Article | Published:

Characterization of the Drosophila melanogaster genome at the nuclear lamina

Abstract

The nuclear lamina binds chromatin in vitro and is thought to function in its organization, but genes that interact with it are unknown. Using an in vivo approach, we identified 500 Drosophila melanogaster genes that interact with B-type lamin (Lam). These genes are transcriptionally silent and late replicating, lack active histone marks and are widely spaced. These factors collectively predict lamin binding behavior, indicating that the nuclear lamina integrates variant and invariant chromatin features. Consistently, proximity of genomic regions to the nuclear lamina is partly conserved between cell types, and induction of gene expression or active histone marks reduces Lam binding. Lam target genes cluster in the genome, and these clusters are coordinately expressed during development. This genome-wide analysis gives clear insight into the nature and dynamic behavior of the genome at the nuclear lamina, and implies that intergenic DNA functions in the global organization of chromatin in the nucleus.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Accession codes

Accessions

Gene Expression Omnibus

References

  1. 1

    Misteli, T. Spatial positioning; a new dimension in genome function. Cell 119, 153–156 (2004).

  2. 2

    Taddei, A., Hediger, F., Neumann, F.R. & Gasser, S.M. The function of nuclear architecture: a genetic approach. Annu. Rev. Genet. 38, 305–345 (2004).

  3. 3

    Marshall, W.F. Order and disorder in the nucleus. Curr. Biol. 12, R185–R192 (2002).

  4. 4

    Gartenberg, M.R., Neumann, F.R., Laroche, T., Blaszczyk, M. & Gasser, S.M. Sir-mediated repression can occur independently of chromosomal and subnuclear contexts. Cell 119, 955–967 (2004).

  5. 5

    Andrulis, E.D., Neiman, A.M., Zappulla, D.C. & Sternglanz, R. Perinuclear localization of chromatin facilitates transcriptional silencing. Nature 394, 592–595 (1998).

  6. 6

    Kosak, S.T. et al. Subnuclear compartmentalization of immunoglobulin loci during lymphocyte development. Science 296, 158–162 (2002).

  7. 7

    Zink, D. et al. Transcription-dependent spatial arrangements of CFTR and adjacent genes in human cell nuclei. J. Cell Biol. 166, 815–825 (2004).

  8. 8

    Mathog, D. & Sedat, J.W. The three-dimensional organization of polytene nuclei in male Drosophila melanogaster with compound XY or ring X chromosomes. Genetics 121, 293–311 (1989).

  9. 9

    Hochstrasser, M. & Sedat, J.W. Three-dimensional organization of Drosophila melanogaster interphase nuclei. II. Chromosome spatial organization and gene regulation. J. Cell Biol. 104, 1471–1483 (1987).

  10. 10

    Hochstrasser, M., Mathog, D., Gruenbaum, Y., Saumweber, H. & Sedat, J.W. Spatial organization of chromosomes in the salivary gland nuclei of Drosophila melanogaster. J. Cell Biol. 102, 112–123 (1986).

  11. 11

    Gerasimova, T.I., Byrd, K. & Corces, V.G. A chromatin insulator determines the nuclear localization of DNA. Mol. Cell 6, 1025–1035 (2000).

  12. 12

    Marshall, W.F., Dernburg, A.F., Harmon, B., Agard, D.A. & Sedat, J.W. Specific interactions of chromatin with the nuclear envelope: positional determination within the nucleus in Drosophila melanogaster. Mol. Biol. Cell 7, 825–842 (1996).

  13. 13

    Taddei, A. & Gasser, S.M. Multiple pathways for telomere tethering: functional implications of subnuclear position for heterochromatin formation. Biochim. Biophys. Acta 1677, 120–128 (2004).

  14. 14

    Casolari, J.M. et al. Genome-wide localization of the nuclear transport machinery couples transcriptional status and nuclear organization. Cell 117, 427–439 (2004).

  15. 15

    Brickner, J.H. & Walter, P. Gene recruitment of the activated INO1 locus to the nuclear membrane. PLoS Biol. 2, e342 (2004).

  16. 16

    Ishii, K., Arib, G., Lin, C., Van Houwe, G. & Laemmli, U.K. Chromatin boundaries in budding yeast: the nuclear pore connection. Cell 109, 551–562 (2002).

  17. 17

    Gruenbaum, Y., Margalit, A., Goldman, R.D., Shumaker, D.K. & Wilson, K.L. The nuclear lamina comes of age. Nat. Rev. Mol. Cell Biol. 6, 21–31 (2005).

  18. 18

    Taniura, H., Glass, C. & Gerace, L. A chromatin binding site in the tail domain of nuclear lamins that interacts with core histones. J. Cell Biol. 131, 33–44 (1995).

  19. 19

    Luderus, M.E. et al. Binding of matrix attachment regions to lamin B1. Cell 70, 949–959 (1992).

  20. 20

    Paddy, M.R., Belmont, A.S., Saumweber, H., Agard, D.A. & Sedat, J.W. Interphase nuclear envelope lamins form a discontinuous network that interacts with only a fraction of the chromatin in the nuclear periphery. Cell 62, 89–106 (1990).

  21. 21

    Belmont, A.S., Zhai, Y. & Thilenius, A. Lamin B distribution and association with peripheral chromatin revealed by optical sectioning and electron microscopy tomography. J. Cell Biol. 123, 1671–1685 (1993).

  22. 22

    Marshall, W.F., Fung, J.C. & Sedat, J.W. Deconstructing the nucleus: global architecture from local interactions. Curr. Opin. Genet. Dev. 7, 259–263 (1997).

  23. 23

    Kosak, S.T. & Groudine, M. Gene order and dynamic domains. Science 306, 644–647 (2004).

  24. 24

    van Steensel, B., Delrow, J. & Henikoff, S. Chromatin profiling using targeted DNA adenine methyltransferase. Nat. Genet. 27, 304–308 (2001).

  25. 25

    van Steensel, B. & Henikoff, S. Identification of in vivo DNA targets of chromatin proteins using tethered dam methyltransferase. Nat. Biotechnol. 18, 424–428 (2000).

  26. 26

    Bianchi-Frias, D. et al. Hairy transcriptional repression targets and cofactor recruitment in Drosophila. PLoS Biol. 2, E178 (2004).

  27. 27

    Greil, F. et al. Distinct HP1 and Su(var)3–9 complexes bind to sets of developmentally coexpressed genes depending on chromosomal location. Genes Dev. 17, 2825–2838 (2003).

  28. 28

    van Steensel, B., Delrow, J. & Bussemaker, H.J. Genomewide analysis of Drosophila GAGA factor target genes reveals context-dependent DNA binding. Proc. Natl. Acad. Sci. USA 100, 2580–2585 (2003).

  29. 29

    Sun, L.V. et al. Protein-DNA interaction mapping using genomic tiling path microarrays in Drosophila. Proc. Natl. Acad. Sci. USA 100, 9428–9433 (2003).

  30. 30

    Kitten, G.T. & Nigg, E.A. The CaaX motif is required for isoprenylation, carboxyl methylation, and nuclear membrane association of lamin B2. J. Cell Biol. 113, 13–23 (1991).

  31. 31

    Schübeler, D. et al. The histone modification pattern of active genes revealed through genome-wide chromatin analysis of a higher eukaryote. Genes Dev. 18, 1263–1271 (2004).

  32. 32

    Elgin, S.C. Heterochromatin and gene regulation in Drosophila. Curr. Opin. Genet. Dev. 6, 193–202 (1996).

  33. 33

    O'Keefe, R.T., Henderson, S.C. & Spector, D.L. Dynamic organization of DNA replication in mammalian cell nuclei: spatially and temporally defined replication of chromosome-specific alpha-satellite DNA sequences. J. Cell Biol. 116, 1095–1110 (1992).

  34. 34

    Schübeler, D. et al. Genome-wide DNA replication profile for Drosophila melanogaster: a link between transcription and replication timing. Nat. Genet. 32, 438–442 (2002).

  35. 35

    MacAlpine, D.M., Rodriguez, H.K. & Bell, S.P. Coordination of replication and transcription along a Drosophila chromosome. Genes Dev. 18, 3094–3105 (2004).

  36. 36

    Stolc, V. et al. A gene expression map for the euchromatic genome of Drosophila melanogaster. Science 306, 655–660 (2004).

  37. 37

    Boutanaev, A.M., Kalmykova, A.I., Shevelyov, Y.Y. & Nurminsky, D.I. Large clusters of co-expressed genes in the Drosophila genome. Nature 420, 666–669 (2002).

  38. 38

    Spellman, P.T. & Rubin, G.M. Evidence for large domains of similarly expressed genes in the Drosophila genome. J. Biol. 1, 5 (2002).

  39. 39

    Echalier, G. in Drosophila Cells in Culture 393–438 (Academic, New York, 1997).

  40. 40

    Marshall, W.F. et al. Interphase chromosomes undergo constrained diffusional motion in living cells. Curr. Biol. 7, 930–939 (1997).

  41. 41

    Heun, P., Laroche, T., Raghuraman, M.K. & Gasser, S.M. The positioning and dynamics of origins of replication in the budding yeast nucleus. J. Cell Biol. 152, 385–400 (2001).

  42. 42

    Vazquez, J., Belmont, A.S. & Sedat, J.W. Multiple regimes of constrained chromosome motion are regulated in the interphase Drosophila nucleus. Curr. Biol. 11, 1227–1239 (2001).

  43. 43

    Walter, J., Schermelleh, L., Cremer, M., Tashiro, S. & Cremer, T. Chromosome order in HeLa cells changes during mitosis and early G1, but is stably maintained during subsequent interphase stages. J. Cell Biol. 160, 685–697 (2003).

  44. 44

    Makatsori, D. et al. The inner nuclear membrane protein lamin B receptor forms distinct microdomains and links epigenetically marked chromatin to the nuclear envelope. J. Biol. Chem. 279, 25567–25573 (2004).

  45. 45

    Somech, R. et al. The nuclear-envelope protein and transcriptional repressor LAP2beta interacts with HDAC3 at the nuclear periphery, and induces histone H4 deacetylation. J. Cell Sci. 118, 4017–4025 (2005).

  46. 46

    Croft, J.A. et al. Differences in the localization and morphology of chromosomes in the human nucleus. J. Cell Biol. 145, 1119–1131 (1999).

  47. 47

    Cremer, M. et al. Inheritance of gene density-related higher order chromatin arrangements in normal and tumor cell nuclei. J. Cell Biol. 162, 809–820 (2003).

  48. 48

    Taddei, A., Roche, D., Bickmore, W.A. & Almouzni, G. The effects of histone deacetylase inhibitors on heterochromatin: implications for anticancer therapy? EMBO Rep 6, 520–524 (2005).

  49. 49

    Stuurman, N., Maus, N. & Fisher, P.A. Interphase phosphorylation of the Drosophila nuclear lamin: site-mapping using a monoclonal antibody. J. Cell Sci. 108, 3137–3144 (1995).

  50. 50

    Smyth, G.K. Linear models and empirical Bayes methods for assessing differential expression in microarray experiments. Stat. Appl. Genet. Mol. Biol. 3, Article3 (2004).

Download references

Acknowledgements

We thank J. Delrow and M. Aronszajn (Genomics facility, Fred Hutchinson Cancer Research Center) for providing and annotating of cDNA arrays; N. Stuurman for plasmids, antibodies and helpful suggestions; F. Greil and C. Moorman for valuable technical advice; G. Hart for advice on statistical methods; M. Heimerikx and the NKI microarray facility for technical support; L. Guelen, H. van der Velde, J. Hendriksen and D. Engelsma for scoring of the FISH images; and J.M. Boer, S. Nijman, F. van Leeuwen, J. Neefjes, M. van Lohuizen and members of the van Steensel and Fornerod labs for helpful discussions and critical reading of the manuscript. This work was supported by a Marie Curie European Community Training and Mobility grant to H.P. and an European Young Investigator Award to B.v.S.

Author information

H.P., M.F. and B.v.S. conceived and designed the project; H.P. performed DamID, expression profiling and microscopy experiments and designed FISH probes; B.K. developed and performed FISH experiments; W.T. performed ChIP; E.d.W., M.F. and B.v.S. designed and performed computational analyses and H.P., M.F. and B.v.S. wrote the manuscript with help from B.K. and E.d.W.

Competing interests

The authors declare no competing financial interests.

Correspondence to Maarten Fornerod or Bas van Steensel.

Supplementary information

Supplementary Fig. 1

Estimate of DNA content in the nuclear periphery. (PDF 273 kb)

Supplementary Fig. 2

Genomic Lamin binding in embryonic cells shows significant correspondence to polytene chromosome nuclear envelope contacts in larvae. (PDF 116 kb)

Supplementary Fig. 3

Genes bound by Lam are repressed, lack active histone marks and are late replicating. (PDF 157 kb)

Supplementary Fig. 4

The binding pattern of Lam is distinct from that of HP1 and Su(var)3-9. (PDF 1100 kb)

Supplementary Fig. 5

Developmental expression profiles of Lam target gene clusters. (PDF 99 kb)

Supplementary Table 1

Excel spreadsheet containing DamID data, clustering data, expression profiling, flanking intergenic region lengths, probe annotation and gene ontology analysis of Lam-binding genes. (XLS 6469 kb)

Supplementary Table 2

Characteristics of FISH probes. (PDF 86 kb)

Supplementary Table 3

Multiple regression analysis of Lam binding. (PDF 64 kb)

Rights and permissions

Reprints and Permissions

About this article

Further reading

Figure 1: Generation of genome-wide binding maps by expression of Dam-fused Lam proteins.
Figure 2: Lam targets identified by DamID are enriched at the nuclear envelope (NE).
Figure 3: Lam-associated genes are repressed.
Figure 4: Lam binding genes are identified by a combination of DNA and chromatin properties.
Figure 5: Global loss of Lam binding upon inhibition of histone deacetylation.
Figure 6: Lamina-associated gene clusters are units of developmental regulation.
Figure 7: Changes in nuclear lamina binding of gene clusters during differentiation.
Figure 8: Model summarizing factors involved in the association of Lam binding genes to the nuclear lamina.