Abstract
In tumor progression definite alterations in nuclear matrix (NM) protein composition as well as in chromatin structure occur. The NM interacts with chromatin via specialized DNA sequences called matrix attachment regions (MARs). In the present study, using a proteomic approach along with a two-dimensional Southwestern assay and confocal laser microscopy, we show that the differentiation of stabilized human prostate carcinoma cells is marked out by modifications both NM protein composition and bond between NM proteins and MARs. Well-differentiated androgen-responsive and slowly growing LNCaP cells are characterized by a less complex pattern and by a major number of proteins binding MAR sequences in comparison to 22Rv1 cells expressing androgen receptor but androgen-independent. Finally, in the poorly differentiated and strongly aggressive androgen-independent PC3 cells the complexity of NM pattern further increases and a minor number of proteins bind the MARs. Furthermore, in this cell line with respect to LNCaP cells, these changes are synchronous with modifications in both the nuclear distribution of the MAR sequences and in the average loop dimensions that significantly increase. Although the expression of many NM proteins changes during dedifferentiation, only a very limited group of MAR-binding proteins seem to play a key role in this process. Variations in the expression of poly (ADP-ribose) polymerase (PARP) and special AT-rich sequence-binding protein-1 (SATB1) along with an increase in the phosphorylation of lamin B represent changes that might trigger passage towards a more aggressive phenotype. These results suggest that elucidating the MAR-binding proteins that are involved in the differentiation of prostate cancer cells could be an important tool to improve our understanding of this carcinogenesis process, and they could also be novel targets for prostate cancer therapy.
Highlights
Abnormal nuclear organization and alterations in the amount and distribution of heterochromatin have long been recognized as hallmarks of human cancer [1]; at present we do not know the exact causes of these modifications, nor do we know how the activity/silencing of thousands of genes is orchestrated
Polypeptides, we further characterized the expression of the nuclear matrix (NM) proteins by 2D-PAGE followed by 2D Southwestern blotting (SWB) analysis
We provide evidence that shows, for the first time, that during Prostate carcinoma (PCa) cell differentiation the binding between the NM proteins and matrix attachment regions (MARs) sequences is dynamic and inversely correlated with the cellular differentiation
Summary
Abnormal nuclear organization and alterations in the amount and distribution of heterochromatin have long been recognized as hallmarks of human cancer [1]; at present we do not know the exact causes of these modifications, nor do we know how the activity/silencing of thousands of genes is orchestrated. The interactions between chromatin and the NM occur via AT-rich DNA sequences called matrix attachment regions (MARs). Not all potential MARs are bound to the NM or participate in the organization of loop attachment regions. Several MAR-binding proteins have been identified, some of which are dramatically deregulated in tumor cells. Often their expression is significantly correlated with aggressive tumor phenotypes. Modifications in the interactions between NM proteins and MARs might be related to the large-scale chromatin reorganization observed during carcinogenesis. This has prompted a rising interest in MARs and MAR-binding proteins as potential targets for antineoplastic drugs [2]
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