Release of epigenetic gene silencing by trans-acting mutations in Arabidopsis - PubMed (original) (raw)
Release of epigenetic gene silencing by trans-acting mutations in Arabidopsis
O Mittelsten Scheid et al. Proc Natl Acad Sci U S A. 1998.
Abstract
Gene silencing in plants inactivates trans-genes introduced into plants and/or endogenous homologous genes. This stable but potentially reversible loss of gene activity resembles epigenetic changes that occur in normal development. The stability of silencing implies the involvement of trans-acting components, although none of them have been identified so far. Here we report the finding of second-site mutations interfering with maintenance of the silent state. We mutagenized Arabidopsis thaliana plants carrying a silent transgene encoding hygromycin phosphotransferase (hpt) and therefore show a heritable hygromycin-sensitive phenotype. The M2 generation was screened for hygromycin resistance. Eight putative mutants (som1 to 8) were found that expressed the transgene and transmitted the expressed state to their progeny. All mutations were shown to reactivate a silent transgenic test locus in trans. The level of DNA methylation at the hpt locus and at centromeric repeats was found to be reduced in the som mutants. Complementation crosses indicated complex epigenetic interactions among the som mutant alleles and with the previously described ddm1 allele, which elicits DNA hypomethylation [Vongs, A., Kakutani, T, Martiensen, R.A. & Richards, E.J. (1993) Science 260, 1926-1928]. Som mutants can be classified into three groups: (i) allelic or interacting with ddm1 and with each other (som 1,4, and 5), (ii) nonallelic with ddm1 and som mutants of group A (som2), and (iii) mutants with slow resilencing after out-crosses, which hinders their classification (som 3, 6, 7 and 8).
Figures
Figure 1
Selection of mutants. Primary selection of mutant candidates: hygromycin-resistant seedling after 8 days selection on 15 mg/l hygromycin (a), Northern (b), and Southern (c) blot analysis of the parental line A with the silent hpt gene and the mutants _som1_-som8. Both membranes were probed with the hpt coding region. Genomic DNA was digested with _Bam_HI.
Figure 2
Reactivation of a silent gene in test crosses with the mutants. (a) Scheme of the crosses for mutants with hemizygous transgene. (b) Scheme of the crosses for mutants with homozygous transgene. (c) F2 populations after 2 weeks on 10 mg/l hygromycin. som, mutant locus; SOM, corresponding wild-type allele; _A_s, hpt gene in silent state; _A_r, hpt gene in reactivated state; −, transgene-free locus allelic to A.
Figure 3
Methylation analysis. The same Southern blot of DNA cut with _Hpa_II was probed with the hpt coding region (a), or with a fragment containing the centromeric 180-bp repeat (b; ref. 20). Lanes: 1, pGl2 plasmid used to generate the parental line A; 2, wt; 3, parental line A; 4–11, som 1–8, 12, mutant ddm1/ddm1 with reduced methylation (20); 13, _ddm1/_A; 14, METIas T3 #10.5 transgenic for a methyltransferase antisense construct (21); 15, METIas T3 #10.5/A.
Figure 4
Complementation analysis. F1 hybrids of som2, som5, and som7 with ddm1 or wt, after 2 weeks on 10 mg/l hygromycin.
Similar articles
- Two regulatory levels of transcriptional gene silencing in Arabidopsis.
Mittelsten Scheid O, Probst AV, Afsar K, Paszkowski J. Mittelsten Scheid O, et al. Proc Natl Acad Sci U S A. 2002 Oct 15;99(21):13659-62. doi: 10.1073/pnas.202380499. Epub 2002 Oct 7. Proc Natl Acad Sci U S A. 2002. PMID: 12370435 Free PMC article. - Formation of stable epialleles and their paramutation-like interaction in tetraploid Arabidopsis thaliana.
Mittelsten Scheid O, Afsar K, Paszkowski J. Mittelsten Scheid O, et al. Nat Genet. 2003 Aug;34(4):450-4. doi: 10.1038/ng1210. Nat Genet. 2003. PMID: 12847525 - Susceptibility of transgene loci to homology-dependent gene silencing.
Neuhuber F, Park YD, Matzke AJ, Matzke MA. Neuhuber F, et al. Mol Gen Genet. 1994 Aug 2;244(3):230-41. doi: 10.1007/BF00285450. Mol Gen Genet. 1994. PMID: 8058034 - Heritable epigenetic mutation of a transposon-flanked Arabidopsis gene due to lack of the chromatin-remodeling factor DDM1.
Saze H, Kakutani T. Saze H, et al. EMBO J. 2007 Aug 8;26(15):3641-52. doi: 10.1038/sj.emboj.7601788. Epub 2007 Jul 12. EMBO J. 2007. PMID: 17627280 Free PMC article. - Transcriptional gene silencing mutants.
Mittelsten Scheid O, Paszkowski J. Mittelsten Scheid O, et al. Plant Mol Biol. 2000 Jun;43(2-3):235-41. doi: 10.1023/a:1006487529698. Plant Mol Biol. 2000. PMID: 10999407 Review.
Cited by
- Molecular and structural basis of the chromatin remodeling activity by Arabidopsis DDM1.
Osakabe A, Takizawa Y, Horikoshi N, Hatazawa S, Negishi L, Sato S, Berger F, Kakutani T, Kurumizaka H. Osakabe A, et al. Nat Commun. 2024 Jul 11;15(1):5187. doi: 10.1038/s41467-024-49465-w. Nat Commun. 2024. PMID: 38992002 Free PMC article. - Tissue-specific silencing of integrated transgenes achieved through endogenous RNA interference in Caenorhabditis elegans.
Chen S, Liu W, Xiong L, Tao Z, Zhao D. Chen S, et al. RNA Biol. 2024 Jan;21(1):1-10. doi: 10.1080/15476286.2024.2332856. Epub 2024 Mar 26. RNA Biol. 2024. PMID: 38531838 Free PMC article. - DDM1-Mediated TE Silencing in Plants.
Akinmusola RY, Wilkins CA, Doughty J. Akinmusola RY, et al. Plants (Basel). 2023 Jan 18;12(3):437. doi: 10.3390/plants12030437. Plants (Basel). 2023. PMID: 36771522 Free PMC article. Review. - Seminars in cell and development biology on histone variants remodelers of H2A variants associated with heterochromatin.
Berger F, Muegge K, Richards EJ. Berger F, et al. Semin Cell Dev Biol. 2023 Feb 15;135:93-101. doi: 10.1016/j.semcdb.2022.02.026. Epub 2022 Mar 4. Semin Cell Dev Biol. 2023. PMID: 35249811 Free PMC article. Review. - Zebularine induces enzymatic DNA-protein crosslinks in 45S rDNA heterochromatin of Arabidopsis nuclei.
Prochazkova K, Finke A, Tomaštíková ED, Filo J, Bente H, Dvořák P, Ovečka M, Šamaj J, Pecinka A. Prochazkova K, et al. Nucleic Acids Res. 2022 Jan 11;50(1):244-258. doi: 10.1093/nar/gkab1218. Nucleic Acids Res. 2022. PMID: 34904670 Free PMC article.
References
- Meyer, P., ed. (1995) Curr. Top. Microbiol. Immunol. 197. - PubMed
- Matzke M, Matzke A J M. Trends Genet. 1995;11:1–3. - PubMed
- Meyer P, Saedler H. Annu Rev Plant Physiol Plant Mol Biol. 1996;47:23–48. - PubMed
Publication types
MeSH terms
Substances
LinkOut - more resources
Full Text Sources
Other Literature Sources
Molecular Biology Databases