Microarray analysis to find new Wingless targets in Drosophila Wing Imaginal disc

Trabajo presentado en el 23rd European Drosophila Research Congress (EDRC 2013), celebrado en Barcelona del 16 al 19 de octubre de 2013.The formation of complex structures from undifferentiated cells has always been a major question for developmental biologists. One good model to study the developmental processes that take part in organ formation is Drosophila melanogaster. In Drosophila, adult wings arise from larval imaginal discs: single-layered sheets of undifferentiated cells. Many signalling pathways play a role in wing formation, the Wingless (Wg) pathway among them. Wg is a secreted protein that binds to its receptor Frizzled at the receiving cells, activating the pathway that ultimately leads to the translocation of the -catenin Armadillo (Arm) to the nucleus, activating the targets of the pathway. Depleting Wg from the wing imaginal disc leads to adult flies without wings. On the contrary, Wg overexpression effects are level-dependant whereas mild levels cause overgrown wings, high levels cause smaller over-differentiated wings. Thus, the mechanisms by which Wg signalling orchestrates wing formation and the effects of Wg signalling in cell differentiation and growth have yet to be elucidated. In our work, we performed microarray analysis of three different conditions for Wg: depletion of Wg, Wg mild overexpression and overexpression of a constitutively activated Arm; in order to identify targets of the pathway for each condition. About 300 genes were differentially expressed in almost one of the conditions; transcription factors, adhesion-related and programmed cell death related-genes, among others. This analysis has been useful to find new targets of the pathway and to be one step closer to answer the question of how Wg regulates morphogenesis.N

FGF coordinates air sac development by activation of the EGF ligand Vein through the transcription factor PntP2

How several signaling pathways are coordinated to generate complex organs through regulation of tissue growth and patterning is a fundamental question in developmental biology. The larval trachea of Drosophila is composed of differentiated functional cells and groups of imaginal tracheoblasts that build the adult trachea during metamorphosis. Air sac primordium cells (ASP) are tracheal imaginal cells that form the dorsal air sacs that supply oxygen to the flight muscles of the Drosophila adult. The ASP emerges from the tracheal branch that connects to the wing disc by the activation of both Bnl-FGF/Btl and EGFR signaling pathways. Together, these pathways promote cell migration and proliferation. In this study we demonstrate that Vein (vn) is the EGF ligand responsible for the activation of the EGFR pathway in the ASP. We also find that the Bnl-FGF/Btl pathway regulates the expression of vn through the transcription factor PointedP2 (PntP2). Furthermore, we show that the FGF target gene escargot (esg) attenuates EGFR signaling at the tip cells of the developing ASP, reducing their mitotic rate to allow proper migration. Altogether, our results reveal a link between Bnl-FGF/Btl and EGFR signaling and provide novel insight into how the crosstalk of these pathways regulates migration and growth.J.C. was supported by BFU2009-08748 from the Spanish MICINN and N.B-R. was supported by FPI from the Spanish MICINN. This work was funded by the Spanish Ministerio de Ciencia e Innovacion (Project BFU2009-08748).Peer reviewe

Contextual cues from cancer cells govern cancer-associated fibroblast heterogeneity

Cancer cells function as primary architects of the tumor microenvironment. However, the molecular features of cancer cells that govern stromal cell phenotypes remain unclear. Here, we show that cancer-associated fibroblast (CAF) heterogeneity is driven by lung adenocarcinoma (LUAD) cells at either end of the epithelial-to-mesenchymal transition (EMT) spectrum. LUAD cells that have high expression of the EMT-activating transcription factor ZEB1 reprogram CAFs through a ZEB1-dependent secretory program and direct CAFs to the tips of invasive projections through a ZEB1-driven CAF repulsion process. The EMT, in turn, sensitizes LUAD cells to pro-metastatic signals from CAFs. Thus, CAFs respond to contextual cues from LUAD cells to promote metastasis

Cancer becomes wasteful: emerging roles of exosomes in cell-fate determination

Extracellular vesicles (EVs), including exosomes, have been widely recognized for their role in intercellular communication of the immune response system. In the past few years, significance has been given to exosomes in the induction and modulation of cell-fate-inducing signalling pathways, such as the Hedgehog (Hh), Wnts, Notch, transforming growth factor (TGF-β), epidermal growth factor (EGF) and fibroblast growth factor (FGF) pathways, placing them in the wider context of development and also of cancer. These protein families induce signalling cascades responsible for tissue specification, homeostasis and maintenance. Exosomes contribute to cell-fate signal secretion, and vice versa exosome secretion can be induced by these proteins. Interestingly, exosomes can also transfer their mRNA to host cells or modulate the signalling pathways directly by the removal of downstream effector molecules from the cell. Surprisingly, much of what we know about the function of exosomes in cell determination is gathered from pathological transformed cancer cells and wound healing while data about their biogenesis and biology in normal developing and adult tissue lag behind. In this report, we will summarize some of the published literature and point to current advances and questions in this fast-developing topic. In a brief foray, we will also update and shortly discuss their potential in diagnosis and targeted cancer treatment

Pathology and Classification of SCLC

Lung cancer is consistently the leading cause of cancer-related death worldwide, and it ranks as the second most frequent type of new cancer cases diagnosed in the United States, both in males and females. One subtype of lung cancer, small cell lung carcinoma (SCLC), is an aggressive, poorly differentiated, and high-grade neuroendocrine carcinoma that accounts for 13% of all lung carcinomas. SCLC is the most frequent neuroendocrine lung tumor, and it is commonly presented as an advanced stage disease in heavy smokers. Due to its clinical presentation, it is typically diagnosed in small biopsies or cytology specimens, with routine immunostaining only. However, immunohistochemistry markers are extremely valuable in demonstrating neuroendocrine features of SCLC and supporting its differential diagnosis. The 2015 WHO classification grouped all pulmonary neuroendocrine carcinomas in one category and maintained the SCLC combined variant that was previously recognized. In this review, we explore multiple aspects of the pathologic features of this entity, as well as clinically relevant immunohistochemistry markers expression and its molecular characteristics. In addition, we will focus on characteristics of the tumor microenvironment, and the latest pathogenesis findings to better understand the new therapeutic options in the current era of personalized therapy

The role of tarsal-less in Drosophila wing imaginal disc development

Trabajo presentado en el 1st PhD and Postdoc Symposium (From Cells to Organs), celebrado en Barcelona el 16 y 17 de abril de 2012.N

Phosphorylation at ser-181 of oncogenic KRAS is required for tumor growth

KRAS phosphorylation has been reported recently to modulate the activity of mutant KRAS protein in vitro. In this study, we defined S181 as a specific phosphorylation site required to license the oncogenic function of mutant KRAS in vivo. The phosphomutant S181A failed to induce tumors in mice, whereas the phosphomimetic mutant S181D exhibited an enhanced tumor formation capacity, compared with the wild-type KRAS protein. Reduced growth of tumors composed of cells expressing the nonphosphorylatable KRAS S181A mutant was correlated with increased apoptosis. Conversely, increased growth of tumors composed of cells expressing the phosphomimetic KRAS S181D mutant was correlated with increased activation of AKT and ERK, two major downstream effectors of KRAS. Pharmacologic treatment with PKC inhibitors impaired tumor growth associated with reduced levels of phosphorylated KRAS and reduced effector activation. In a panel of human tumor cell lines expressing various KRAS isoforms, we showed that KRAS phosphorylation was essential for survival and tumorigenic activity. Furthermore, we identified phosphorylated KRAS in a panel of primary human pancreatic tumors. Taken together, our findings establish that KRAS requires S181 phosphorylation to manifest its oncogenic properties, implying that its inhibition represents a relevant target to attack KRAS-driven tumors. © 2014 American Association for Cancer Research.This study was supported by MICINN-Spain (SAF2010-20712) and RTICC (MINECO-Spain; groups RD 12/0036/0049 and RD 12/0036/0008). C. Barceló and N. Paco are recipients of predoctoral fellowships from MEC-Spain and the Catalan Government, respectively.http://dx.doi.org/10.1158/0008-5472.CAN-13-1750Peer Reviewe

Pro-metastatic collagen lysyl hydroxylase dimer assemblies stabilized by Fe2+-binding

Collagen lysyl hydroxylases promote cancer progression. Here the authors present the crystal structure of the lysyl hydroxylase domain of L230 from Acanthamoeba polyphaga mimivirus, which is of interest for LH inhibitor development, and show that ectopic expression of L230 in tumors promotes collagen cross-linking and metastasis

A collagen glucosyltransferase drives lung adenocarcinoma progression in mice.

Cancer cells are a major source of enzymes that modify collagen to create a stiff, fibrotic tumor stroma. High collagen lysyl hydroxylase 2 (LH2) expression promotes metastasis and is correlated with shorter survival in lung adenocarcinoma (LUAD) and other tumor types. LH2 hydroxylates lysine (Lys) residues on fibrillar collagens amino- and carboxy-terminal telopeptides to create stable collagen cross-links. Here, we show that electrostatic interactions between the LH domain active site and collagen determine the unique telopeptidyl lysyl hydroxylase (tLH) activity of LH2. However, CRISPR/Cas-9-mediated inactivation of tLH activity does not fully recapitulate the inhibitory effect of LH2 knock out on LUAD growth and metastasis in mice, suggesting that LH2 drives LUAD progression, in part, through a tLH-independent mechanism. Protein homology modeling and biochemical studies identify an LH2 isoform (LH2b) that has previously undetected collagen galactosylhydroxylysyl glucosyltransferase (GGT) activity determined by a loop that enhances UDP-glucose-binding in the GLT active site and is encoded by alternatively spliced exon 13 A. CRISPR/Cas-9-mediated deletion of exon 13 A sharply reduces the growth and metastasis of LH2b-expressing LUADs in mice. These findings identify a previously unrecognized collagen GGT activity that drives LUAD progression