Poly(ADPribosyl)ation of CTCF: role in breast tumourigenesis

F Docquier, D Farrar, I Chernukhin and E Klenova

Department of Biological Sciences, University of Essex, Colchester, UK

from Breast cancer research: the past and the future
London, UK. 1 November 2006

Breast Cancer Research 2006, 8(Suppl 2):P7doi:10.1186/bcr1562

Published:

1 November 2006

Background

CTCF is a conserved, ubiquitous and multifunctional 11 Zn finger (ZF) transcription factor with features of a tumour suppressor. CTCF regulates transcription in diverse modes, such as promoter activation and repression, silencing, constitutive and methylation-dependent chromatin insulation. We have previously reported that CTCF can be post-translationally regulated by poly(ADPribosyl)ation and that this modification modulates the insulator function of CTCF [1,2]. The purpose of the present study is to investigate the role of CTCF poly(ADPribosyl)ation in normal and breast cancer cells.

Methods

The following techniques have been used in this investigation: western analysis, mass spectrometry, immunoprecipitation, cell cultures, transient transfection, primary cultures from normal and tumour tissues, cellular fractionation and laser capture microdissection.

Results

Using a large panel of breast tumours and paired peripheral tissues, we have discovered that only the poly(ADPribosyl)ated isoform of CTCF (called CTCF180) is detected in normal breast tissues, whereas the other isoform of CTCF (called CTCF130) only appears in breast tumour tissues and immortalised cell lines (see Figure 1). The identity of the poly(ADPribosyl)ated isoform of CTCF was further verified by mass spectrometry. We are currently establishing primary cultures from normal and tumour tissues in order to investigate whether the appearance of CTCF130 is linked to immortalisation. The histological type of cells containing CTCF180 and CTCF130 is being determined by cellular fractionation and laser capture microdissection of breast tissues. Finally, experiments using cell culture models suggest that the generation of the CTCF180 can be associated with cell cycle arrest.

Figure 1. Western blot analysis of six paired samples (1–6) of normal and tumour tissues (a representative western blot of total 56 paired samples is shown). Fifty micrograms of the total protein were used in this assay. The bands of the CTCF 180 kDa and 130 kDa isoforms were quantified and their additive values were normalized to the corresponding values of α-Tubulin used as loading control (bottom panel).

Conclusion

This research addresses the molecular mechanisms of breast tumourigenesis: CTCF180 and CTCF130 may regulate different sets of genes and/or different cell functions specific for normal and cancer cells, respectively. The loss of CTCF poly(ADPribosyl)ation could also lead to epigenetic disturbances. Our data obtained so far indicate that the transition from CTCF180 to CTCF130 could be a hallmark of tumour development. We envisage the potential use of both CTCF isoforms as biological markers for breast tumourigenesis.

Acknowledgements

This work was funded by Breast Cancer Campaign, The Medical Research Council, and The University of Essex.

References

Yu W, Ginjala V, Pant V, Chernukhin I, Whitehead J, Docquier F, Farrar D, Tavoosidana R, Mukhopadhyay R, Kanduri C, et al.: Poly(ADPribosyl)ation regulates CTCF dependent chromatin insulation.
Nat Genet 2004, 36:1105-1110. PubMed Abstract | Publisher Full Text
Klenova E, Ohlsson R: Poly(ADPribosyl)ation and epigenetics: is CTCF PARt of the plot?
Cell Cycle 2005, 4:96-101. PubMed Abstract | Publisher Full Text

This article is part of the supplement: Breast cancer research: the past and the future