Genome-wide analysis of genetic alterations in acute lymphoblastic leukaemia

Chromosomal aberrations are a hallmark of acute lymphoblastic leukaemia (ALL) but alone fail to induce leukaemia. To identify cooperating oncogenic lesions, we performed a genome-wide analysis of leukaemic cells from 242 paediatric ALL patients using high-resolution, single-nucleotide polymorphism arrays and genomic DNA sequencing. Our analyses revealed deletion, amplification, point mutation and structural rearrangement in genes encoding principal regulators of B lymphocyte development and differentiation in 40% of B-progenitor ALL cases. The PAX5 gene was the most frequent target of somatic mutation, being altered in 31.7% of cases. The identified PAX5 mutations resulted in reduced levels of PAX5 protein or the generation of hypomorphic alleles. Deletions were also detected in TCF3 (also known as E2A), EBF1, LEF1, IKZF1 (IKAROS) and IKZF3 (AIOLOS). These findings suggest that direct disruption of pathways controlling B-cell development and differentiation contributes to B-progenitor ALL pathogenesis. Moreover, these data demonstrate the power of high-resolution, genome-wide approaches to identify new molecular lesions in cancer.     

Notch1 functions as a tumor suppressor in mouse skin

Notch proteins are important in binary cell-fate decisions and inhibiting differentiation in many developmental systems, and aberrant Notch signaling is associated with tumorigenesis. The role of Notch signaling in mammalian skin is less well characterized and is mainly based on in vitro studies, which suggest that Notch signaling induces differentiation in mammalian skin. Conventional gene targeting is not applicable to establishing the role of Notch receptors or ligands in the skin because Notch1-/- embryos die during gestation. Therefore, we used a tissue-specific inducible gene-targeting approach to study the physiological role of the Notch1 receptor in the mouse epidermis and the corneal epithelium of adult mice. Unexpectedly, ablation of Notch1 results in epidermal and corneal hyperplasia followed by the development of skin tumors and facilitated chemical-induced skin carcinogenesis. Notch1 deficiency in skin and in primary keratinocytes results in increased and sustained expression of Gli2, causing the development of basal-cell carcinoma-like tumors. Furthermore, Notch1 inactivation in the epidermis results in derepressed beta-catenin signaling in cells that should normally undergo differentiation. Enhanced beta-catenin signaling can be reversed by re-introduction of a dominant active form of the Notch1 receptor. This leads to a reduction in the signaling-competent pool of beta-catenin, indicating that Notch1 can inhibit beta-catenin-mediated signaling. Our results indicate that Notch1 functions as a tumor-suppressor gene in mammalian skin.     

An endoribonuclease-prepared siRNA screen in human cells identifies genes essential for cell division

RNA interference (RNAi) is an evolutionarily conserved defence mechanism whereby genes are specifically silenced through degradation of messenger RNAs; this process is mediated by homologous double-stranded (ds)RNA molecules. In invertebrates, long dsRNAs have been used for genome-wide screens and have provided insights into gene functions. Because long dsRNA triggers a nonspecific interferon response in many vertebrates, short interfering (si)RNA or short hairpin (sh)RNAs must be used for these organisms to ensure specific gene silencing. Here we report the generation of a genome-scale library of endoribonuclease-prepared short interfering (esi)RNAs from a sequence-verified complementary DNA collection representing 15,497 human genes. We used 5,305 esiRNAs from this library to screen for genes required for cell division in HeLa cells. Using a primary high-throughput cell viability screen followed by a secondary high content videomicroscopy assay, we identified 37 genes required for cell division. These include several splicing factors for which knockdown generates mitotic spindle defects. In addition, a putative nuclear-export terminator was found to speed up cell proliferation and mitotic progression after knockdown. Thus, our study uncovers new aspects of cell division and establishes esiRNA as a versatile approach for genomic RNAi screens in mammalian cells.     

Brain homeostasis: VEGF receptor 1 and 2—two unequal brothers in mind

Vascular endothelial growth factors (VEGFs), initially thought to act specifically on the vascular system, exert trophic effects on neural cells during development and adulthood. Therefore, the VEGF system serves as a promising therapeutic target for brain pathologies, but its simultaneous action on vascular cells paves the way for harmful side effects. To circumvent these deleterious effects, many studies have aimed to clarify whether VEGFs directly affect neural cells or if the effects are mediated secondarily via other cell types, like vascular cells. A great number of reports have shown the expression and function of VEGF receptors (VEGFRs), mainly VEGFR-1 and -2, in neural cells, where VEGFR-2 has been described as the major mediator of VEGF-A signals. This review aims to summarize and compare the divergent roles of VEGFR-1 and -2 during CNS development and homeostasis.     

Notch and cancer: a double-edged sword

The highly conserved Notch signaling pathway plays pleiotropic roles during embryonic development and is important for the regulation of selfrenewing tissues. The physiological functions of this signaling cascade range from stem cell maintenance and influencing cell fate decisions of barely differentiated progenitor cells, to the induction of terminal differentiation processes, all of which have been found to be recapitulated in different forms of cancers. Although Notch signaling has mostly been associated with oncogenic and growth-promoting roles, depending on the tissue type it can also function as a tumor suppressor. Here we describe recent findings on Notch signaling in cancer and tumor angiogenesis, and highlight some of the therapeutic approaches that are currently being developed to interfere with tumor growth and progression. Received 2 April 2007; received after revision 29 June 2007; accepted 2 July 2007     

Notch-dependent VEGFR3 upregulation allows angiogenesis without VEGF-VEGFR2 signalling

Developing tissues and growing tumours produce vascular endothelial growth factors (VEGFs), leading to the activation of the corresponding receptors in endothelial cells. The resultant angiogenic expansion of the local vasculature can promote physiological and pathological growth processes. Previous work has uncovered that the VEGF and Notch pathways are tightly linked. Signalling triggered by VEGF-A (also known as VEGF) has been shown to induce expression of the Notch ligand DLL4 in angiogenic vessels and, most prominently, in the tip of endothelial sprouts. DLL4 activates Notch in adjacent cells, which suppresses the expression of VEGF receptors and thereby restrains endothelial sprouting and proliferation. Here we show, by using inducible loss-of-function genetics in combination with inhibitors in vivo, that DLL4 protein expression in retinal tip cells is only weakly modulated by VEGFR2 signalling. Surprisingly, Notch inhibition also had no significant impact on VEGFR2 expression and induced deregulated endothelial sprouting and proliferation even in the absence of VEGFR2, which is the most important VEGF-A receptor and is considered to be indispensable for these processes. By contrast, VEGFR3, the main receptor for VEGF-C, was strongly modulated by Notch. VEGFR3 kinase-activity inhibitors but not ligand-blocking antibodies suppressed the sprouting of endothelial cells that had low Notch signalling activity. Our results establish that VEGFR2 and VEGFR3 are regulated in a highly differential manner by Notch. We propose that successful anti-angiogenic targeting of these receptors and their ligands will strongly depend on the status of endothelial Notch signalling.     

Maternal signals for progeny prevention against allergy and asthma

Allergy and asthma are chronic inflammatory diseases which result from complex gene–environment interactions. Recent evidence indicates the importance of prenatal and postnatal developmental processes in terms of maturation of balanced immune responses. According to the current view, gene–environment interactions during a restricted time frame are responsible for programming of the immune system in favor of allergic immune mechanisms later in life. The interaction between genes and environment is complex and only partially understood; however, heritable epigenetic modifications including chemical additions in and alternative packaging of the DNA have been shown to play a crucial role in this context. Novel data indicate that epigenetic mechanisms contribute to the development of T-helper cell function. Environmental factors, including diesel exhaust particles (DEP), vitamins and tobacco smoke, operate through such mechanisms. Furthermore, the role of environmental microbes provides another and maybe even more important group of exogenous exposures which operates in this critical time frame.