Skip to main content
SpringerLink
Log in

Regional differences in immunostainability of isolated metaphase chromosomes of Indian muntjac with anti-Z-DNA antibody

  • Published:
Chromosoma Aims and scope Submit manuscript

Abstract

The distribution of Z-form DNA along the length of metaphase chromosomes of Indian muntjac was studied by indirect immunofluorescence procedures using an antibody specific to the Z-DNA conformation. Several fixation conditions were compared for reproducible detection of Z-DNA in isolated metaphase chromosomes. Fixation of chromosomes with 45% acetic acid alone gave reproducible reactivity with the antibody. When fixation was done either with Carnoy's solution (3:1 methanol:acetic acid) or with 75% alcohol alone, the antibody binding was at background level. Acetic acid-fixed chromosomes exhibited intense fluorescence both at C-band heterochromatin and at nucleolus organizer regions (NORs). The euchromatic regions had weakly, but clearly, stained bands, which were quite similar to the chromomycin A3 R-bands. After treatment with topoisomerase I, the immunofluorescence at NORs and R-bands disappeared, but only a slight decrease in immunofluorescence intensity was observed at C-band regions. We suggest that this difference in the immunoreactivity of NORs and R-bands from C-bands reflects a difference in gene activity among these regions. Possible molecular mechanisms involved in Z-DNA immunoreactivity are discussed, based on SDS-polyacrylamide gel electrophoretic analysis of chromosomal proteins after extraction of metaphase chromosomes with different fixative solutions.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

Abbreviations

PI :

propidium iodide

NOR(s) :

nucleolus organizer region(s)

SDS-PAGE :

sodium dodecyl sulfate polyacrylamide gel electrophoresis

References

  • Arndt-Jovin DJ, Robert-Nicoud M, Zarling DA, Greider C, Weimer E, Jovin TM (1983) Left-handed Z-DNA in bands of acid-fixed polytene chromosomes. Proc Natl Acad Sci USA 80:4344–4348

    Google Scholar 

  • Brat SV, Verma RS, Dosik H (1979) Structural organization of chromosomes of the indian muntjac (Muntiacus muntjak). Cytogenet Cell Genet 24:201–208

    Google Scholar 

  • Burkholder GD, Latimer LJP, Lee JS (1988) Immunofluorescent staining of mammalian nuclei and chromosomes with a monoclonal antibody to triplex DNA. Chromosoma 97:185–192

    Google Scholar 

  • Comings DE (1978) Mechanisms of chromosome banding and implications for chromosome structure. Annu Rev Genet 12:25–46

    Google Scholar 

  • Hamada H, Kakunaga T (1982) Potential Z-DNA forming sequences are highly dispersed in the human genome. Nature 298:396–398

    Google Scholar 

  • Haniford DB, Pulleyblank DE (1983) The in vivo occurence of Z-DNA. J Biomol Struct Dynam 1:593–609

    Google Scholar 

  • Hill RJ, Stollar BD (1983) Dependence of Z-DNA antibody binding to polytene chromosomes on acid fixation and DNA torsional strain. Nature 305:338–340

    Google Scholar 

  • Irie S, Sezaki M (1983) A quantitative determination of the relative amount of histones in polyacrylamide gel by silver stain. Anal Biochem 134:471–478

    Google Scholar 

  • Jimenez-Ruiz A, Requena JM, Lancillotti F, Morales G, Lopez MC, Alonso C (1989) Molecular cloning of a Drosophila potential Z-DNA forming sequence hybridizing in situ to a developmentally regulated subdivision of the polytene chromosomes. Nucleic Acids Res 17:4579–4588

    Google Scholar 

  • Kmiec EB, Angelides KJ, Holloman WK (1985) Left-handed DNA and the synaptic pairing reaction promoted by Ustilago rec1 protein. Cell 40:139–145

    Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685

    Google Scholar 

  • Lafer E, Sausa R, Rich A (1985) Anti-Z-DNA antibody binding can stabilize Z-DNA in relaxed and linear plasmids under physiological conditions. EMBO J 4:3655–3660

    Google Scholar 

  • Lancillotti F, Lopez MC, Alonso C, Stollar BD (1985) Locations of Z-DNA in polytene chromosomes. J Cell Biol 100:1759–1766

    Google Scholar 

  • Lancillotti F, Lopez MC, Arias P, Alonso C (1987) Z-DNA in transcriptionally active chromosomes. Proc Natl Acad Sci USA 84:1560–1564

    Google Scholar 

  • Lemeunier F, Derbin C, Malfoy B, Leng M, Taillandier E (1982) Identification of left-handed Z-DNA by indirect immunofluorescence in polytene chromosomes ofChironomus thummi thummi. Exp Cell Res 141:508–513

    Google Scholar 

  • Lipps HJ, Nordheim A, Lafer EM, Ammermann D, Stollar BD, Rich A (1983) Antibodies against Z DNA react with the macronucleus but not the micronucleus of the hypotrichous ciliateStylonychia mytilus. Cell 32:435–441

    Google Scholar 

  • Morgenegg G, Celio MR, Malfoy B, Leng M, Kuenzle CC (1983) Z-DNA immunoreactivity in rat tissues. Nature 303:540–543

    Google Scholar 

  • Nordheim A, Rich A (1983) Negatively supercoiled simian virus 40 DNA contains Z-DNA segments within transcriptional enhancer sequences. Nature 303:674–679

    Google Scholar 

  • Nordheim A, Pardue ML, Lafer EM, Moller A, Stollar BD, Rich A (1981) Antibodies to left-handed Z-DNA bind to interband regions ofDrosophila polytene chromosomes. Nature 294:417–422

    Google Scholar 

  • Nordheim A, Tesser P, Azorin F, Kwon YH, Moller A, Rich A (1982) Isolation ofDrosophila proteins that bind selectively to left-handed Z-DNA. Proc Natl Acad Sci USA 79:7729–7733

    Google Scholar 

  • Pardue ML, Hsu TC (1975) Locations of 18S and 28S ribosomal genes on the chromosomes of the Indian muntjac. J Cell Biol 64:251–254

    Google Scholar 

  • Pardue ML, Lowenhaupt K, Rich A, Nordheim A (1987) (dC-dA)n·(dG-dT)n sequences have evolutionarily conserved chromosomal locations in Drosophila with implications for roles in chromosome structure and function. EMBO J 6:1781–1789

    Google Scholar 

  • Pulleyblank DE, Haniford DB, Morgan AR (1985) A structural basis for S1 nuclease sensitivity of double-stranded DNA. Cell 42:271–280

    Google Scholar 

  • Rich A, Nordheim A, Wang AHJ (1984) The chemistry and biology of left-handed Z-DNA. Annu Rev Biochem 53:791–846

    Google Scholar 

  • Robert-Nicoud M, Arndt-Jovin DJ, Zarling DA, Jovin TM (1984) Immunological detection of left-handed Z DNA in isolated polytene chromosomes. Effects of ionic strength, pH, temperature and topological stress. EMBO J 3:721–731

    Google Scholar 

  • Ueda T, Irie S, Kato Y (1987) Longitudinal differentiation of metaphase chromosomes of Indian muntjac as studied by restriction enzyme digestion, in situ hybridization with cloned DNA probes and distamycin A plus DAPI fluorescence staining. Chromosoma 95:251–257

    Google Scholar 

  • Viegas-Pequignot E, Derbin C, Malfoy B, Taillandier E, Leng M, Dutrillaux B (1983) Z-DNA immunoreactivity in fixed metaphase chromosomes of primates. Proc Natl Acad Sci USA 80:5890–5894

    Google Scholar 

  • Wang AH-J, Quigley GJ, Kolpak FJ, Crawford JL, van Boom JH, van der Marel G, Rich A (1979) Molecular structure of a left-handed double helical DNA fragment at atomic resolution. Nature 282:680–686

    Google Scholar 

  • Weinreb A, Collier D, Birshtein BK, Wells RD (1988) Intramolecular triplex and left-handed Z-DNA at a site of unequal sister chromatid exchange. J Cell Biol 107:p90a

    Google Scholar 

  • Wittig B, Dorbic T, Rich A (1989) The level of Z-DNA in metabolically active, permealized mammalian cell nuclei is regulated by torsional strain. J Cell Biol 108:755–764

    Google Scholar 

  • Wray W, Stubblefield E (1970) A new method for the rapid isolation of chromosomes, mitotic apparatus, or nuclei from mammalian fibroblasts at neutral pH. Exp Cell Res 59:469–478

    Google Scholar 

  • Zarling DA, Arndt-Jobin DJ, Robert-Nicoud M, McIntosh LP, Thomae R, Jovin TM (1984) Immunoglobulin recognition of synthetic and natural left-handed Z-DNA conformations and sequences. J Mol Biol 176:369–377

    Google Scholar 

  • Zimmerman SB (1982) The three-dimensional structure of DNA. Annu Rev Biochem 51:395–427

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Laboratory of Developmental Biology, Mitsubishi-Kasei Institute of Life Sciences, 11 Minamiooya, Machida-shi, 194, Tokyo, Japan

    Takayuki Ueda, Yoshihiro Kato & Shinkichi Irie

Authors
  1. Takayuki Ueda
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yoshihiro Kato
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Shinkichi Irie
    View author publications

    You can also search for this author in PubMed Google Scholar

Additional information

Deceased, April 23, 1988

Rights and permissions

Reprints and permissions

About this article

Cite this article

Ueda, T., Kato, Y. & Irie, S. Regional differences in immunostainability of isolated metaphase chromosomes of Indian muntjac with anti-Z-DNA antibody. Chromosoma 99, 161–168 (1990). https://doi.org/10.1007/BF01731126

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF01731126

Keywords

Search

Navigation