The endocytic matrix.

Endocytosis has long been thought of as simply a way for cells to internalize nutrients and membrane-associated molecules. But an explosive growth in knowledge has given a new dimension to our understanding of this process. It now seems that endocytosis is a master organizer of signalling circuits, with one of its main roles being the resolution of signals in space and time.

Disease modeling in zebrafish: cancer and immune responses--a report on a workshop held in Spoleto, Italy, July 20-22, 2009.

The growing interest in using zebrafish for genetic and functional dissection of malignancy and infection was highlighted by the second international workshop on Zebrafish Models of Cancer and the Immune Response in Spoleto, Italy (July 20-22, 2009). The overarching theme of the state-of-the-art reports featured the unique suitability of zebrafish for in vivo monitoring of fundamental biologic and pathologic processes. For example, in vivo imaging was employed for the first demonstration of direct development of hematopoietic stem cells from hemogenic epithelium and for visualization of T-cell homing and interaction with thymic epithelial cells.

Global repression of cancer gene expression in a zebrafish model of melanoma is linked to epigenetic regulation.

We have established a model of melanoma progression in zebrafish through the generation of transgenic lines specifically expressing oncogenic human HRAS in the melanocytic lineage. In these tumors we have carried out quantitative expression analysis of several putative cancer genes, from known and predicted cancer gene lists. In particular, we analyzed 39 out of 101 putative cancer genes identified with a bioinformatics approach and selected for the low frequency of duplication and the high connectivity in protein networks.

SASPense and DDRama in cancer and ageing.

Senescent cells alter their microenvironment by secreting a growing collection of factors, a phenomenon termed the senescence-associated secretory phenotype (SASP). Cellular senescence is often the result of nuclear DNA damage fuelling a chronic DNA damage response (DDR). Upstream elements of the DDR cascade are necessary for full blown SASP, and additional crosstalk occurs between the DDR and cytokine secretion.

The checkpoint response to replication stress.

Genome instability is a hallmark of cancer cells, and defective DNA replication, repair and recombination have been linked to its etiology. Increasing evidence suggests that proteins influencing S-phase processes such as replication fork movement and stability, repair events and replication completion, have significant roles in maintaining genome stability. DNA damage and replication stress activate a signal transduction cascade, often referred to as the checkpoint response.

Replicon dynamics, dormant origin firing, and terminal fork integrity after double-strand break formation.

In response to replication stress, the Mec1/ATR and SUMO pathways control stalled- and damaged-fork stability. We investigated the S phase response at forks encountering a broken template (termed the terminal fork). We show that double-strand break (DSB) formation can locally trigger dormant origin firing.

Transcription factor binding sites are genetic determinants of retroviral integration in the human genome.

Gamma-retroviruses and lentiviruses integrate non-randomly in mammalian genomes, with specific preferences for active chromatin, promoters and regulatory regions. Gene transfer vectors derived from gamma-retroviruses target at high frequency genes involved in the control of growth, development and differentiation of the target cell, and may induce insertional tumors or pre-neoplastic clonal expansions in patients treated by gene therapy. The gene expression program of the target cell is apparently instrumental in directing gamma-retroviral integration, although the molecular basis of this phenomenon is poorly understood.

The Saccharomyces cerevisiae Esc2 and Smc5-6 proteins promote sister chromatid junction-mediated intra-S repair.

Recombination is important for DNA repair, but it can also contribute to genome rearrangements. RecQ helicases, including yeast Sgs1 and human BLM, safeguard genome integrity through their functions in DNA recombination. Sgs1 prevents the accumulation of Rad51-dependent sister chromatid junctions at damaged replication forks, and its functionality seems to be regulated by Ubc9- and Mms21-dependent sumoylation.

Expression of H-RASV12 in a zebrafish model of Costello syndrome causes cellular senescence in adult proliferating cells.

Constitutively active, 'oncogenic' H-RAS can drive proliferation and transformation in human cancer, or be a potent inducer of cellular senescence. Moreover, aberrant activation of the Ras pathway owing to germline mutations can cause severe developmental disorders. In this study we have generated transgenic zebrafish that constitutively express low levels, or can be induced to express high levels, of oncogenic H-RAS.

SUMOylation regulates Rad18-mediated template switch.

Replication by template switch is thought to mediate DNA damage-bypass and fillings of gaps. Gap-filling repair requires homologous recombination as well as Rad18- and Rad5-mediated proliferating cell nuclear antigen (PCNA) polyubiquitylation. However, it is unclear whether these processes are coordinated, and the physical evidence for Rad18-Rad5-dependent template switch at replication forks is still elusive.

Ubiquitin in trafficking: the network at work.

Targeting of membrane proteins to their proper destination requires specific mechanisms. Protein cargos are included in vesicles that bud off a donor organelle and ultimately fuse with a target organelle, where the cargos are delivered. Endocytosis of transmembrane receptors (e.

DNA damage response activation in mouse embryonic fibroblasts undergoing replicative senescence and following spontaneous immortalization.

Primary mouse embryonic fibroblasts (MEFs) are a popular tool for molecular and cell biology studies. However, when MEFs are grown in vitro under standard tissue culture conditions, they proliferate only for a limited number of population doublings (PD) and eventually undergo cellular senescence. Presently, the molecular mechanisms halting cell cycle progression and establishing cellular senescence under these conditions are unclear.

Alignment of protein structures in the presence of domain motions.

Background: Structural alignment is an important step in protein comparison. Well-established methods exist for solving this problem under the assumption that the structures under comparison are considered as rigid bodies. However, proteins are flexible entities often undergoing movements that alter the positions of domains or subdomains with respect to each other.

Living on a break: cellular senescence as a DNA-damage response.

Cellular senescence is associated with ageing and cancer in vivo and has a proven tumour-suppressive function. Common to both ageing and cancer is the generation of DNA damage and the engagement of the DNA-damage response pathways. In this Review, the diverse mechanisms that lead to DNA-damage generation and the activation of DNA-damage-response signalling pathways are discussed, together with the evidence for their contribution to the establishment and maintenance of cellular senescence in the context of organismal ageing and cancer development.

RAPIDO: a web server for the alignment of protein structures in the presence of conformational changes.

Rapid alignment of proteins in terms of domains (RAPIDO) is a web server for the 3D alignment of crystal structures of different protein molecules in the presence of conformational change. The structural alignment algorithm identifies groups of equivalent atoms whose interatomic distances are constant (within a defined tolerance) in the two structures being compared and considers these groups of atoms as rigid bodies. In addition to the functionalities provided by existing tools, RAPIDO can identify structurally equivalent regions also when these consist of fragments that are distant in terms of sequence and separated by other movable domains.

How to create the vascular tree? (Latest) help from the zebrafish.

The cardiovascular system provides oxygen, nutrients and hormones to organs, it directs traffic of metabolites and it maintains tissue homeostasis. It is one of the first organs assembled during vertebrate development and it is essential to life from early stages to adult. For these reasons, the process of vessel formation has being studied for more than a century, but it is only in the late eighties that there has been an explosion of research in the field with the employment of various in vitro and in vivo model systems.

Regulation of DNA repair throughout the cell cycle.

The repair of DNA lesions that occur endogenously or in response to diverse genotoxic stresses is indispensable for genome integrity. DNA lesions activate checkpoint pathways that regulate specific DNA-repair mechanisms in the different phases of the cell cycle. Checkpoint-arrested cells resume cell-cycle progression once damage has been repaired, whereas cells with unrepairable DNA lesions undergo permanent cell-cycle arrest or apoptosis.

IRSp53: crossing the road of membrane and actin dynamics in the formation of membrane protrusions.

A tight spatiotemporal coordination of the machineries controlling membrane bending and trafficking, and actin dynamics is crucial for the generation of cellular protrusions. Proteins that are simultaneously capable of regulating actin dynamics and sensing or inducing membrane curvature are predicted to have a prominent role. A prototypical example of this type of proteins is the insulin receptor tyrosine kinase substrate of 53kDa, the founding member of a recently discovered family of proteins, including missing-in-metastasis and ABBA (actin-bundling protein with BAIAP2 homology).

The primate-specific protein TBC1D3 is required for optimal macropinocytosis in a novel ARF6-dependent pathway.

The generation of novel genes and proteins throughout evolution has been proposed to occur as a result of whole genome and gene duplications, exon shuffling, and retrotransposition events. The analysis of such genes might thus shed light into the functional complexity associated with highly evolved species. One such case is represented by TBC1D3, a primate-specific gene, harboring a TBC domain.

NUMB controls p53 tumour suppressor activity.

NUMB is a cell fate determinant, which, by asymmetrically partitioning at mitosis, controls cell fate choices by antagonising the activity of the plasma membrane receptor of the NOTCH family. NUMB is also an endocytic protein, and the NOTCH-NUMB counteraction has been linked to this function. There might be, however, additional functions of NUMB, as witnessed by its proposed role as a tumour suppressor in breast cancer.

Topological properties of co-occurrence networks in published gene expression signatures.

Meta-analysis of high-throughput gene expression data is often used for the interpretation of proprietary gene expression data sets. We have recently shown that co-occurrence patterns of gene expression in published cancer-related gene expression signatures are reminiscent of several cancer signaling pathways. Indeed, significant co-occurrence of up to ten genes in published gene expression signatures can be exploited to build a co-occurrence network from the sets of co-occurring genes ("co-occurrence modules").

Template switching: from replication fork repair to genome rearrangements.

Genome rearrangements are a hallmark of human genomic disorders and occur largely through recombination mechanisms. In this issue, Lee et al. (2007) show that the complex nonrecurrent rearrangements observed in the dysmyelinating disorder Pelizaeus-Merzbacher disease (PMD) are likely to be caused by a replication mechanism involving template switching.

Breaking news: high-speed race ends in arrest--how oncogenes induce senescence.

Oncogene activation in normal cells induces a permanent proliferative arrest known as cellular senescence. This phenomenon restrains the expansion of cells that bear an activated oncogene and acts as a powerful tumor-suppressive process. Although the full molecular mechanisms are still being elucidated, it has been observed recently that some oncogenes alter the DNA-replication process and cause DNA-damage accumulation.

Top1- and Top2-mediated topological transitions at replication forks ensure fork progression and stability and prevent DNA damage checkpoint activation.

DNA topoisomerases solve topological problems during chromosome metabolism. We investigated where and when Top1 and Top2 are recruited on replicating chromosomes and how their inactivation affects fork integrity and DNA damage checkpoint activation. We show that, in the context of replicating chromatin, Top1 and Top2 act within a 600-base-pair (bp) region spanning the moving forks.