Abstract
The activity of hemimethylated herpes simplex virus thymidine kinase DNA and chromatin was analyzed by microinjection and thymidine incorporation into the DNA of thymidine kinase-negative Rat2 cells. Hemimethylated DNA was obtained by in vitro replication of single-stranded M13 DNA constructs and of chromatin produced by in vitro reconstitution of the DNA with purified chicken histone octamers. We found that methylation of either the coding or the noncoding DNA strand was sufficient to block expression of the hemimethylated chromatin. In contrast, the hemimethylated DNA was as active as the unmethylated control DNA after microinjection until chromatin formation occurred in the recipient cells. Microinjection of chromatin hemimethylated by bacterial Hae III methyltransferase excluded the possibility that inactivation was caused by symmetrical methylation of the injected molecules.
Full text
PDF




Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Bender T. P., Thompson C. B., Kuehl W. M. Differential expression of c-myb mRNA in murine B lymphomas by a block to transcription elongation. Science. 1987 Sep 18;237(4821):1473–1476. doi: 10.1126/science.3498214. [DOI] [PubMed] [Google Scholar]
- Bentley D. L., Groudine M. A block to elongation is largely responsible for decreased transcription of c-myc in differentiated HL60 cells. Nature. 1986 Jun 12;321(6071):702–706. doi: 10.1038/321702a0. [DOI] [PubMed] [Google Scholar]
- Bestor T. H., Hellewell S. B., Ingram V. M. Differentiation of two mouse cell lines is associated with hypomethylation of their genomes. Mol Cell Biol. 1984 Sep;4(9):1800–1806. doi: 10.1128/mcb.4.9.1800. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bestor T. H., Ingram V. M. Two DNA methyltransferases from murine erythroleukemia cells: purification, sequence specificity, and mode of interaction with DNA. Proc Natl Acad Sci U S A. 1983 Sep;80(18):5559–5563. doi: 10.1073/pnas.80.18.5559. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bird A. P. DNA methylation--how important in gene control? Nature. 1984 Feb 9;307(5951):503–504. doi: 10.1038/307503a0. [DOI] [PubMed] [Google Scholar]
- Buschhausen G., Graessmann M., Graessmann A. Inhibition of herpes simplex thymidine kinase gene expression by DNA methylation is an indirect effect. Nucleic Acids Res. 1985 Aug 12;13(15):5503–5513. doi: 10.1093/nar/13.15.5503. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Buschhausen G., Wittig B., Graessmann M., Graessmann A. Chromatin structure is required to block transcription of the methylated herpes simplex virus thymidine kinase gene. Proc Natl Acad Sci U S A. 1987 Mar;84(5):1177–1181. doi: 10.1073/pnas.84.5.1177. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Doerfler W. DNA methylation and gene activity. Annu Rev Biochem. 1983;52:93–124. doi: 10.1146/annurev.bi.52.070183.000521. [DOI] [PubMed] [Google Scholar]
- Eick D., Bornkamm G. W. Transcriptional arrest within the first exon is a fast control mechanism in c-myc gene expression. Nucleic Acids Res. 1986 Nov 11;14(21):8331–8346. doi: 10.1093/nar/14.21.8331. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Felsenfeld G., Nickol J., Behe M., McGhee J., Jackson D. Methylation and chromatin structure. Cold Spring Harb Symp Quant Biol. 1983;47(Pt 2):577–584. doi: 10.1101/sqb.1983.047.01.068. [DOI] [PubMed] [Google Scholar]
- Graessmann A., Bumke-Vogt C., Buschhausen G., Bauer M., Graessmann M. SV40 chromatin structure is not essential for viral gene expression. FEBS Lett. 1985 Jan 1;179(1):41–45. doi: 10.1016/0014-5793(85)80187-3. [DOI] [PubMed] [Google Scholar]
- Graessmann A., Graessmann M. DNA methylation, chromatin structure and regulation of Herpes simplex virus tk gene expression. Gene. 1988 Dec 25;74(1):135–137. doi: 10.1016/0378-1119(88)90269-7. [DOI] [PubMed] [Google Scholar]
- Graessmann M., Graessmann A. Microinjection of tissue culture cells. Methods Enzymol. 1983;101:482–492. doi: 10.1016/0076-6879(83)01033-2. [DOI] [PubMed] [Google Scholar]
- Graessmann M., Graessmann A., Wagner H., Werner E., Simon D. Complete DNA methylation does not prevent polyoma and simian virus 40 virus early gene expression. Proc Natl Acad Sci U S A. 1983 Nov;80(21):6470–6474. doi: 10.1073/pnas.80.21.6470. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Han M., Grunstein M. Nucleosome loss activates yeast downstream promoters in vivo. Cell. 1988 Dec 23;55(6):1137–1145. doi: 10.1016/0092-8674(88)90258-9. [DOI] [PubMed] [Google Scholar]
- Hare J. T., Taylor J. H. One role for DNA methylation in vertebrate cells is strand discrimination in mismatch repair. Proc Natl Acad Sci U S A. 1985 Nov;82(21):7350–7354. doi: 10.1073/pnas.82.21.7350. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Harrington M. A., Jones P. A., Imagawa M., Karin M. Cytosine methylation does not affect binding of transcription factor Sp1. Proc Natl Acad Sci U S A. 1988 Apr;85(7):2066–2070. doi: 10.1073/pnas.85.7.2066. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Jones N. C., Rigby P. W., Ziff E. B. Trans-acting protein factors and the regulation of eukaryotic transcription: lessons from studies on DNA tumor viruses. Genes Dev. 1988 Mar;2(3):267–281. doi: 10.1101/gad.2.3.267. [DOI] [PubMed] [Google Scholar]
- Jones P. A., Taylor S. M. Cellular differentiation, cytidine analogs and DNA methylation. Cell. 1980 May;20(1):85–93. doi: 10.1016/0092-8674(80)90237-8. [DOI] [PubMed] [Google Scholar]
- Keshet I., Lieman-Hurwitz J., Cedar H. DNA methylation affects the formation of active chromatin. Cell. 1986 Feb 28;44(4):535–543. doi: 10.1016/0092-8674(86)90263-1. [DOI] [PubMed] [Google Scholar]
- Keshet I., Yisraeli J., Cedar H. Effect of regional DNA methylation on gene expression. Proc Natl Acad Sci U S A. 1985 May;82(9):2560–2564. doi: 10.1073/pnas.82.9.2560. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kovesdi I., Reichel R., Nevins J. R. Role of an adenovirus E2 promoter binding factor in E1A-mediated coordinate gene control. Proc Natl Acad Sci U S A. 1987 Apr;84(8):2180–2184. doi: 10.1073/pnas.84.8.2180. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lorch Y., LaPointe J. W., Kornberg R. D. Nucleosomes inhibit the initiation of transcription but allow chain elongation with the displacement of histones. Cell. 1987 Apr 24;49(2):203–210. doi: 10.1016/0092-8674(87)90561-7. [DOI] [PubMed] [Google Scholar]
- Lorch Y., LaPointe J. W., Kornberg R. D. On the displacement of histones from DNA by transcription. Cell. 1988 Dec 2;55(5):743–744. doi: 10.1016/0092-8674(88)90128-6. [DOI] [PubMed] [Google Scholar]
- Morse R. H. Nucleosomes inhibit both transcriptional initiation and elongation by RNA polymerase III in vitro. EMBO J. 1989 Aug;8(8):2343–2351. doi: 10.1002/j.1460-2075.1989.tb08362.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Morse R. H., Simpson R. T. DNA in the nucleosome. Cell. 1988 Jul 29;54(3):285–287. doi: 10.1016/0092-8674(88)90190-0. [DOI] [PubMed] [Google Scholar]
- Ogden G. B., Pratt M. J., Schaechter M. The replicative origin of the E. coli chromosome binds to cell membranes only when hemimethylated. Cell. 1988 Jul 1;54(1):127–135. doi: 10.1016/0092-8674(88)90186-9. [DOI] [PubMed] [Google Scholar]
- Razin A., Szyf M., Kafri T., Roll M., Giloh H., Scarpa S., Carotti D., Cantoni G. L. Replacement of 5-methylcytosine by cytosine: a possible mechanism for transient DNA demethylation during differentiation. Proc Natl Acad Sci U S A. 1986 May;83(9):2827–2831. doi: 10.1073/pnas.83.9.2827. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rougvie A. E., Lis J. T. The RNA polymerase II molecule at the 5' end of the uninduced hsp70 gene of D. melanogaster is transcriptionally engaged. Cell. 1988 Sep 9;54(6):795–804. doi: 10.1016/s0092-8674(88)91087-2. [DOI] [PubMed] [Google Scholar]
- Russell D. W., Zinder N. D. Hemimethylation prevents DNA replication in E. coli. Cell. 1987 Sep 25;50(7):1071–1079. doi: 10.1016/0092-8674(87)90173-5. [DOI] [PubMed] [Google Scholar]
- Saluz H. P., Jiricny J., Jost J. P. Genomic sequencing reveals a positive correlation between the kinetics of strand-specific DNA demethylation of the overlapping estradiol/glucocorticoid-receptor binding sites and the rate of avian vitellogenin mRNA synthesis. Proc Natl Acad Sci U S A. 1986 Oct;83(19):7167–7171. doi: 10.1073/pnas.83.19.7167. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Simpson R. T., Thoma F., Brubaker J. M. Chromatin reconstituted from tandemly repeated cloned DNA fragments and core histones: a model system for study of higher order structure. Cell. 1985 Oct;42(3):799–808. doi: 10.1016/0092-8674(85)90276-4. [DOI] [PubMed] [Google Scholar]
- Tanaka K., Appella E., Jay G. Developmental activation of the H-2K gene is correlated with an increase in DNA methylation. Cell. 1983 Dec;35(2 Pt 1):457–465. doi: 10.1016/0092-8674(83)90179-4. [DOI] [PubMed] [Google Scholar]
- Thompson J. P., Granoff A., Willis D. B. Infection with frog virus 3 allows transcription of DNA methylated at cytosine but not adenine residues. Virology. 1987 Sep;160(1):275–277. doi: 10.1016/0042-6822(87)90073-0. [DOI] [PubMed] [Google Scholar]
- 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]
- Vardimon L., Kressmann A., Cedar H., Maechler M., Doerfler W. Expression of a cloned adenovirus gene is inhibited by in vitro methylation. Proc Natl Acad Sci U S A. 1982 Feb;79(4):1073–1077. doi: 10.1073/pnas.79.4.1073. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wagner M. J., Sharp J. A., Summers W. C. Nucleotide sequence of the thymidine kinase gene of herpes simplex virus type 1. Proc Natl Acad Sci U S A. 1981 Mar;78(3):1441–1445. doi: 10.1073/pnas.78.3.1441. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Weintraub H. Assembly and propagation of repressed and depressed chromosomal states. Cell. 1985 Oct;42(3):705–711. doi: 10.1016/0092-8674(85)90267-3. [DOI] [PubMed] [Google Scholar]
- Wigler M., Levy D., Perucho M. The somatic replication of DNA methylation. Cell. 1981 Apr;24(1):33–40. doi: 10.1016/0092-8674(81)90498-0. [DOI] [PubMed] [Google Scholar]
- Workman J. L., Roeder R. G. Binding of transcription factor TFIID to the major late promoter during in vitro nucleosome assembly potentiates subsequent initiation by RNA polymerase II. Cell. 1987 Nov 20;51(4):613–622. doi: 10.1016/0092-8674(87)90130-9. [DOI] [PubMed] [Google Scholar]
- Yisraeli J., Frank D., Razin A., Cedar H. Effect of in vitro DNA methylation on beta-globin gene expression. Proc Natl Acad Sci U S A. 1988 Jul;85(13):4638–4642. doi: 10.1073/pnas.85.13.4638. [DOI] [PMC free article] [PubMed] [Google Scholar]