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. 1990 May;10(5):2302–2307. doi: 10.1128/mcb.10.5.2302

The chicken lysozyme 5' matrix attachment region increases transcription from a heterologous promoter in heterologous cells and dampens position effects on the expression of transfected genes.

L Phi-Van 1, J P von Kries 1, W Ostertag 1, W H Strätling 1
PMCID: PMC360577  PMID: 2325653

Abstract

Matrix attachment regions (MARs) are DNA elements that dissect the genome into topologically separated domains by binding to a chromosomal skeleton. This study explored the putative influence of the MAR located 5' of the chicken lysozyme gene on expression of heterologous genes in heterologous cell systems. Expression of a construct with the chloramphenicol acetyltransferase (CAT) indicator gene controlled by the herpes simplex virus thymidine kinase promoter (TC) and a construct in which the same transcriptional unit is flanked by chicken lysozyme 5' MARs (MTCM) was assayed after stable transfection into rat fibroblasts. Median CAT activity per copy number in MTCM transfectants was elevated approximately 10-fold relative to that in TC transfectants. Total variation in normalized CAT activity decreased from more than 100-fold among TC transfectants to nearly 6-fold among MTCM transfectants. The steady-state level of transcripts and the relative rate of transcription were increased in MTCM transfectants, as shown by S1 nuclease and run-on transcription assays, respectively. The chicken lysozyme 5' MAR thus can confer elevated, less position-dependent expression on a heterologous promoter in cells of a different species by increasing the density of transcribing RNA polymerase molecules. MAR-mediated transcriptional enhancement suggests that MARs are important for gene expression and not just for DNA packaging.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Benyajati C., Worcel A. Isolation, characterization, and structure of the folded interphase genome of Drosophila melanogaster. Cell. 1976 Nov;9(3):393–407. doi: 10.1016/0092-8674(76)90084-2. [DOI] [PubMed] [Google Scholar]
  2. Berezney R., Coffey D. S. Identification of a nuclear protein matrix. Biochem Biophys Res Commun. 1974 Oct 23;60(4):1410–1417. doi: 10.1016/0006-291x(74)90355-6. [DOI] [PubMed] [Google Scholar]
  3. Blom van Assendelft G., Hanscombe O., Grosveld F., Greaves D. R. The beta-globin dominant control region activates homologous and heterologous promoters in a tissue-specific manner. Cell. 1989 Mar 24;56(6):969–977. doi: 10.1016/0092-8674(89)90630-2. [DOI] [PubMed] [Google Scholar]
  4. Bode J., Maass K. Chromatin domain surrounding the human interferon-beta gene as defined by scaffold-attached regions. Biochemistry. 1988 Jun 28;27(13):4706–4711. doi: 10.1021/bi00413a019. [DOI] [PubMed] [Google Scholar]
  5. Canaani D., Berg P. Regulated expression of human interferon beta 1 gene after transduction into cultured mouse and rabbit cells. Proc Natl Acad Sci U S A. 1982 Sep;79(17):5166–5170. doi: 10.1073/pnas.79.17.5166. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  7. Cockerill P. N., Garrard W. T. Chromosomal loop anchorage of the kappa immunoglobulin gene occurs next to the enhancer in a region containing topoisomerase II sites. Cell. 1986 Jan 31;44(2):273–282. doi: 10.1016/0092-8674(86)90761-0. [DOI] [PubMed] [Google Scholar]
  8. Cockerill P. N., Garrard W. T. Chromosomal loop anchorage sites appear to be evolutionarily conserved. FEBS Lett. 1986 Aug 11;204(1):5–7. doi: 10.1016/0014-5793(86)81377-1. [DOI] [PubMed] [Google Scholar]
  9. Cockerill P. N., Yuen M. H., Garrard W. T. The enhancer of the immunoglobulin heavy chain locus is flanked by presumptive chromosomal loop anchorage elements. J Biol Chem. 1987 Apr 15;262(11):5394–5397. [PubMed] [Google Scholar]
  10. Cook P. R., Brazell I. A. Spectrofluorometric measurement of the binding of ethidium to superhelical DNA from cell nuclei. Eur J Biochem. 1978 Mar 15;84(2):465–477. doi: 10.1111/j.1432-1033.1978.tb12188.x. [DOI] [PubMed] [Google Scholar]
  11. Favaloro J., Treisman R., Kamen R. Transcription maps of polyoma virus-specific RNA: analysis by two-dimensional nuclease S1 gel mapping. Methods Enzymol. 1980;65(1):718–749. doi: 10.1016/s0076-6879(80)65070-8. [DOI] [PubMed] [Google Scholar]
  12. Friel J., Stocking C., Stacey A., Ostertag W. A temperature-sensitive mutant of the myeloproliferative sarcoma virus, altered by a point mutation in the mos oncogene, has been modified as a selectable retroviral vector. J Virol. 1987 Mar;61(3):889–897. doi: 10.1128/jvi.61.3.889-897.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Fritton H. P., Igo-Kemenes T., Nowock J., Strech-Jurk U., Theisen M., Sippel A. E. Alternative sets of DNase I-hypersensitive sites characterize the various functional states of the chicken lysozyme gene. Nature. 1984 Sep 13;311(5982):163–165. doi: 10.1038/311163a0. [DOI] [PubMed] [Google Scholar]
  14. Gasser S. M., Laemmli U. K. Cohabitation of scaffold binding regions with upstream/enhancer elements of three developmentally regulated genes of D. melanogaster. Cell. 1986 Aug 15;46(4):521–530. doi: 10.1016/0092-8674(86)90877-9. [DOI] [PubMed] [Google Scholar]
  15. Hwang L. H., Gilboa E. Expression of genes introduced into cells by retroviral infection is more efficient than that of genes introduced into cells by DNA transfection. J Virol. 1984 May;50(2):417–424. doi: 10.1128/jvi.50.2.417-424.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Igó-Kemenes T., Zachau H. G. Domains in chromatin structure. Cold Spring Harb Symp Quant Biol. 1978;42(Pt 1):109–118. doi: 10.1101/sqb.1978.042.01.012. [DOI] [PubMed] [Google Scholar]
  17. Izaurralde E., Mirkovitch J., Laemmli U. K. Interaction of DNA with nuclear scaffolds in vitro. J Mol Biol. 1988 Mar 5;200(1):111–125. doi: 10.1016/0022-2836(88)90337-3. [DOI] [PubMed] [Google Scholar]
  18. Jarman A. P., Higgs D. R. Nuclear scaffold attachment sites in the human globin gene complexes. EMBO J. 1988 Nov;7(11):3337–3344. doi: 10.1002/j.1460-2075.1988.tb03205.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Käs E., Chasin L. A. Anchorage of the Chinese hamster dihydrofolate reductase gene to the nuclear scaffold occurs in an intragenic region. J Mol Biol. 1987 Dec 20;198(4):677–692. doi: 10.1016/0022-2836(87)90209-9. [DOI] [PubMed] [Google Scholar]
  20. Lebkowski J. S., Laemmli U. K. Evidence for two levels of DNA folding in histone-depleted HeLa interphase nuclei. J Mol Biol. 1982 Apr 5;156(2):309–324. doi: 10.1016/0022-2836(82)90331-x. [DOI] [PubMed] [Google Scholar]
  21. Loc P. V., Strätling W. H. The matrix attachment regions of the chicken lysozyme gene co-map with the boundaries of the chromatin domain. EMBO J. 1988 Mar;7(3):655–664. doi: 10.1002/j.1460-2075.1988.tb02860.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Luckow B., Schütz G. CAT constructions with multiple unique restriction sites for the functional analysis of eukaryotic promoters and regulatory elements. Nucleic Acids Res. 1987 Jul 10;15(13):5490–5490. doi: 10.1093/nar/15.13.5490. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Mirkovitch J., Gasser S. M., Laemmli U. K. Scaffold attachment of DNA loops in metaphase chromosomes. J Mol Biol. 1988 Mar 5;200(1):101–109. doi: 10.1016/0022-2836(88)90336-1. [DOI] [PubMed] [Google Scholar]
  24. Mirkovitch J., Mirault M. E., Laemmli U. K. Organization of the higher-order chromatin loop: specific DNA attachment sites on nuclear scaffold. Cell. 1984 Nov;39(1):223–232. doi: 10.1016/0092-8674(84)90208-3. [DOI] [PubMed] [Google Scholar]
  25. Mirkovitch J., Spierer P., Laemmli U. K. Genes and loops in 320,000 base-pairs of the Drosophila melanogaster chromosome. J Mol Biol. 1986 Jul 20;190(2):255–258. doi: 10.1016/0022-2836(86)90296-2. [DOI] [PubMed] [Google Scholar]
  26. Paulson J. R., Laemmli U. K. The structure of histone-depleted metaphase chromosomes. Cell. 1977 Nov;12(3):817–828. doi: 10.1016/0092-8674(77)90280-x. [DOI] [PubMed] [Google Scholar]
  27. Penman S. RNA metabolism in the HeLa cell nucleus. J Mol Biol. 1966 May;17(1):117–130. doi: 10.1016/s0022-2836(66)80098-0. [DOI] [PubMed] [Google Scholar]
  28. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  29. Stief A., Winter D. M., Strätling W. H., Sippel A. E. A nuclear DNA attachment element mediates elevated and position-independent gene activity. Nature. 1989 Sep 28;341(6240):343–345. doi: 10.1038/341343a0. [DOI] [PubMed] [Google Scholar]
  30. Strätling W. H., Dölle A., Sippel A. E. Chromatin structure of the chicken lysozyme gene domain as determined by chromatin fractionation and micrococcal nuclease digestion. Biochemistry. 1986 Jan 28;25(2):495–502. doi: 10.1021/bi00350a033. [DOI] [PubMed] [Google Scholar]
  31. Topp W. C. Normal rat cell lines deficient in nuclear thymidine kinase. Virology. 1981 Aug;113(1):408–411. doi: 10.1016/0042-6822(81)90168-9. [DOI] [PubMed] [Google Scholar]
  32. Weaver R. F., Weissmann C. Mapping of RNA by a modification of the Berk-Sharp procedure: the 5' termini of 15 S beta-globin mRNA precursor and mature 10 s beta-globin mRNA have identical map coordinates. Nucleic Acids Res. 1979 Nov 10;7(5):1175–1193. doi: 10.1093/nar/7.5.1175. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Weber J., Jelinek W., Darnell J. E., Jr The definition of a large viral transcription unit late in Ad2 infection of HeLa cells: mapping of nascent RNA molecules labeled in isolated nuclei. Cell. 1977 Apr;10(4):611–616. doi: 10.1016/0092-8674(77)90093-9. [DOI] [PubMed] [Google Scholar]
  34. von Kries J. P., Strätling W. H. Lysozyme gene specific transcription in isolated hen oviduct nuclei. Int J Biochem. 1988;20(6):633–637. doi: 10.1016/0020-711x(88)90103-6. [DOI] [PubMed] [Google Scholar]

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