274

BIN1

AMPH2|AMPHL|SH3P9;bridging integrator 1;BIN1;bridging integrator 1

amphiphysin II|amphiphysin-like protein|box dependant MYC interacting protein 1|box-dependent myc-interacting protein 1|myc box-dependent-interacting protein 1

2q14

protein-coding

Loss of tumor suppressors that restrain important oncoproteins such as c-Myc may contribute to malignant progression. Bin1 is an adapter protein with features of a tumor suppressor that was identified through its interaction with and inhibition of the oncogenic properties of c-Myc. In this study, we analyzed the patterns of Bin1 expression in normal melanocytes and melanoma cells at different stages of tumor progression. Evidence is provided that Bin1 function is abrogated in melanoma cells by a mechanism based on aberrant splicing of a tissue-specific exon. Specifically, most melanoma cells inappropriately expressed exon 12A, which is spliced alternately into Bin1 isoforms found in brain but not into isoforms found in melanocytes and many other nonneuronal cells. Exon 12A sequences abolished the ability of Bin1 to inhibit malignant transformation by c-Myc or adenovirus E1A. Similarly, these sequences abolished the ability of Bin1 to induce programmed cell death in melanoma cells that endogenously expressed exon 12A. Our findings suggest that aberrant splicing of Bin1 may contribute to melanoma progression, and they define a mechanism by which the activity of a tumor suppressor can be eliminated in cells.

Oncogenic activation of MYC occurs often in breast carcinoma and is associated with poor prognosis. Loss or inactivation of mechanisms that restrain MYC may therefore be involved in tumor progression. In this study, we show that the MYC-interacting adaptor protein BIN1 is frequently missing in malignant breast cells and that this loss is functionally significant. BIN1 was expressed in normal and benign cells and tissues but was undetectable in 6/6 estrogen receptor-positive or estrogen receptor-negative carcinoma cell lines examined. Similarly, complete or partial losses of BIN1 were documented in 30/50 (60%) cases of malignant breast tissue analyzed by immuno-histochemistry or RT-PCR. Abnormalities in the organization of the BIN1 gene were apparent in only a minority of these cases, suggesting that most losses were due to epigenetic causes. Nevertheless, they were functionally significant because ectopic BIN1 induced programmed cell death in malignant cells lacking endogenous BIN1 but had no effect on the viability of benign cells. We propose that loss of BIN1 may contribute to breast cancer progression by eliminating a mechanism that restrains the ability of activated MYC to drive cell division inappropriately.

The genetic events underlying the development of prostate cancer are poorly defined. c-Myc is often activated in tumors that have progressed to metastatic status, so events that promote this process may be important. Bin1 is a nucleocytoplasmic adaptor protein with features of a tumor suppressor that was identified through its ability to interact with and inhibit malignant transformation by c-Myc. We investigated a role for Bin1 loss or inactivation in prostate cancer because the human Bin1 gene is located at chromosome 2q14 within a region that is frequently deleted in metastatic prostate cancer but where no tumor suppressor candidate has been located. A novel polymorphic microsatellite marker located within intron 5 of the human Bin1 gene was used to demonstrate loss of heterozygosity and coding alteration in 40% of informative cases of prostate neoplasia examined. RNA and immunohistochemical analyses indicated that Bin1 was expressed in most primary tumors, even at slightly elevated levels relative to benign tissues, but that it was frequently missing or inactivated by aberrant splicing in metastatic tumors and androgen-independent tumor cell lines. Ectopic expression of Bin1 suppressed the growth of prostate cancer lines in vitro. Our findings support the candidacy of Bin1 as the chromosome 2q prostate tumor suppressor gene.

The E2F1 transcription factor, which was originally identified as a cell-cycle initiator, mediates apoptosis in response to DNA damage. As E2F1-induced apoptosis is an attractive mechanism for cancer therapy, it is critical to fully elucidate its effector pathways. Here, we show that the c-MYC-interacting proapoptotic tumor suppressor, BIN1, is transcriptionally activated by E2F1 and mediates E2F1-induced apoptosis in response to DNA damage. Acting through the DNA-binding and transactivation domains, ectopically expressed E2F1 activated the human BIN1 promoter, which contains canonical E2F-recognition sites. Conversely, depletion of E2F1 by small interfering RNA or germline deletion led to BIN1 deficiency. DNA-damaging agents (which included etoposide) increased BIN1 levels, unless E2F1 was deficient. Moreover, endogenous E2F1 protein interacted directly with the BIN1 gene promoter in chromatin, particularly after etoposide treatment. Notably, suppression of BIN1 expression using an antisense (AS) technique attenuated the cell death mediated by E2F1 and etoposide. Although the p53 tumor suppressor, its sibling protein p73, and caspases are well-known E2F1 effectors for DNA damage-induced apoptosis, AS-BIN1 did not compromise their apoptotic functions. Our results collectively suggest that BIN1 is a novel transcriptional target of E2F1 that triggers a unique mode of cell death in response to DNA damage.

Indoleamine 2,3-dioxygenase (IDO) is generally considered to be immunosuppressive but recent findings suggest this characterization oversimplifies its role in disease pathogenesis. Recently, we showed that IDO is essential for tumor outgrowth in the classical two-stage model of inflammatory skin carcinogenesis. Here, we report that IDO loss did not exacerbate classical inflammatory responses. Rather, IDO induction could be elicited by environmental signals and tumor promoters as an integral component of the inflammatory tissue microenvironment even in the absence of cancer. IDO loss had limited impact on tumor outgrowth in carcinogenesis models that lacked an explicit inflammatory tumor promoter. In the context of inflammatory carcinogenesis where IDO was critical to tumor development, the most important source of IDO was radiation-resistant non-hematopoietic cells, consistent with evidence that loss of the IDO regulatory tumor suppressor gene Bin1 in transformed skin cells facilitates IDO-mediated immune escape by a cell autonomous mechanism. Taken together, our results identify IDO as an integral component of cancer-associated inflammation that tilts the immune system toward tumor support. More generally, they promote the concept that mediators of immune escape and cancer-associated inflammation may be genetically synonymous.

Src homology 3 (SH3) domains are widely known for their ability to interact with other proteins using the canonical PxxP binding motif. Besides those well-characterized interaction modes, there is an increasing number of SH3 domain-containing complexes that lack this motif. Here we characterize the interaction of SH3 domains, in particular the Bin1-SH3 domain, with the intrinsically disordered part of nonstructural protein 5A of the hepatitis C virus using noncanonical binding sites in addition to its PxxP motif. These binding regions partially overlap with regions that have previously been identified as having an increased propensity to form alpha-helices. Remarkably, upon interaction with the Bin1-SH3 domain, the alpha-helical propensity decreases and a fuzzy complex is formed.

BIN1 is a novel protein that interacts with the functionally critical Myc box regions at the N terminus of the MYC oncoprotein. BIN1 is structurally related to amphiphysin, a breast cancer-associated autoimmune antigen, and RVS167, a negative regulator of the yeast cell cycle, suggesting roles in malignancy and cell cycle control. Consistent with this likelihood, BIN1 inhibited malignant cell transformation by MYC. Although BIN1 is expressed in many normal cells, its levels were greatly reduced or undetectable in 14/27 carcinoma cell lines and 3/6 primary breast tumours. Deficits were functionally significant because ectopic expression of BIN1 inhibited the growth of tumour cells lacking endogenous message. We conclude that BIN1 is an MYC-interacting protein with features of a tumour suppressor.

Bin1 is a Myc-interacting protein with features of a tumor suppressor. The high level of Bin1 expression in skeletal muscle prompted us to investigate its role in muscle differentiation. Significant levels of Bin1 were observed in undifferentiated C2C12 myoblasts, a murine in vitro model system. Induction of differentiation by growth factor withdrawal led to an upregulation of Bin1 mRNA and to the generation of higher-molecular-weight forms of Bin1 protein by alternate splicing. While Bin1 in undifferentiated cells was localized exclusively in the nucleus, differentiation-associated isoforms of Bin1 were found in the cytoplasm as well. To examine the function of Bin1 during differentiation, we generated stable cell lines that express exogenous human Bin1 cDNA in the sense or antisense orientation. Cells overexpressing Bin1 grew more slowly than control cells and differentiated more rapidly when deprived of growth factors. In contrast, C2C12 cells expressing antisense Bin1 showed an impaired ability to undergo differentiation. Taken together, the results indicated that Bin1 expression, structure, and localization are tightly regulated during muscle differentiation and suggested that Bin1 plays a functional role in the differentiation process.