We discovered and cloned mouse and human BORIS, a CTCF-paralogous gene that emerged in a vertebrate ancestor from a duplication of CTCF locus on a different chromosome. In mammals, BORIS evolved into germ cell specific factor with the same DNA binding specificity as CTCF due to the preservation of all 11 ZFs and their coding exons. However, BORIS underwent substantial divergence in its amino and carboxy termini and corresponding exons, as well as in flanking non-coding regulatory regions. As a result, BORIS attained the expression pattern and biological properties that are both distinct from CTCF and specific for germ cells. At the same time BORIS is strictly repressed in somatic cells, where it would interfere with essential CTCF functions. We named BORIS the Brother Of the Regulator of Imprinted Sites because CTCF plays a critical role in imprinting-mediated gene regulation in somatic cells of placental mammals. However, the involvement and mechanistic role of BORIS and/or of BORIS+ CTCF heterodimers in re-establishing imprinting marks in germ cells remains to be elucidated. The 11ZF region was previously identified in the lab as a multivalent DBD, which is able to recognize and bind extended (around 50bp) target sequences. By virtue of sharing the identical DBD, CTCF and BORIS can recognize the same DNA sequences. Outside of the identical 11 ZF and AT-hook domains, flanking carboxy- and N- terminal regions of CTCF orthologs have >90% identical a.a sequences in all vertebrates from frogs to humans. In contrast, a pair wise comparison of these regions in mouse and human BORIS proteins and in corresponding CTCF proteins revealed no obvious homologies of one to another and no significant similarities to other proteins. Thus, while mammalian CTCF and BORIS proteins will recognize the same or an overlapping spectrum of DNA sequences, the dissimilar flanking regions indicate that the functional consequences of DNA binding by these two proteins is likely to be different. Indeed, the two factors are largely antagonistic, as expression of BORIS in normally BORIS-negative cells promotes cell growth, which can lead to transformation, while CTCF overexpression blocks cell proliferation. Human BORIS gene maps to the 20q13 region well known for frequent gains and/or amplification observed in many of the same types of tumors that also show often LOH at CTCF locus on 16q22, including breast cancers. Unlike CTCF gene, BORIS is expressed in multiple isoforms generated by combinatorial usage of alternative promoters coupled with alternative splicing. Remarkably, several BORIS isoforms contain DBDs with variable number of ZFs and with novel ZF-types not found in CTCF. This explains how BORIS isoforms can recognize DNA sequences distinct from CTCF target sites (CTSes), other isoforms may potentially and may. Furthermore, it is now apparent that mouse is not completely adequate model to study human BORIS, because mBoris isoforms are significantly different from those found in humans and in primates. Therefore, a primate animal model should subserve better for GNW profiling CTCF vs. BORIS sites and for recapitulating human functions of the gene. (1) BORIS in germline development. Limiting BORIS expression to testicular germ cells is important for normal spermeiogenesis when proper epigenetic regulation by certain CTCF sites requires their in vivo binding to BORIS rather than to CTCF. Indeed, in addition to one recently published example of consensus CTCF site that may never bind to CTCF, it is likely that switching to BORIS-occupancy allows many other CTCF-sites to turn into mediators of germline specific functions of BORIS that are distinct from known functions of CTCF. We identified multiple spermatogenesis-specific pathways that depend on BORIS as a transcriptional factor, including cancer-testis antigens. We are testing if BORIS binding to a subset of CTSes, - that are normally not supposed to be re-methylated upon BER-like methylation erasure in haploid epigenomes, - may serve as a surveillance sensor factor responsible for epigenetic integrity of CTCF sites networks, preventing thereby epimutations in mature spermatozoa from transgenerational transfer. (2) BORIS and cancer, In addition to its role in germ cell development, BORIS likely plays a key role in oncogenesis. Indeed, while BORIS expression is silenced in normal somatic cells, it is activated in cancer cells;i.e. BORIS is a so-called cancer-testis (CT) gene. We and others previously characterized BORIS expression in uterine cancers, breast cancers, osteosarcomas, lung cancers, and prostate cancers. However, as BORIS is itself a gene expression regulator, it was hypothesized that BORIS-mediated regulation of promoters is the regulatory network responsible for the expression of multiple CT genes. Using the Boris KO model we demonstrated that BORIS directly regulates the testis-specific protease gene TSP50, which is in turn negatively regulated by p53. In another line of experiments, we previously discovered that DNA methylation plays dual role in the regulation of human telomerase gene, hTERT, one of the key cell immortalization factors. (3) BORIS and viruses. We demonstrated that acute infections by viruses, such as Epstein-Barr virus (EBV), activate BORIS expression. Furthermore, it appears that the optimal level of BORIS expression is required for cell immortalization by EBV. Overexpression of BORIS is toxic for cells, and the excess if BORIS is inactivated, likely by the process of autophagy. CTCF is involved in establishment and maintenance of latent infection of gammaherpesviruses, EBV, and Kaposi's sarcoma-associated herpesvirus (KSHV). By virtue of direct binding to virus-borne CTSes, CTCF enables the integration of latent virus into cellular machinery. It is thus conceivable that BORIS might be also involved in viral latency by interfering with CTCF functions in this pathway. The mechanism of this interaction remains to be elucidated. Recently we demonstrated that acute HIV infection of T-cells induces expression of a 64-KD protein that is recognized by anti-BORIS monoclonal antibody. MPS was awarded a supplementary grant from the Office of AIDS Research to characterize this protein, likely an isoform of BORIS.
Showing the most recent 10 out of 21 publications
