RECERCAT - Genètica

La Genética forense y sus aplicaciones

La Genética forense y sus aplicaciones Mestres Naval, Francesc La Genética forense es una disciplina que en la actualidad goza de una gran popularidad debido a su reiterada aparición en la prensa escrita, series de televisión y producciones cinematográfi cas. Aunque la palabra"forense" se asocia comúnmente con"médico forense" y el estudio de los cadáveres, técnicamente deriva de la palabra latina"forum", el foro, donde los romanos llevaban a cabo sus juicios. De forma que"forense" es un adjetivo que indica referencia a temas judiciales. Por tanto, la Genética forense no es más que la utilización de esta rama de la ciencia para resolver temas judiciales. Los ámbitos principales de actuación de la Genética forense son: la identificación de individuos para resolver diferentes tipos de delitos (asesinatos, robos, etc.), la identifi cación de individuos desaparecidos o de cuerpos seriamente dañados como consecuencia de una gran catástrofe (natural o producida por el hombre), los estudios de paternidad (o de otros grados de parentesco) y por último la identifi cación de diferentes especies o de individuos concretos de ciertas especies (para resolver temas de fraudes alimentarios, casos criminales, ataques terroristas mediante microorganismos, etc.).

RNA interference is ineffective as a routine method for gene silencing in chick embryos as monitored by fgf8 silencing.

RNA interference is ineffective as a routine method for gene silencing in chick embryos as monitored by fgf8 silencing. Hernández Hernández, Victor; Bueno i Torrens, David, 1965- The in vivo accessibility of the chick embryo makes it a favoured model system for experimental developmental biology. Although the range of available techniques now extends to miss-expression of genes through in ovo electroporation, it remains difficult to knock out individual gene expression. Recently, the possibility of silencing gene expression by RNAi in chick embryos has been reported. However, published studies show only discrete quantitative differences in the expression of the endogenous targeted genes and unclear morphological alterations. To elucidate whether the tools currently available are adequate to silence gene expression sufficiently to produce a clear and specific null-like mutant phenotype, we have performed several experiments with different molecules that trigger RNAi: dsRNA, siRNA, and shRNA produced from a plasmid coexpressing green fluorescent protein as an internal marker. Focussing on fgf8 expression in the developing isthmus, we show that no morphological defects are observed, and that fgf8 expression is neither silenced in embryos microinjected with dsRNA nor in embryos microinjected and electroporated with a pool of siRNAs. Moreover, fgf8 expression was not significantly silenced in most isthmic cells transformed with a plasmid producing engineered shRNAs to fgf8. We also show that siRNA molecules do not spread significantly from cell to cell as reported for invertebrates, suggesting the existence of molecular differences between different model systems that may explain the different responses to RNAi. Although our results are basically in agreement with previously reported studies, we suggest, in contrast to them, that with currently available tools and techniques the number of cells in which fgf8 gene expression is decreased, if any, is not sufficient to generate a detectable mutant phenotype, thus making RNAi useless as a routine method for functional gene analysis in chick embryos.

The embryonic blood-CSF barrier has molecular elements for specific glucose transport and for the general transport of molecules via transcellular routes.

The embryonic blood-CSF barrier has molecular elements for specific glucose transport and for the general transport of molecules via transcellular routes. Parvas, Maryam; Bueno i Torrens, David, 1965- In vertebrates, early brain development takes place at the expanded anterior end of the neural tube, which is filled with embryonic cerebrospinal fluid (E-CSF). We have recently identified a transient blood
CSF barrier that forms between embryonic days E3 and E4 in chick embryos and that is responsible for the transport of proteins and control of E-CSF homeostasis, including osmolarity. Here we examined the presence of glucose transporter GLUT-1 as well the presence of caveolae-structural protein Caveolin1 (CAV-1) in the embryonic blood-CSF barrier which may be involved in the transport of glucose and of proteins, water and ions respectively across the neuroectoderm. In this paper we demonstrate the presence of GLUT-1 and CAV-1 in endothelial cells of blood vessels as well as in adjacent neuroectodermal cells, located in the embryonic blood
CSF barrier. In blood vessels, these proteins were detected as early as E4 in chick embryos and E12.7 in rat embryos, i.e. the point at which the embryonic blood
CSF barrier acquires this function. In the neuroectoderm of the embryonic blood-CSF barrier, GLUT-1 was also detected at E4 and E12.7 respectively, and CAV-1 was detected shortly thereafter in both experimental models. These experiments contribute to delineating the extent to which the blood
CSF embryonic barrier controls E-CSF composition and homeostasis during early stages of brain development in avians and mammals. Our results suggest the regulation of glucose transport to the E-CSF by means of GLUT-1 and also suggest a mechanism by which proteins are transported via transcellular routes across the neuroectoderm, thus reinforcing the crucial role of E-CSF in brain development.

Disruption of embryonic blood-CSF barrier in chick embryos reveals the actual importance of this barrier to control E-CSF composition and homeostasis in early brain development

Disruption of embryonic blood-CSF barrier in chick embryos reveals the actual importance of this barrier to control E-CSF composition and homeostasis in early brain development Parvas, Maryam; Bueno Torrens, David, 1965- In vertebrates, early brain development takes place at the expanded anterior end of the neural tube. After closure of the anterior neuropore, the brain wall forms a physiologically sealed cavity that encloses embryonic cerebrospinal fluid (E-CSF), a complex and protein-rich fluid that is initially composed of trapped amniotic fluid. E-CSF has several crucial roles in brain anlagen development. Recently, we reported the presence of transient blood
CSF barrier located in the brain stem lateral to the ventral midline, at the mesencephalon and prosencephalon level, in chick and rat embryos by transporting proteins, water, ions and glucose in a selective manner via transcellular routes. To test the actual relevance of the control of E-CSF composition and homeostasis on early brain development by this embryonic blood-CSF barrier, we block the activity of this barrier by treating the embryos with 6-aminonicotinamide gliotoxin (6-AN). We demonstrate that 6-AN treatment in chick embryos blocks protein transport across the embryonic blood-CSF barrier, and that the disruption of the barrier properties is due to the cease transcellular caveolae transport, as detected by CAV-1 expression cease. We also show that the lack of protein transport across the embryonic blood-CSF barrier influences neuroepithelial cell survival, proliferation and neurogenesis, as monitored by neurepithelial progenitor cells survival, proliferation and neurogenesis. The blockage of embryonic blood-CSF transport also disrupts water influx to the E-CSF, as revealed by an abnormal increase in brain anlagen volume. These experiments contribute to delineate the actual extent of this blood-CSF embryonic barrier controlling E-CSF composition and homeostasis and the actual important of this control for early brain development, as well as to elucidate the mechanism by which proteins and water are transported thought transcellular routes across the neuroectoderm, reinforcing the crucial role of E-CSF for brain development.

El líquid cerebroespinal. Un fluid vital per al sistema nerviós central

El líquid cerebroespinal. Un fluid vital per al sistema nerviós central Bueno Torrens, David, 1965-; Parvas, Maryam Moltes civilitzacions s"han desenvolupat al costat de rius i mars, uns medis fluids que han servit de vehicle de cohesió i transport i que han afavorit la supervivència de les persones que vivien a les seves ribes. Per als egipcis va ser el Nil; per als mesopotamis, el Tigris i l"Eufrates, i per als grecs i fenicis, la Mediterrània. Un dels sistemes dels animals més complexos funcionalment és el nerviós, el qual assoleix l"expressió màxima en els vertebrats, molt especialment en òrgans com el cervell. Sorprenentment, el cervell també s"organitza, des de l"inici embrionari i durant tota la vida adulta, al voltant d"un fluid extraordinàriament dinàmic i complex: el líquid cerebrospinal.

Cèl·lules mare i malalties neurodegeneratives

Cèl·lules mare i malalties neurodegeneratives Parada, Carolina; Bueno Torrens, David, 1965- L"augment en l"esperança de vida va associat a un increment significatiu en la incidència de determinades malalties, entre les quals cal destacar les malalties neurodegeneratives per l"alt cost personal, social i econòmic que representen.

Most of the Abundant Protein Fractions of Embryonic Cerebrospinal Fluid are Produced Out of the Brain Anlagen

Most of the Abundant Protein Fractions of Embryonic Cerebrospinal Fluid are Produced Out of the Brain Anlagen Parvas, Maryam; Rius Viladomiu, Marc; Bueno Torrens, David, 1965- The microenvironment of the central nervous system is important for neuronal function and development. During the early stages of embryo development the cephalic vesicles are filled by embryonic cerebrospinal fluid, a complex fluid containing different protein fractions, which contributes to the regulation of the survival, proliferation and neurogenesis of neuroectodermal stem cells. The protein content of embryonic cerebrospinal fluid from chick and rat embryos at the start of neurogenesis has already been determined. Most of the identified gene products are thought to be involved in the regulation of developmental processes during embryogenesis. However, due to the crucial roles played by embryonic cerebrospinal fluid during brain development, the embryological origin of the gene products it contains remains an intriguing question. According to the literature most of these products are synthesised in embryonic tissues other than the neuroepithelium. In this study we examined the embryological origin of the most abundant embryonic cerebrospinal fluid protein fractions by means of slot-blot analysis and by using several different embryonic and extraembryonic protein extracts, immunodetected with polyclonal antibodies. This first attempt to elucidate their origin is not based on the proteins identified by proteomic methods, but rather on crude protein fractions detected by SDS-PAGE analysis and to which polyclonal antibodies were specifically generated. Despite some of the limitations of this study, i.e. that one protein fraction may contain more than one gene product, and that a specific gene product may be contained in different protein fractions depending on post-translational modifications, our results show that most of the analysed protein fractions are not produced by the cephalic neuroectoderm but are rather stored in the egg reservoir; furthermore, few are produced by embryo tissues, thus indicating that they must be transported from their production or storage sites to the cephalic cavities, most probably via embryonic serum. These results raise the question as to whether the transfer of proteins from these two embryo compartments is regulated at this early developmental stage.

Difusió de la ciència: repte i responsabilitat

Difusió de la ciència: repte i responsabilitat Bueno Torrens, David, 1965- La ciència, definida com el conjunt de coneixences d"un cert ordre de coses, i la tècnica, que es nodreix dels avenços científics, tenen un impacte cabdal en tots els aspectes de la nostra vida, i hi confiem bona part del nostre futur. Només cal que fem una ullada al nostre voltant. Des dels objectes més quotidians fins als més sofisticats tractaments biomèdics tenen el seu origen en la ciència o bé en són conseqüència. Paradoxalment, però, per a moltes persones la ciència és una gran desconeguda, la qual cosa genera incomptables contradiccions. D"una banda, perquè de vegades
o sovint
la tècnica es percep com un element deshumanitzador, i hi ha qui associa progrés a destrucció, malgrat pràcticament ningú renunciï al progrés. D"altra, perquè no sempre resulta senzill explicar conceptes aparentment complexos amb un llenguatge planer sense que es perdi rigor i precisió, i sense induir interpretacions capcioses. És aquí on la difusió de la ciència troba el seu camp natural d"actuació, com el repte d"assolir el nivell adequat i també com la responsabilitat del divulgador envers la societat.

Aggressivity, violence, sociability and conflict resolution: What genes can tell us.

Aggressivity, violence, sociability and conflict resolution: What genes can tell us. Bueno Torrens, David, 1965- Conflicts are inherent to the human condition, as they are for all living beings. Disputes about resources or access to mating partners are among the most common causes of conflict. Conflict is herein defined as a struggle or contest between individuals or parties, and may involve a variety of aggressive behaviours. In humans, aggressiveness, violence and conflicts, including individual predisposal to conflict resolution, have traditionally been said to have deep cultural roots, but recent research in both neuroscience and genetics has shown the influence of genes on such complex behavioural traits. In this paper, recent data on the genetic aspects of these interrelated behaviours will be put together, including the effects of particular genes, the influence of stress and gender on gene regulation, and gene
environment interactions, all of which may influence biological predisposal to conflict resolution. Other genetically influenced behavioural aspects involved in conflicts and conflict resolution, such as sociability, will also be discussed. The importance of taking into account genetic and biological data to provide strategies for conflict resolution will be highlighted.

Sex Reversal in Zebrafish fancl Mutants is Caused by Tp53-Mediated Germ Cell Apoptosis

Sex Reversal in Zebrafish fancl Mutants is Caused by Tp53-Mediated Germ Cell Apoptosis Rodríguez Marí, Adriana; Cañestro García, Cristian; BreMiller, R.A.; Nguyen-Johnson, A.; Asakawa, K.; Kawakami, K.; Postlethwait, J.H. The molecular genetic mechanisms of sex determination are not known for most vertebrates, including zebrafish. We identified a mutation in the zebrafish fancl gene that causes homozygous mutants to develop as fertile males due to female-to-male sex reversal. Fancl is a member of the Fanconi Anemia/BRCA DNA repair pathway. Experiments showed that zebrafish fancl was expressed in developing germ cells in bipotential gonads at the critical time of sexual fate determination. Caspase-3 immunoassays revealed increased germ cell apoptosis in fancl mutants that compromised oocyte survival. In the absence of oocytes surviving through meiosis, somatic cells of mutant gonads did not maintain expression of the ovary gene cyp19a1a and did not down-regulate expression of the early testis gene amh; consequently, gonads masculinized and became testes. Remarkably, results showed that the introduction of a tp53 (p53) mutation into fancl mutants rescued the sex-reversal phenotype by reducing germ cell apoptosis and, thus, allowed fancl mutants to become fertile females. Our results show that Fancl function is not essential for spermatogonia and oogonia to become sperm or mature oocytes, but instead suggest that Fancl function is involved in the survival of developing oocytes through meiosis. This work reveals that Tp53-mediated germ cell apoptosis induces sex reversal after the mutation of a DNA
repair pathway gene by compromising the survival of oocytes and suggests the existence of an oocyte-derived signal that biases gonad fate towards the female developmental pathway and thereby controls zebrafish sex determination.

Transcription factor map alignment of promoter regions.

Transcription factor map alignment of promoter regions. Blanco García, Enrique; Messeguer Peypoch, Xavier; Smith, T.F.; Guigó Serra, Roderic We address the problem of comparing and characterizing the promoter regions of genes with similar expression patterns. This remains a challenging problem in sequence analysis, because often the promoter regions of co-expressed genes do not show discernible sequence conservation. In our approach, thus, we have not directly compared the nucleotide sequence of promoters. Instead, we have obtained predictions of transcription factor binding sites, annotated the predicted sites with the labels of the corresponding binding factors, and aligned the resulting sequences of labels to which we refer here as transcription factor maps (TF-maps). To obtain the global pairwise alignment of two TF-maps, we have adapted an algorithm initially developed to align restriction enzyme maps. We have optimized the parameters of the algorithm in a small, but well-curated, collection of human
mouse orthologous gene pairs. Results in this dataset, as well as in an independent much larger dataset from the CISRED database, indicate that TF-map alignments are able to uncover conserved regulatory elements, which cannot be detected by the typical sequence alignments.

Roles of brca2 (fancd1) in Oocyte Nuclear Architecture, Gametogenesis, Gonad Tumors, and Genome Stability in Zebrafish

Roles of brca2 (fancd1) in Oocyte Nuclear Architecture, Gametogenesis, Gonad Tumors, and Genome Stability in Zebrafish Rodríguez Marí, Adriana; Wilson, C.; Titus, T.A.; Cañestro García, Cristian; BreMiller, R.A.; Yan, Y.L.; Nanda, I.; Johnston, A.; Kanki, J.P.; Grayl, E.M.; He, X.; Spitsbergen, J.; Schindeler, D.; Postlethwait, J.H. Mild mutations in BRCA2 (FANCD1) cause Fanconi anemia (FA) when homozygous, while severe mutations cause common cancers including breast, ovarian, and prostate cancers when heterozygous. Here we report a zebrafish brca2 insertional mutant that shares phenotypes with human patients and identifies a novel brca2 function in oogenesis. Experiments showed that mutant embryos and mutant cells in culture experienced genome instability, as do cells in FA patients. In wild-type zebrafish, meiotic cells expressed brca2; and, unexpectedly, transcripts in oocytes localized asymmetrically to the animal pole. In juvenile brca2 mutants, oocytes failed to progress through meiosis, leading to female-to-male sex reversal. Adult mutants became sterile males due to the meiotic arrest of spermatocytes, which then died by apoptosis, followed by neoplastic proliferation of gonad somatic cells that was similar to neoplasia observed in ageing dead end (dnd)-knockdown males, which lack germ cells. The construction of animals doubly mutant for brca2 and the apoptotic gene tp53 (p53) rescued brca2-dependent sex reversal. Double mutants developed oocytes and became sterile females that produced only aberrant embryos and showed elevated risk for invasive ovarian tumors. Oocytes in double-mutant females showed normal localization of brca2 and pou5f1 transcripts to the animal pole and vasa transcripts to the vegetal pole, but had a polarized rather than symmetrical nucleus with the distribution of nucleoli and chromosomes to opposite nuclear poles; this result revealed a novel role for Brca2 in establishing or maintaining oocyte nuclear architecture. Mutating tp53 did not rescue the infertility phenotype in brca2 mutant males, suggesting that brca2 plays an essential role in zebrafish spermatogenesis. Overall, this work verified zebrafish as a model for the role of Brca2 in human disease and uncovered a novel function of Brca2 in vertebrate oocyte nuclear architecture.

Consequences of lineage-specific gene loss on functional evolution of surviving paralogs: ALDH1A and retinoic acid signaling in vertebrate genomes

Consequences of lineage-specific gene loss on functional evolution of surviving paralogs: ALDH1A and retinoic acid signaling in vertebrate genomes Cañestro García, Cristian; Catchen, J.; Rodríguez Marí, Adriana; Yokoi, H.; Postlethwait, J.H. Genome duplications increase genetic diversity and may facilitate the evolution of gene subfunctions. Little attention, however, has focused on the evolutionary impact of lineage-specific gene loss. Here, we show that identifying lineage-specific gene loss after genome duplication is important for understanding the evolution of gene subfunctions in surviving paralogs and for improving functional connectivity among human and model organism genomes. We examine the general principles of gene loss following duplication, coupled with expression analysis of the retinaldehyde dehydrogenase Aldh1a gene family during retinoic acid signaling in eye development as a case study. Humans have three ALDH1A genes, but teleosts have just one or two. We used comparative genomics and conserved syntenies to identify loss of ohnologs (paralogs derived from genome duplication) and to clarify uncertain phylogenies. Analysis showed that Aldh1a1 and Aldh1a2 form a clade that is sister to Aldh1a3-related genes. Genome comparisons showed secondarily loss of aldh1a1 in teleosts, revealing that Aldh1a1 is not a tetrapod innovation and that aldh1a3 was recently lost in medaka, making it the first known vertebrate with a single aldh1a gene. Interestingly, results revealed asymmetric distribution of surviving ohnologs between co-orthologous teleost chromosome segments, suggesting that local genome architecture can influence ohnolog survival. We propose a model that reconstructs the chromosomal history of the Aldh1a family in the ancestral vertebrate genome, coupled with the evolution of gene functions in surviving Aldh1a ohnologs after R1, R2, and R3 genome duplications. Results provide evidence for early subfunctionalization and late subfunction-partitioning and suggest a mechanistic model based on altered regulation leading to heterochronic gene expression to explain the acquisition or modification of subfunctions by surviving ohnologs that preserve unaltered ancestral developmental programs in the face of gene loss.

Acoel flatworms are not Platyhelminthes: evidence from phylogenomics

Acoel flatworms are not Platyhelminthes: evidence from phylogenomics Philippe, H.; Brinkmann, H.; Martínez Serra, Pedro; Riutort León, Marta; Baguñà Monjo, Jaume Acoel flatworms are small marine worms traditionally considered to belong to the phylum Platyhelminthes. However, molecular phylogenetic analyses suggest that acoels are not members of Platyhelminthes, but are rather extant members of the earliest diverging Bilateria. This result has been called into question, under suspicions of a long branch attraction (LBA) artefact. Here we re-examine this problem through a phylogenomic approach using 68 different protein-coding genes from the acoel Convoluta pulchra and 51 metazoan species belonging to 15 different phyla. We employ a mixture model, named CAT, previously found to overcome LBA artefacts where classical models fail. Our results unequivocally show that acoels are not part of the classically defined Platyhelminthes, making the latter polyphyletic. Moreover, they indicate a deuterostome affinity for acoels, potentially as a sister group to all deuterostomes, to Xenoturbellida, to Ambulacraria, or even to chordates. However, the weak support found for most deuterostome nodes, together with the very fast evolutionary rate of the acoel Convoluta pulchra, call for more data from slowly evolving acoels (or from its sister-group, the Nemertodermatida) to solve this challenging phylogenetic problem.

The genetic content of chromosomal inversions across a wide latitudinal gradient

The genetic content of chromosomal inversions across a wide latitudinal gradient Simões, Pedro; Calabria Garcia, Gemma; Picão Osório, J.; Balanyà i Maymó, Joan; Pascual Berniola, Marta There is increasing evidence regarding the role of chromosomal inversions in relevant biological processes such as local adaptation and speciation. A classic example of the adaptive role of chromosomal polymorphisms is given by the clines of inversion frequencies in Drosophila subobscura, repeatable across continents. Nevertheless, not much is known about the molecular variation associated with these polymorphisms. We characterized the genetic content of ca. 600 individuals from nine European populations following a latitudinal gradient by analysing 19 microsatellite loci from two autosomes (J and U) and the sex chromosome (A), taking into account their chromosomal inversions. Our results clearly demonstrate the molecular genetic uniformity within a given chromosomal inversion across a large latitudinal gradient, particularly from Groningen (Netherlands) in the north to Málaga (Spain) in the south, experiencing highly diverse environmental conditions. This low genetic differentiation within the same gene arrangement across the nine European populations is consistent with the local adaptation hypothesis for th evolutionof chromosomal polymorphisms. We also show the effective role of chromosomal inversions in maintaining different genetic pools within these inverted genomic regions even in the presence of high gene flow. Inversions represent thus an important barrier to gene flux and can help maintain specific allelic combinations with positive effects on fitness. Consistent patterns of microsatellite allele-inversion linkage disequilibrium particularly in loci within inversions were also observed. Finally, we identified areas within inversions presenting clinal variation that might be under selection.

Ubiquitin-specific protease USP25 functions in endoplasmic reticulum-associated degradation

Ubiquitin-specific protease USP25 functions in endoplasmic reticulum-associated degradation Blount, J.R.; Burr, A.A.; Denuc Isern, Amanda; Marfany i Nadal, Gemma; Todi, S.V. Endoplasmic Reticulum (ER)-associated degradation (ERAD) discards abnormal proteins synthesized in the ER. Through coordinated actions of ERAD components, misfolded/anomalous proteins are recognized, ubiquitinated, extracted from the ER and ultimately delivered to the proteasome for degradation. It is not well understood how ubiquitination of ERAD substrates is regulated. Here, we present evidence that the deubiquitinating enzyme Ubiquitin-Specific Protease 25 (USP25) is involved in ERAD. Our data support a model where USP25 counteracts ubiquitination of ERAD substrates by the ubiquitin ligase HRD1, rescuing them from degradation by the proteasome.

Tracking Invasion Histories in the Sea: Facing Complex Scenarios Using Multilocus Data

Tracking Invasion Histories in the Sea: Facing Complex Scenarios Using Multilocus Data Rius Viladomiu, Marc; Turon Barrera, Xavier; Ordóñez, V.; Pascual Berniola, Marta In recent years, new analytical tools have allowed researchers to extract historical information contained in molecular data, which has fundamentally transformed our understanding of processes ruling biological invasions. However, the use of these new analytical tools has been largely restricted to studies of terrestrial organisms despite the growing recognition that the sea contains ecosystems that are amongst the most heavily affected by biological invasions, and that marine invasion histories are often remarkably complex. Here, we studied the routes of invasion and colonisation histories of an invasive marine invertebrate Microcosmus squamiger (Ascidiacea) using microsatellite loci, mitochondrial DNA sequence data and 11 worldwide populations. Discriminant analysis of principal components, clustering methods and approximate Bayesian computation (ABC) methods showed that the most likely source of the introduced populations was a single admixture event that involved populations from two genetically differentiated ancestral regions - the western and eastern coasts of Australia. The ABC analyses revealed that colonisation of the introduced range of M. squamiger consisted of a series of non-independent introductions along the coastlines of Africa, North America and Europe. Furthermore, we inferred that the sequence of colonisation across continents was in line with historical taxonomic records - first the Mediterranean Sea and South Africa from an unsampled ancestral population, followed by sequential introductions in California and, more recently, the NE Atlantic Ocean. We revealed the most likely invasion history for world populations of M. squamiger, which is broadly characterized by the presence of multiple ancestral sources and non-independent introductions within the introduced range. The results presented here illustrate the complexity of marine invasion routes and identify a cause-effect relationship between human-mediated transport and the success of widespread marine non-indigenous species, which benefit from stepping-stone invasions and admixture processes involving different sources for the spread and expansion of their range.

Large-scale evidence for the effect of the COL1A1 Sp1 polymorphism on osteoporosis outcomes: the GENOMOS study

Large-scale evidence for the effect of the COL1A1 Sp1 polymorphism on osteoporosis outcomes: the GENOMOS study Ralston, S.H.; Uitterlinden, A.G.; Brandi, M.L.; Balcells Comas, Susana; Langdahl, B. L.; Lips, P.; Lorenc, R.; Obermayer-Pietsch, B.; Scollen, S.; Bustamante, M.; Bjerre Husted, L.; Carey, A. H.; Diez Perez, A.; Dunning, A.M; Falchetti, A.; Karczmarewicz, E.; Kruk, M. Osteoporosis and fracture risk are considered to be under genetic control. Extensive work is being performed to identify the exact genetic variants that determine this risk. Previous work has suggested that a G/T polymorphism affecting an Sp1 binding site in the COLIA1 gene is a genetic marker for low bone mineral density (BMD) and osteoporotic fracture, but there have been no very-large-scale studies of COLIA1 alleles in relation to these phenotypes.

In memoriam Antonio Prevosti (1919-2011), a pioneer of genetics of natural populations

In memoriam Antonio Prevosti (1919-2011), a pioneer of genetics of natural populations Mestres i Naval, Francesc; Serra i Camó, Lluís; Sperlich, D. Prof. Dr. Antonio Prevosti was born on the 15th of February, 1919, at Barcelona where he grew up and lived with his wife Maria Monclús and his family until his death. Between the years 1939 and 1942 he registered at the University of Barcelona and finished his studies there with the"University Degree in Natural Science" obtaining the extraordinary award of his promotion. Immediately afterwards he started his research about the growth rate of Barcelona school children from two very different social groups at the Anthropology Laboratory of the University of Barcelona. The obtained results were published in his Doctoral Thesis (1948). Further investigations on quantitative traits in human populations improved his knowledge of statistical analyses and provided the basis for a scholarship from the Italian Ministry of Foreign Affairs to work at the Institute of Statistical and Demographical Sciences in Rome directed by Prof. C. Gini.

The planarian flatworm: an in vivo model for stem cell biology and nervous system regeneration

The planarian flatworm: an in vivo model for stem cell biology and nervous system regeneration Gentile, L.; Cebrià Sánchez, Francesc; Bartscherer, K. Planarian flatworms are an exception among bilaterians in that they possess a large pool of adult stem cells that enables them to promptly regenerate any part of their body, including the brain. Although known for two centuries for their remarkable regenerative capabilities, planarians have only recently emerged as an attractive model for studying regeneration and stem cell biology. This revival is due in part to the availability of a sequenced genome and the development of new technologies, such as RNA interference and next-generation sequencing, which facilitate studies of planarian regeneration at the molecular level. Here, we highlight why planarians are an exciting tool in the study of regeneration and its underlying stem cell biology in vivo, and discuss the potential promises and current limitations of this model organism for stem cell research and regenerative medicine.

The Saccharomyces cerevisiae Hot1p regulated gene YHR087W (HGI1) has a role in translation upon high glucose concentration stress.

The Saccharomyces cerevisiae Hot1p regulated gene YHR087W (HGI1) has a role in translation upon high glucose concentration stress. Gomar-Alba, M.; Jiménez Martí, Elena; Del Olmo, M. Background While growing in natural environments yeasts can be affected by osmotic stress provoked by high glucose concentrations. The response to this adverse condition requires the HOG pathway and involves transcriptional and posttranscriptional mechanisms initiated by the phosphorylation of this protein, its translocation to the nucleus and activation of transcription factors. One of the genes induced to respond to this injury is YHR087W. It encodes for a protein structurally similar to the N-terminal region of human SBDS whose expression is also induced under other forms of stress and whose deletion determines growth defects at high glucose concentrations. Results In this work we show that YHR087W expression is regulated by several transcription factors depending on the particular stress condition, and Hot1p is particularly relevant for the induction at high glucose concentrations. In this situation, Hot1p, together to Sko1p, binds to YHR087W promoter in a Hog1p-dependent manner. Several evidences obtained indicate Yhr087wp"s role in translation. Firstly, and according to TAP purification experiments, it interacts with proteins involved in translation initiation. Besides, its deletion mutant shows growth defects in the presence of translation inhibitors and displays a slightly slower translation recovery after applying high glucose stress than the wild type strain. Analyses of the association of mRNAs to polysome fractions reveals a lower translation in the mutant strain of the mRNAs corresponding to genes GPD1, HSP78 and HSP104. Conclusions The data demonstrates that expression of Yhr087wp under high glucose concentration is controlled by Hot1p and Sko1p transcription factors, which bind to its promoter. Yhr087wp has a role in translation, maybe in the control of the synthesis of several stress response proteins, which could explain the lower levels of some of these proteins found in previous proteomic analyses and the growth defects of the deletion strain. Keywords: Saccharomyces cerevisiae; High glucose osmotic stress; Gene YHR087W; Gene expression; Translation; Hot1p; Hog1p; Polysomes

Genetic markers of the Va/Ba balanced lethal strain of Drosophila subobscura

Genetic markers of the Va/Ba balanced lethal strain of Drosophila subobscura Araúz, P.A.; Pegueroles Queralt, Cinta; Calabria Garcia, Gemma; Serra i Camó, Lluís; Balanyà i Maymó, Joan; Pascual Berniola, Marta; Mestres i Naval, Francesc The Va/Ba strain, constructed by Sperlich et al. (1977), is the only balanced lethal strain in D. subobscura. It allows the production of homozygous O chromosomes and has been a useful tool not only to analyse chromosomal viabilities but also to obtain homokaryotypic lines (Mestres and Serra, 2008). Besides the morphological dominant mutations Va (Varicose) and Ba (Bare), other genetic markers have been characterized in this strain, some of them by our group and not described previously. Here we present a list of these markers.

White eye phenotypes and their genetic analysis

White eye phenotypes and their genetic analysis Mestres i Naval, Francesc; Arenas Solà, Concepción An interesting case for undergraduate students of general Genetics is to consider that different genes can produce the same or similar phenotypes. We present here an experiment to discover that the same phenotype could be produced by different genes, and then, to carry out the genetic analysis of these genes. For this laboratory study we have used the following Drosophila melanogaster strains: white (white eyes) and scarlet
brown (white eyes).

Genome-wide chromatin occupancy reveals a role for ASH2 in transcriptional pausing

Genome-wide chromatin occupancy reveals a role for ASH2 in transcriptional pausing Perez-Lluch, Silvia; Blanco García, Enrique; Carbonell Sanroma, Albert; Raha, D.; Snyder, M.; Serras Rigalt, Florenci; Corominas, Montserrat (Corominas Guiu) While it is widely acknowledged that the ubiquitin
proteasome system plays an important role in transcription, little is known concerning the mechanistic basis, in particular the spatial organization of proteasome-dependent proteolysis at the transcription site. Here, we show that proteasomal activity and tetraubiquitinated proteins concentrate to nucleoplasmic microenvironments in the euchromatin. Such proteolytic domains are immobile and distinctly positioned in relation to transcriptional processes. Analysis of gene arrays and early genes in Caenorhabditis elegans embryos reveals that proteasomes and proteasomal activity are distantly located relative to transcriptionally active genes. In contrast, transcriptional inhibition generally induces local overlap of proteolytic microdomains with components of the transcription machinery and degradation of RNA polymerase II. The results establish that spatial organization of proteasomal activity differs with respect to distinct phases of the transcription cycle in at least some genes, and thus might contribute to the plasticity of gene expression in response to environmental stimuli.

Gene flow and gene flux shape evolutionary patterns of variation in Drosophila subobscura

Gene flow and gene flux shape evolutionary patterns of variation in Drosophila subobscura Pegueroles Queralt, Cinta; Aquadro, C.F.; Mestres i Naval, Francesc; Pascual Berniola, Marta Gene flow (defined as allele exchange between populations) and gene flux (defined as allele exchange during meiosis in heterokaryotypic females) are important factors decreasing genetic differentiation between populations and inversions. Many chromosomal inversions are under strong selection and their role in recombination reduction enhances the maintenance of their genetic distinctness. Here we analyze levels and patterns of nucleotide diversity, selection and demographic history, using 37 individuals of Drosophila subobscura from Mount Parnes (Greece) and Barcelona (Spain). Our sampling focused on two frequent O-chromosome arrangements that differ by two overlapping inversions (OST and O3+4), which are differentially adapted to the environment as observed by their opposing latitudinal clines in inversion frequencies. The six analyzed genes (Pif1A, Abi, Sqd, Yrt, Atpa and Fmr1) were selected for their location across the O-chromosome and their implication in thermal adaptation. Despite the extensive gene flux detected outside the inverted region, significant genetic differentiation between both arrangements was found inside it. However, high levels of gene flow were detected for all six genes when comparing the same arrangement among populations. These results suggest that the adaptive value of inversions is maintained, regardless of the lack of genetic differentiation within arrangements from different populations, and thus favors the Local Adaptation hypothesis over the Coadapted Genome hypothesis as the basis of the selection acting on inversions in these populations.

Ash2 acts as an Ecdysone Receptor coactivator by stabilizing the histone methyltransferase Trr.

Ash2 acts as an Ecdysone Receptor coactivator by stabilizing the histone methyltransferase Trr. Carbonell Sanroma, Albert; Mazo, A.; Serras Rigalt, Florenci; Corominas, Montserrat (Corominas Guiu) The molting hormone ecdysone triggers chromatin changes via histone modifica- tions that are important for gene regulation. On hormone activation, the ecdysone receptor (EcR) binds to the SET domain
containing histone H3 methyltransferase trithorax-related protein (Trr). Methylation of histone H3 at lysine 4 (H3K4me), which is associated with tran- scriptional activation, requires several cofactors, including Ash2. We find that ash2 mutants have severe defects in pupariation and metamorphosis due to a lack of activation of ecdy- sone-responsive genes. This transcriptional defect is caused by the absence of the H3K4me3 marks set by Trr in these genes. We present evidence that Ash2 interacts with Trr and is re- quired for its stabilization. Thus we propose that Ash2 functions together with Trr as an ecdysone receptor coactivator.

CBS: an open platform that integrates predictive methods and epigenetics information to characterize conserved regulatory features in multiple Drosophila genomes.

CBS: an open platform that integrates predictive methods and epigenetics information to characterize conserved regulatory features in multiple Drosophila genomes. Blanco García, Enrique; Corominas, Montserrat (Corominas Guiu) Background: Information about the composition of regulatory regions is of great value for designing experiments to functionally characterize gene expression. The multiplicity of available applications to predict transcription factor binding sites in a particular locus contrasts with the substantial computational expertise that is demanded to manipulate them, which may constitute a potential barrier for the experimental community. Results: CBS (Conserved regulatory Binding Sites, http://compfly.bio.ub.es/CBS) is a public platform of evolutionarily conserved binding sites and enhancers predicted in multiple Drosophila genomes that is furnished with published chromatin signatures associated to transcriptionally active regions and other experimental sources of information. The rapid access to this novel body of knowledge through a user-friendly web interface enables non-expert users to identify the binding sequences available for any particular gene, transcription factor, or genome region. Conclusions: The CBS platform is a powerful resource that provides tools for data mining individual sequences and groups of co-expressed genes with epigenomics information to conduct regulatory screenings in Drosophila.

dKDM5/LID regulates H3K4me3 dynamics at the transcription-start site (TSS) of actively transcribed developmental genes

dKDM5/LID regulates H3K4me3 dynamics at the transcription-start site (TSS) of actively transcribed developmental genes Lloret-Llinares, Marta; Perez-Lluch, Silvia; Rossell, David; Morán, Tomás; Ponsà-Cobas, Joan; Auer, Herbert; Corominas, Montserrat (Corominas Guiu); Azorin, Fernando H3K4me3 is a histone modification that accumulates at the transcription-start site (TSS) of active genes and is known to be important for transcription activation. The way in which H3K4me3 is regulated at TSS and the actual molecular basis of its contribution to transcription remain largely unanswered. To address these questions, we have analyzed the contribution of dKDM5/LID, the main H3K4me3 demethylase in Drosophila, to the regulation of the pattern of H3K4me3. ChIP-seq results show that, at developmental genes, dKDM5/LID localizes at TSS and regulates H3K4me3. dKDM5/LID target genes are highly transcribed and enriched in active RNApol II and H3K36me3, suggesting a positive contribution to transcription. Expression-profiling show that, though weakly, dKDM5/LID target genes are significantly downregulated upon dKDM5/LID depletion. Furthermore, dKDM5/LID depletion results in decreased RNApol II occupancy, particularly by the promoter-proximal Pol lloser5 form. Our results also show that ASH2, an evolutionarily conserved factor that locates at TSS and is required for H3K4me3, binds and positively regulates dKDM5/LID target genes. However, dKDM5/LID and ASH2 do not bind simultaneously and recognize different chromatin states, enriched in H3K4me3 and not, respectively. These results indicate that, at developmental genes, dKDM5/LID and ASH2 coordinately regulate H3K4me3 at TSS and that this dynamic regulation contributes to transcription.

Competition between SOCS36E and Drk modulates Sevenless receptor tyrosine kinase activity

Competition between SOCS36E and Drk modulates Sevenless receptor tyrosine kinase activity Almudi, I.; Corominas, Montserrat (Corominas Guiu); Serras Rigalt, Florenci Modulation of signalling pathways can trigger different cellular responses, including differences in cell fate. This modulation can be achieved by controlling the pathway activity with great precision to ensure robustness and reproducibility of the specification of cell fate. The development of the photoreceptor R7 in the Drosophila melanogasterretina has become a model in which to investigate the control of cell signalling. During R7 specification, a burst of Ras small GTPase (Ras) and mitogen-activated protein kinase (MAPK) controlled by Sevenless receptor tyrosine kinase (Sev) is required. Several cells in each ommatidium express sev. However, the spatiotemporal expression of the boss ligand and the action of negative regulators of the Sev pathway will restrict the R7 fate to a single cell. The Drosophila suppressor of cytokine signalling 36E (SOCS36E) protein contains an SH2 domain and acts as a Sev signalling attenuator. By contrast, downstream of receptor kinase (Drk), the fly homolog of the mammalian Grb2 adaptor protein, which also contains an SH2 domain, acts as a positive activator of the pathway. Here, we apply the Förster resonance energy transfer (FRET) assay to transfected Drosophila S2 cells and demonstrate that Sev binds directly to either the suppressor protein SOCS36E or the adaptor protein Drk. We propose a mechanistic model in which the competition between these two proteins for binding to the same docking site results in either attenuation of the Sev transduction in cells that should not develop R7 photoreceptors or amplification of the Ras
MAPK signal only in the R7 precursor.

The CdiTESK1 kinase is required for sevenless signaling and photoreceptor cell shape in the Drosophila eye

The CdiTESK1 kinase is required for sevenless signaling and photoreceptor cell shape in the Drosophila eye Sesé, M.; Corominas, Montserrat (Corominas Guiu); Stocker, H.; Heino, T.I.; Hafen, E.; Serras Rigalt, Florenci How cellular behaviors such as cell-to-cell communication, epithelial organization and cell shape reorganization are coordinated during development is poorly understood. The developing Drosophila eye offers an ideal model system to study these processes. Localized actin polymerization is required to constrict the apical surface of epithelial cells of the eye imaginal disc to maintain the reﬁned arrangement of the developing ommatidia. The identity of each photoreceptor cell within the epithelium is determined by cell-to-cell contacts involving signal transduction events. The R7 photoreceptor cell requires the activity of the Sevenless RTK to adopt a proper cell fate. We performed an EP screen for negative regulators of this inductive process, and we identiﬁed the serine/threonine kinase Center divider (cdi) as a suppressor of the phenotype caused by an activated Sevenless receptor. Cdi is homologous to the human testis-speciﬁc kinase 1 (TESK1), a member of the LIM kinases involved in cytoskeleton control through ADF/coﬁlin phosphorylation. We have analyzed the effects of gain- and loss-of-function of cdi and found alterations in actin organization and in the adherens junctions proteins DE-cadherin and -catenin, as well as in Sevenless apical localization. Interference with the function of the ADF/coﬁlin phosphatase Slingshot (ssh), which antagonizes Cdi, also results in a suppression of signaling triggered by the Sevenless RTK. Our results reveal a critical interplay between the localization of molecules involved in epithelial organization and signal transduction. Key words: cdi, Sevenless, Photoreceptor, Eye, Actin