Neurogenomics: An opportunity to integrate neuroscience, genomics and bioinformatics research in Africa

AbstractModern genomic approaches have made enormous contributions to improving our understanding of the function, development and evolution of the nervous system, and the diversity within and between species. However, most of these research advances have been recorded in countries with advanced scientific resources and funding support systems. On the contrary, little is known about, for example, the possible interplay between different genes, non-coding elements and environmental factors in modulating neurological diseases among populations in low-income countries, including many African countries. The unique ancestry of African populations suggests that improved inclusion of these populations in neuroscience-related genomic studies would significantly help to identify novel factors that might shape the future of neuroscience research and neurological healthcare. This perspective is strongly supported by the recent identification that diseased individuals and their kindred from specific sub-Saharan African populations lack common neurological disease-associated genetic mutations. This indicates that there may be population-specific causes of neurological diseases, necessitating further investigations into the contribution of additional, presently-unknown genomic factors. Here, we discuss how the development of neurogenomics research in Africa would help to elucidate disease-related genomic variants, and also provide a good basis to develop more effective therapies. Furthermore, neurogenomics would harness African scientists' expertise in neuroscience, genomics and bioinformatics to extend our understanding of the neural basis of behaviour, development and evolution

The role of Dichaete in transcriptional regulation during Drosophila embryonic development

Sox domain genes encode a family of developmentally important transcription factors conserved throughout the Metazoa. The subgroup B, which includes the mammalian Sox1, 2 and 3 proteins and their Drosophila counterparts Dichaete and SoxNeuro, are particularly important for the development of the nervous system where they appear to play conserved roles in neuronal specification and differentiation. Despite years of detailed study we still have a relatively poor idea of how Sox proteins function on a genome wide scale and the aim of my PhD work was to explore this aspect using the fly group B protein, Dichaete. A number of studies have shown that Dichaete performs a variety of critical functions during development and a few individual regulatory targets have been defined, however, at the start of my work no genome-wide data on Dichaete action were available. While such data emerged from large scale initiatives during my work, a systematic analysis of Dichaete action was lacking. Here I describe the first detailed genomic analysis of Dichaete activity, with a particular focus on three areas: finding the locations of Dichaete binding in the genome, a prediction of potential Dichaete cofactors and an analysis of Dichaete effects on gene expression. To address the issue of where Dichaete binds in the genome, I generated whole genome DamID data for embryos and followed this with a detailed comparative analysis, combining my data with three newly published ChIP-chip datasets. The combined studies identify thousands of binding regions, mostly in the vicinity of developmentally important genes. The binding profiles were found to be consistent with Dichaete acting on enhancer regions and also suggest a role in facilitating RNA Polymerase II pausing. The analysis also identified a Dichaete binding motif closely matching that found with in vitro studies. By combined ChIP and DamID datasets I generated a very high confidence core Dichaete binding dataset, which should be of considerable use in future studies. To identify potential Dichaete cofactors, I compiled the available embryonic transcription factor binding data from the Berkeley Drosophila Transcription Network and mod- ENCODE projects, and identified significant overlaps with the core Dichaete binding data. A number of the proteins highlighted in this analysis have known roles during neuroblast development, including Hunchback and Krüppel, transcription factors involved in temporal specification of neuroblast division, and Prospero, which plays a key role in neuroblast differentiation. The analysis suggests that Dichaete has a role during early neuroblast divisions, where it likely interacts with Hb and Kr to maintain neuroblast pluripotency. This is a role consistent with previous studies in Drosophila larval neuroblasts and is analogous to neural functions of Sox2 in mammals. My analysis suggests that Dichaete acts on the same target genes as Prospero but in an antagonistic role, with Dichaete preventing stem cell differentiation and Prospero promoting it. To examine the effects of Dichaete on gene expression, a number of microarray transcript profiling studies were performed, including a global study with Dichaete null mutants, and tissue specific studies in the CNS midline and neuroblasts via the use of dominant negative constructs. Whole transcriptome expression profiling data was combined with the binding data to establish a set of high confidence potential Dichaete targets, both for specific tissues and more globally during neurogenesis. Specific high confidence targets were found, including bancal during nervous system development. It was also concluded that Dichaete is likely to prevent cell cycle exit by repressing the apoptosis genes grim, hid and reaper, as well as the differentiation genes prospero and miranda. An extensive list of potential Dichaete direct targets was generated and can be used for validation and future research.This work was supported by a Medical Research Council scholarship

Identifying targets of the Sox domain protein Dichaete in the Drosophila CNS via targeted expression of dominant negative proteins.

BACKGROUND: Group B Sox domain transcription factors play important roles in metazoan central nervous system development. They are, however, difficult to study as mutations often have pleiotropic effects and other Sox family members can mask phenotypes due to functional compensation. In Drosophila melanogaster, the Sox gene Dichaete is dynamically expressed in the embryonic CNS, where it is known to have functional roles in neuroblasts and the ventral midline. In this study, we use inducible dominant negative proteins in combination with ChIP, immunohistochemistry and genome-wide expression profiling to further dissect the role of Dichaete in these two tissues. RESULTS: We generated two dominant negative Dichaete constructs, one lacking a DNA binding domain and the other fused to the Engrailed transcriptional repressor domain. We expressed these tissue-specifically in the midline and in neuroblasts using the UAS/GAL4 system, validating their use at the phenotypic level and with known target genes. Using ChIP and immunohistochemistry, we identified two new likely direct Dichaete target genes, commisureless in the midline and asense in the neuroectoderm. We performed genome-wide expression profiling in stage 8-9 embryos, identifying almost a thousand potential tissue-specific Dichaete targets, with half of these genes showing evidence of Dichaete binding in vivo. These include a number of genes with known roles in CNS development, including several components of the Notch, Wnt and EGFR signalling pathways. CONCLUSIONS: As well as identifying commisureless as a target, our data indicate that Dichaete helps establish its expression during early midline development but has less effect on its established later expression, highlighting Dichaete action on tissue specific enhancers. An analysis of the broader range of candidate Dichaete targets indicates that Dichaete plays diverse roles in CNS development, with the 500 or so Dichaete-bound putative targets including a number of transcription factors, signalling pathway components and terminal differentiation genes. In the early neurectoderm we implicate Dichaete in the lateral inhibition pathway and show that Dichaete acts to repress the proneural gene asense. Our analysis also reveals that dominant negatives cause off-target effects, highlighting the need to use other experimental data for validating findings from dominant negative studies.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

Randman trogodišnjeg šarana iz kaveznog sistema uzgoja

Uzgoj šarana (Cyprinus carpio L.) u kaveznom sistemu predstavlja posebnu vrstu intenzivne proizvodnje. Odlikuje se malim početnim ulaganjima u izgradnju kaveznog ribnjaka, ne zahteva značajno angažovanje radne snage i obezbeđuje veliku proizvodnju po jedinici zapremine. Izbor lokacije je od velikog značaja jer utiče na ekonomsku održivost, proizvodnju i mortalitet šarana. U odnosu na druge uzgojne sisteme, utvrđeno je da u kaveznom sistemu morbiditet i mortalitet mogu značajno varirati i u slučaju odstupanja vrednosti parametara kvaliteta vode od optimalnih, gubici mogu nastupiti brzo i biti dramatični. Nedostatak kaveznog sistema gajenja ogleda se u olakšanoj transmisiji bolesti i zagađenju vodenog ekosistema u kome je kavezni sistem postavljen, što se može sprečiti uvođenjem zaštitnih sistema. Uz poštovanje načela dobre proizvođačke prakse, održavanjem dobrog kvaliteta vode i optimalno izbalansiranu hranu primerenu starosnoj kategoriji uzgajanog šarana, očekivan mortalitet je 1-5%. Zahtevi savremenog tržišta su sve više usmereni ka obrađenoj ribi, posebno filetima. Uklanjanjem kože i odstranjivanjem unutrašnjih organa i intramuskularnih kostiju, fileti šarana i ostalih ciprinida koje se gaje u polikulturi sa šaranom, postaju visokovredni obroci, koji se lako i brzo spremaju. Randman riba je ekonomski parametar čija je vrednost značajna pri svim tehnološkim operacijama vezanim za preradu ribe. Iskazuje se kao odnos primarno obrađenog trupa u odnosu na masu trupa žive ribe. Odnos jestivog i nejestivog dela trupa ribe može značajno da varira u zavisnosti od vrste, mase i veličine ribe i sezone ulova). Od njega direktno zavisi ekonomičnost proizvodnje i neopravdano je zanemaren kao parameter u istraživanjima. Razlika u randmanu između različitih linija i njihovih hibrida javlja se zbog različitog oblika, dužine glave, debljine i širine tela. Randman riba značajno je povoljniji od randmana kod ostalih vrsta životinja. Delovi koji se odbacuju mogu sadržati sve ili samo neke delove glave, krljušti, kože, creva, gonada i peraja. Cilj istraživanja bio je određivanje randmana kod trogodišnjeg šarana poreklom iz kaveznog sistema uzgoja. Uzorkovanje je obavljeno iz dva kavezna sistema na Tikveškom jezeru (Makedonija) (41°20′51″N 21°57′58″E) i iz jednog na Bilećkom jezeru (Bosna i Hercegovina) (42°49′31″N 18°26′17″E). Ishrana riba vršena je kompletnim krmnim smešama različitih proizvođača. Iz svakog kaveznog sistema uzorkovano je po 8 jedinki muškog i 8 jedinki ženskog pola (ukupno po 24 jedinke svakog pola). Morfometrijske osobine riba utvrđene su sa ciljem da se ispita uticaj pola na randman trogodišnjeg šarana gajenog u kaveznom sistemu. Nakon što su izlovljeni, šarani su držani na ledu. Krljušt je skinuta ručno nazubljenim nožem. Glava je odsečena cirkularnim rezom ispred pojasa pektoralnog peraja tako da je peraje ostalo na trupu. Peraja su odsečena na početku perajnih žbica. Egzenteracija organa zajedno sa gonadama obavljena je ručno. Dobijeni su obrađen trup, koji podrazumeva trup ribe bez krljušti, peraja, unutrašnjih organa i glave. Posle odstranjivanja krljušti, unutrašnji oragni, gonade, glava, peraja i obrađen trup su izmereni. Rezultati su obrađeni t-testom kako bi se utvrdio uticaj pola na randman trogodišnjeg šarana gajenog u kaveznom sistemu proizvodnje. U odnosu na pol (ženke naspram mužjaka šarana) pažnja je usmerena na parametre koji su važni sa aspekta gajenja i što povoljnijeg randmana. U našem istraživanju randman je određen kao odnos mase cele ribe i mase trupa bez glave, krljušti, peraja i unutrašnjih organa. Rezultati dobijeni u našem istraživanju pokazuju veću težinu ženskih u odnosu na muške jedinke (FW), ali nemaju statističku značajnost. Ukupna dužina (TL), standardna dužina (SL) i dužina trupa (CL) ženki u odnosu na mužjake imaju statistički značajnu razliku (p < 0,01), dok razlike u telesnoj masi ženki i mužjaka nisu statistički značajne. Rezultati dobijeni merenjem težine trupa (CW) i težine unutrašnjih organa (VW) ženki u odnosu na iste parametre kod mužjaka imaju statistički značajnu razliku (p < 0,01). Vrednosti dobijene merenjem težine glave (HW), težine gonada (GW) mužjaka, kao i vrednosti gonadosomatskog indeksa (GSI) u odnosu na vrednosti istih parametara kod ženki imaju statistički značaj (p < 0,01). Razlike u dužini glave (HL), visini tela (BH), masi peraja (FW), masi jedinki (FW) i obrađenih fileta (FilletW) mužjaka i ženki, nemaju statistički značaj. Vrednosti dobijene izračunavanjem relativne težine fileta i randmana uzorkovanih trogodišnjih jedinki šarana muškog i ženskog pola nemaju statistički značajnu razliku

Biases in Drosophila melanogaster protein trap screens.

BACKGROUND: The ability to localise or follow endogenous proteins in real time in vivo is of tremendous utility for cell biology or systems biology studies. Protein trap screens utilise the random genomic insertion of a transposon-borne artificial reporter exon (e.g. encoding the green fluorescent protein, GFP) into an intron of an endogenous gene to generate a fluorescent fusion protein. Despite recent efforts aimed at achieving comprehensive coverage of the genes encoded in the Drosophila genome, the repertoire of genes that yield protein traps is still small. RESULTS: We analysed the collection of available protein trap lines in Drosophila melanogaster and identified potential biases that are likely to restrict genome coverage in protein trap screens. The protein trap screens investigated here primarily used P-element vectors and thus exhibit some of the same positional biases associated with this transposon that are evident from the comprehensive Drosophila Gene Disruption Project. We further found that protein trap target genes usually exhibit broad and persistent expression during embryonic development, which is likely to facilitate better detection. In addition, we investigated the likely influence of the GFP exon on host protein structure and found that protein trap insertions have a significant bias for exon-exon boundaries that encode disordered protein regions. 38.8% of GFP insertions land in disordered protein regions compared with only 23.4% in the case of non-trapping P-element insertions landing in coding sequence introns (p < 10(-4)). Interestingly, even in cases where protein domains are predicted, protein trap insertions frequently occur in regions encoding surface exposed areas that are likely to be functionally neutral. Considering the various biases observed, we predict that less than one third of intron-containing genes are likely to be amenable to trapping by the existing methods. CONCLUSION: Our analyses suggest that the utility of P-element vectors for protein trap screens has largely been exhausted, and that approximately 2,800 genes may still be amenable using piggyBac vectors. Thus protein trap strategies based on current approaches are unlikely to offer true genome-wide coverage. We suggest that either transposons with reduced insertion bias or recombineering-based targeting techniques will be required for comprehensive genome coverage in Drosophila.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

The role of Dichaete in transcriptional regulation during Drosophila embryonic development.

BACKGROUND: Group B Sox domain transcription factors play conserved roles in the specification and development of the nervous system in higher metazoans. However, we know comparatively little about how these transcription factors regulate gene expression, and the analysis of Sox gene function in vertebrates is confounded by functional compensation between three closely related family members. In Drosophila, only two group B Sox genes, Dichaete and SoxN, have been shown to function during embryonic CNS development, providing a simpler system for understanding the functions of this important class of regulators. RESULTS: Using a combination of transcriptional profiling and genome-wide binding analysis we conservatively identify over 1000 high confidence direct Dichaete target genes in the Drosophila genome. We show that Dichaete plays key roles in CNS development, regulating aspects of the temporal transcription factor sequence that confer neuroblast identity. Dichaete also shows a complex interaction with Prospero in the pathway controlling the switch from stem cell self-renewal to neural differentiation. Dichaete potentially regulates many more genes in the Drosophila genome and was found to be associated with over 2000 mapped regulatory elements. CONCLUSIONS: Our analysis suggests that Dichaete acts as a transcriptional hub, controlling multiple regulatory pathways during CNS development. These include a set of core CNS expressed genes that are also bound by the related Sox2 gene during mammalian CNS development. Furthermore, we identify Dichaete as one of the transcription factors involved in the neural stem cell transcriptional network, with evidence supporting the view that Dichaete is involved in controlling the temporal series of divisions regulating neuroblast identity.This work was supported by a Medical Research Council scholarship

Honey bee viruses in Serbian colonies of different strength

Protection of honey bees is of great economic importance because of their role in pollination. Crucial steps towards this goal are epidemiological surveys of pathogens connected with honey bee losses. In this study deformed wing virus (DWV), chronic bee paralysis virus (CBPV), acute bee paralysis virus (ABPV) and sacbrood virus (SBV) were investigated in colonies of different strength located in five regions of Serbia. The relationship between colony strength and virus occurrence/infection intensity were assessed as well as the genetic relationship between virus sequences from Serbia and worldwide. Real-time RT-PCR analyses detected at least one virus in 87.33% of colonies. Single infection was found in 28.67% colonies (21.33%, 4.00%, 2.67% and 0.67% in cases of DWV, ABPV, SBV and CBPV, respectively). In the majority of colonies (58.66%) more than one virus was found. The most prevalent was DWV (74%), followed by ABPV, SBV and CBPV (49.30%, 24.00% and 6.70%, respectively). Except for DWV, the prevalence of the remaining three viruses significantly varied between the regions. No significant differences were found between colony strength and either (i) the prevalence of DWV, ABPV, SBV, CBPV and their combinations, or (ii) DWV infection levels. The sequences of honey bee viruses obtained from bees in Serbia were 93–99% identical with those deposited in GenBank