluorescent staining experiments were performed on Drosophila polytene chromosomes. Antisera have been prepared against three individual NHC proteins which were isolated by sequential preparative slab gel isoelectric focusing and SDS polyacrylamide gel electrophoresis. In two cases, immunofluorescent staining of the chromosomes indicated a specific limited distribution pattern; apparently the antigen in each case is present at a reproducible and distinct subset of chromomeres. This type of pattern has also been obtained with antisera prepared against molecular weight subfractions of NHC proteins (Silver and Elgin, 1977). Each selective fluorescence distribution pattern obtained so far is reproducible and unique to the antiserum under study. In a third case, an antiserum caused prominant staining at dense chromomeres and the chromocenter in a pattern mimicking DNA (and presumably histone) distribution. Indirect radioimmunostaining of SDS and isoelectric focusing gels on which total NHC proteins had been separated confirmed that this antiserum reacted specifically with a protein(s) of molecular weight 21,000 D and pI 5.2. The data in conjunction with absorption experiments indicates that the chromosomal staining is due to an interaction of antibodies with NHC protein(s) and not with histones. This finding suggests that at least one major acidic NHC protein plays a very general role (comparable to that of the histones) in maintaining chromatin structure.
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Alfageme, C.R., Rudkin, G.T., Cohen, L.H.: Locations of chromosomal proteins in polytene chromosomes. Proc. nat. Acad. Sci. (Wash) 73, 2038–42 (1976)
Beermann, W.: Chromomeres and genes. In: Developmental studies in giant chromosomes (W. Beermann, ed.), pp. 1–33. New York: Springer 1972
Burridge, K.: Changes in cellular glycoproteins after transformation. Proc. nat. Acad. Sci. (Wash.) 73, 4457–61 (1976)
Efstratiadis, A., Kafatos, F.C.: The chorion of insects. In: Methods in molecular biology Vol. 8, (J. Last, ed.), pp. 1–41. New York: Dekker 1975
Elgin, S.C.R., Hood, L.E.: Chromosomal proteins of Drosophila embryos. Biochemistry 12, 4984–4991 (1973)
Elgin, S.C.R., Serunian, L.A., Silver, L.M.: Distribution patterns of Drosophila nonhistone chromosomal proteins. Cold Spr. Harb. Symp. quant. Biol. 42 (in press 1978)
Elgin, S.C.R., Silver, L.M., Wu, C.E.C.: The in situ distribution of Drosophila nonhistone chromosomal proteins. In: The molecular biology of the mammalian genetic apparatus (P.O.P. T'so, ed.), pp. 127–141. Amsterdam, New York: Elsevier/North Holland Biomedical Press 1977
Elgin, S.C.R., Weintraub, H.: Chromosomal proteins and chromatin structure. Ann. Rev. Biochem. 44, 725–74 (1975)
Fairbanks, G., Steck, T.L., Wallach, D.F.H.: Electrophoratic analysis of the major polypeptides of the human erythrocyte membrane. Biochemistry 10, 2606–2617 (1971)
Gonzalez-Mujica, F., Mathias, A.P.: Protein from different classes of liver nuclei in normal and thioacetamide-treated rats. Biochem. J. 133, 441–55 (1973)
Gronow, M., Griffiths, G.: Rapid isolation and separation of the non-histone proteins of rat liver nuclei. FEBS Letters 15, 340 (1971)
Hill, R.J., Watt, F.: Chromosomal proteins of Drosophila melanogaster and an approach for their localization on polytene chromosomes. Chromosoma (Berl.) 63, 57–78 (1977)
Laemmli, U.K.: Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature (Lond.) 227, 680–685 (1970)
Lea, M.A., Youngsworth, L.A., Morris, H.P.: Isoelectric focusing of non-histone nuclear proteins of Morris hepatomas and normal rat liver. Proc. Amer. Ass. Cancer Res. 15, 25 (1974)
MacGillivray, A.J.: Preparation and properties of chromatin non-histone proteins. In: Subnuclear components: Preparation and fractionation (G.D. Birnie, ed.), pp. 209–266. London: Butterworths 1976
Pardue, M.L., Gall, J.G.: Nucleic acid hybridization to the DNA of cytological preparations. Methods in Cell Biol. X, 1–16 (1975)
Pardue, M.L., Gerbi, S.A., Eckhardt, R.A., Gall, J.G.: Cytological localization of DNA complimentary to ribosomal RNA in polytene chromosomes of Diptera. Chromosoma (Berl.) 29, 268–70 (1970)
Pelling, C.: Ribonukleinsäure-Synthese der Riesenchromosomen. Autoradiographische Untersuchungen an Chironomus tentans. Chromosoma (Berl.) 15, 71–122 (1964)
Peterson, J.L., McConkey, E.H.: Non-histone chromosomal proteins from HeLa cells. A survey by high resolution, two-dimensional Electrophoresis. J. biol. Chem. 251, 548–54 (1976)
Plagens, U., Greenleaf, A.L., Bautz, E.K.F.: Distribution of RNA polymerase on Drosophila polytene chromosomes as studied by indirect immunofluorescence. Chromosoma (Berl.) 59, 157–65 (1976)
Righetti, P.G., Drysdale, J.W.: Isoelectric focusing. In: Laboratory techniques in biochemistry. and molecular biology, Vol. 5, (T.S. Work and E. Work, eds.), pp. 337–590 New York: Elsevier (1976b)
Silver, L.M.: In situ distributions of chromosomal proteins. Ph. D. Thesis, Biophysics, Harvard University (1978)
Silver, L.M., Elgin, S.C.R.: A method for the determination of the in situ distribution of chromosomal proteins. Proc. nat. Acad. Sci. (Wash.) 73, 423–7 (1976)
Silver, L.M., Elgin, S.C.R.: Distribution patterns of three subfractions of Drosophila nonhistone chromosomal proteins: Possible correlations with gene activity. Cell 11, 971–83 (1977)
Silver, L.M., Wu, C.E.C., Elgin, S.C.R.: Immunofluorescent techniques in the analysis of chromosomal proteins. In: Methods in chromosomal protein research (G. Stein, J. Stein, and L. Kleinsmith, eds.), pp. 151–167 New York: Academic Press 1978
Sonoda, S., Schlamowitz, M.: Studies of 125-I trace labeling of immunoglobin G by chloramine T. Immunochemistry 7, 885–98 (1970)
Studier, F.W.: Analysis of bacteriophage T7 early RNAs and proteins on slab gels. J. molec. Biol. 79, 237–248 (1973)
Stumpf, W.E., Elgin, S.C.R., Hood, L.: Antibodies to proteins dissolved in sodium dodecyl sulfate. J. Immunol. 113, 1752–6 (1974)
Suria, D., Liew, C.C.: Isolation and analysis of non-histone chromatin proteins from rat-liver nuclei by three different methods. Canad. J. Biochem. 52, 1143–53 (1974)
Williamson, A.R.: Antibody isoelectric spectra. Europ. J. Immunol. 1, 390–4 (1971)
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Silver, L.M., Elgin, S.C.R. Production and characterization of antisera against three individual NHC proteins; a case of a generally distributed NHC protein. Chromosoma 68, 101–114 (1978). https://doi.org/10.1007/BF00287143
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DOI: https://doi.org/10.1007/BF00287143