Heparan sulfate regulates ADAM12 through a molecular switch mechanism.

The disintegrin and metalloproteases (ADAMs) are emerging as therapeutic targets in human disease, but specific drug design is hampered by potential redundancy. Unlike other metzincins, ADAM prodomains remain bound to the mature enzyme to regulate activity. Here ADAM12, a protease that promotes tumor progression and chondrocyte proliferation in osteoarthritic cartilage, is shown to possess a prodomain/catalytic domain cationic molecular switch, regulated by exogenous heparan sulfate and heparin but also endogenous cell surface proteoglycans and the polyanion, calcium pentosan polysulfate. Sheddase functions of ADAM12 are regulated by the switch, as are proteolytic functions in placental tissue and sera of pregnant women. Moreover, human heparanase, an enzyme also linked to tumorigenesis, can promote ADAM12 sheddase activity at the cell surface through cleavage of the inhibitory heparan sulfate. These data present a novel concept that might allow targeting of ADAM12 and suggest that other ADAMs may have specific regulatory activity embedded in their prodomain and catalytic domain structures.

The Notch pathway in cancer: differentiation gone awry.

The Notch signalling cascade influences several key aspects of normal development by regulating differentiation, proliferation and apoptosis. Its association to human cancer is firmly established in T-cell leukaemia where point mutations or chromosomal translocations lead to constitutive signalling. Accumulating data indicate that deregulated Notch activity is involved also in the genesis of other human cancers, such as pancreatic cancer, medulloblastoma and mucoepidermoid carcinoma. In these tumours, the oncogenic effect of Notch signalling reflects an aberrant recapitulation of the highly tissue-specific function of the cascade during normal development and in tissue homeostasis.

High ID2 protein expression correlates with a favourable prognosis in patients with primary breast cancer and reduces cellular invasiveness of breast cancer cells.

ID proteins have been implicated in the regulation of cell proliferation and differentiation in various cell types during normal development as well as in the formation of cancer. Our aim was to delineate the expression of ID2 by immunohistochemistry in primary breast cancer in order to detect potential associations with cell cycle regulatory proteins and/or clinicopathologic parameters. We further overexpressed ID2 in a breast cancer cell line to elaborate potential effects on proliferation and invasiveness. We observed large variations in ID2 expression in primary breast cancer, and the protein was localised to both the nucleus and cytoplasm. Interestingly, a high cytoplasmic ID2 protein level correlated with a favourable prognosis. Overexpressing ID2 in the MDA-MB-468 breast cancer cell line generated a marked cytoplasmic localisation of the protein and reduced the invasive capacity of cells. Modest enhancement of cell proliferation was further detected in ID2-overexpressing cells. In conclusion, ID2 protein expression varies substantially within primary breast tumours and high cytoplasmic levels of ID2 might reflect a less aggressive breast tumour phenotype.

Neuroblastoma and pre-B lymphoma cells share expression of key transcription factors but display tissue restricted target gene expression.

BACKGROUND: Transcription factors are frequently involved in the process of cellular transformation, and many malignancies are characterized by a distinct genetic event affecting a specific transcription factor. This probably reflects a tissue specific ability of transcription factors to contribute to the generation of cancer but very little is known about the precise mechanisms that governs these restricted effects. METHODS: To investigate this selectivity in target gene activation we compared the overall gene expression patterns by micro-array analysis and expression of target genes for the transcription factor EBF in lymphoma and neuroblastoma cells by RT-PCR. The presence of transcription factors in the different model cell lines was further investigated by EMSA analysis. RESULTS: In pre-B cells mb-1 and CD19 are regulate by EBF-1 in collaboration with Pax-5 and E-proteins. We here show that neuroblastoma cells express these three, for B cell development crucial transcription factors, but nevertheless fail to express detectable levels of their known target genes. Expression of mb-1 could, however, be induced in neuroblastoma cells after disruption of the chromatin structure by treatment with 5-azacytidine and Trichostatin A. CONCLUSION: These data suggest that transcription factors are able to selectively activate target genes in different tissues and that chromatin structure plays a key role in the regulation of this activity.

Olf/EBF proteins are expressed in neuroblastoma cells: potential regulators of the Chromogranin A and SCG10 promoters.

The childhood malignancy neuroblastoma is derived from developmentally arrested sympathetic nervous system precursor cells. To obtain further insight into the molecular processes involved in the formation of these tumors, we decided to investigate the functional role of Olf/EBF (O/E) transcription factors in human neuroblastoma cells. We here report that O/E-1 and O/E-2 are expressed at variable levels in neuroblastoma cell lines and that O/E proteins could be identified by electrophoretic mobility shift assays. To identify potential neuronal target genes for O/E proteins in neuroblastoma cells we investigated the ability of a set of neuronal promoters to interact with O/E-1 in electrophoretic mobility shift assays. This analysis suggested that the Chromogranin A (CgA) and SCG10 promoters both contained binding sites for O/E-1. O/E-1 was able to activate the CgA promoter in vivo and mutation of the O/E-1 binding site in the CgA promoter reduced the functional activity of the element to about 60% of the wild-type in neuroblastoma cells, supporting the idea that O/E proteins may be involved in the control of the CgA promoter. Furthermore, overexpression of O/E-1 in hippocampal progenitor cells led to neurite outgrowth, indicative of a role for O/E proteins in neuronal differentiation.

The basic helix-loop-helix transcription factor TAL1/SCL inhibits the expression of the p16INK4A and pTalpha genes.

The Tal1 gene (also called Scl or TCL5) encodes a basic helix-loop-helix transcription factor required for hematopoiesis and vasculogenesis. Additionally, aberrant transcriptional activation of the Tal1 gene is a frequent event in human T cell acute lymphoblastic leukemia (T-ALL). T cell specific expression of TAL1 in mice induces aggressive T cell malignancies, demonstrating the oncogenic potential of TAL1. Yet, the underlying mechanisms of TAL1 induced tumorigenesis are poorly understood. By inhibiting E protein mediated transcription of the pTalpha gene, TAL1 can interfere with the T cell differentiation program. In addition, several studies suggest that TAL1 expression might also enhance proliferation rate. We report here that TAL1 can bind the E boxes in both the p16 and the pTalpha promoters, and functionally suppress the activity of both promoters. These results indicate that TAL1 can affect both T cell proliferation and differentiation. Moreover, we show that overexpression of TAL1 in hematopoietic progenitor cells promotes cell cycle division.

The LIM-only protein LMO4 modulates the transcriptional activity of HEN1.

The basic helix-loop-helix protein HEN1 and the LIM-only proteins LMO2 and LMO4 are expressed in neuronal cells. HEN1 was cloned by virtue of its homology to TAL1, a bHLH protein important for early hematopoiesis. Since it has been shown that TAL1 forms complex with LMO proteins in erythroid and leukemic cells we investigated the capacity of HEN1 to form complex with LMO2 and LMO4. By mammalian two-hybrid analysis, we show that HEN1 interacts with both LMO2 and LMO4. To characterize the transcriptional capacity of HEN1 alone or together with LMO2 and LMO4, we performed reporter gene assays. In comparison with the ubiquitously expressed bHLH protein E47, HEN1 is a very modest transcriptional activator and titration experiments indicate that HEN1, like TAL1, represses E47 mediated transcriptional activation. Furthermore, LMO4 but not LMO2 was able to augment this effect. Overexpression of HEN1 in hippocampal precursor cells resulted in neurite extension, which could be prevented by LMO4. Taken together, these results indicate that LMO proteins can modulate the transcriptional activity of HEN1.