The Role of MAR-Binding Protein in Breast Cancer

Institution: Lawrence Berkeley National Laboratory
Investigator(s): Terumi Kohwi-Shigematsu, Ph.D. - Terumi Kohwi-Shigematsu, Ph.D. -
Award Cycle: 1995 (Cycle I) Grant #: 1RB-0381A Award: $328,849
Award Type: Research Project Awards
Research Priorities
Biology of the Breast Cell>Pathogenesis: understanding the disease

This is a collaboration with: 1RB-0381 -

Initial Award Abstract (1995)
The nucleus of a cell contains the genetic information of each individual, such as DNA. This DNA material is organized upon a framework within the nucleus called the nuclear matrix, which is made of protein and resembles a mesh in design. Some segments of the DNA, called matrix-attachment regions (MARs), anchor themselves onto the nuclear matrix. Our laboratory has isolated a protein called p114 that binds to MARs. The MAR-binding activity of this protein is found only in malignant human breast tumor specimens, not in normal breast tissues or benign breast disease tissues. This fact indicates that p114 might be an excellent diagnostic tool for the detection of cancerous cells. In addition, strong p114 MAR-binding activity has been detected in aggressive tumors that are more fully developed, while significantly weaker p114 activity has been observed in less aggressive tumors.

In this project, we will clone the gene for p114 and develop antibodies specific for p114 to evaluate the use of p114 as a diagnostic/prognostic marker for breast cancer. The information generated from this study may help us to understand how normal cells become cancerous and to develop better tools for the early detection and prevention of breast cancer.

Final Report (1999)
Note: This project was extended for an additional year.

In this project we focused on proteins that potentially modulate DNA organization (i.e., DNA packaging) in the nuclei, because such proteins can potentially regulate multiple genes. In breast cancer, there might be a change in the amount and/or function of DNA packaging proteins to sustain uncontrolled cell growth. Mammalian DNA is organized in the nuclei through many layers of complexity. At the higher level of packaging, DNA fibers are organized into loop domain structures that make attachments to the skeletal framework of the nuclei. These attachment sites of DNA (or bases of the DNA loops) are called matrix attachment regions (MARs).

We have identified a protein whose MAR-binding activity can be highly detected in breast carcinomas and virtually undetectable in breast lesions or in normal breast tissues. Unexpectedly, this protein was identified as poly(ADP-ribose) polymerase (PARP), which is a nuclear enzyme that participates in the cellular response to DNA damage. We also found that PARP associates with DNA-dependent protein kinase (DNA-PK), which consists of the catalytic subunit and the heterodimeric DNA-binding subunit, Ku70/86. Collectively, the association of PARP and DNA-PK binds with high affinity and specificity to the key structural element of MARs. This results in regions with high unwinding propensity (BUR: base unpairing region).

Our discoveries on PARP and DNA-PK were unexpected, and they are in contrast to the long held notion that PARP/DNA-PK bind mainly to nicks and ends of DNA. Furthermore, autoribosylation of PARP abolished its association with Ku 70/86 and BUR-binding activity, which separates PARP with and without BUR-binding activity. To establish the link of PARP to breast cancer, we succeeded in reducing the level of PARP in an aggressive breast cancer cell line, MDA-MB-231 cells, by treating them with an antisense construct. After this treatment the cells lost their invasive characteristics in vitro and showed a reduced ability to form tumors in mice. Therefore, a high level expression of PARP is important to maintain the aggressive phenotype of breast cancer cells.

These findings have provided a molecular link between PARP and DNA-PK complex to breast carcinogenesis and suggest, for the first time, a novel role for these enzymes in the modulation of chromatin structure and function.

Poly (ADP-ribose) polymerase and Ku autoantigen form a complex and synergistically bind to matrix attachment sequences
Periodical:Journal of Biological Chemistry
Index Medicus: J Biol Chem
Authors: Galande S and Kohwi-Shigematsu T
Yr: 1999 Vol: 274 Nbr: 29 Abs: Pg:20521-20528

A matrix attachment region (MAR)-binding activity due to a p114 kilodalton protein is found only in human breast carcinomas and not in normal and benign breast disease tissues
Periodical:Cancer Research
Index Medicus: Cancer Res
Authors: Yanagisawa J, Ando J, Nakayama J, Kohwi Y, and Kohwi-Shigematsu T
Yr: 1996 Vol: 56 Nbr: Abs: Pg:457-462