1,721,021 research outputs found
Blue luminescent amino-functionalized graphene quantum dots as a responsive material for potential detection of metal ions and malathion
Full text linkLarge amounts of hazardous and toxic substances in the environment require non-toxic, cheap, easy, rapid, and sensitive methods for their detection. Blue luminescent graphene quantum dots (GQDs) were produced by electrochemical cleavage of graphite electrodes followed by gamma irradiation in the presence of ethylenediamine (EDA). Modified dots were able to detect metal ions (Co2+, Pd2+, Fe3+) due to photoluminescence quenching. The highest sensitivity was detected for the sample irradiated at a dose of 25 kGy. The limits of detection (LODs) were 1.79, 2.55, and 0.66 μmol L−1 for Co2+, Fe3+, and Pd2+, respectively. It was observed that GQDs irradiated at 200 kGy act as an ultra-sensitive turn-on probe for Malathion detection with LOD of 94 nmol L−1. Atomic force microscopy images proved the aggregation of GQDs in the presence of the investigated metal ions. Results obtained by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and LIVE/DEAD cytotoxicity test indicated that GQDs irradiated with EDA are not toxic towards MRC-5 cells, which makes them a promising, eco-friendly and safe material for sensing application
The role of TALE transcription factors in the retinoic acid induced activation of SOX3 gene
Full text linkSox3/SOX3 gen je jedan od najranijih markera neuralnog razvića kičmenjaka, koji ima ključnu ulogu u održavanju populacije ćelija neuralnih progenitora. Naša ranija istraživanja dovela su do prve karakterizacije promotora humanog SOX3 gena i identifikacije kontrolnih elemenata preko kojih su opšti transkripcioni faktori NF-Y, Sp1 i USF uključeni u regulaciju ekspresije ovog gena. Rezultati predstavljeni u ovom radu po prvi put pokazuju da su TALE transkripcioni faktori PBX1, MEIS1 i TGIF uključeni u regulaciju ekspresije humanog SOX3 gena.
PBX1 i MEIS1 proteini direktno interaguju sa konsenzusnim mestom za vezivanje Pbx/Meis heterodimera koje je visoko evolutivno očuvano u bazalnom promotoru Sox3 gena sisara. PBX1 je prisutan u proteinskim kompleksima formiranim na ovom mestu sa jedarnim proteinima iz neindukovanih ćelija, dok su i PBX1 i MEIS1 prisutni u kompleksima formiranim sa jedarnim proteinima iz NT2/D1 ćelija indukovanih retinoičnom kiselinom. Zanimljivo je da je MEIS1 jedini do sada identifikovani faktor koji interaguje sa kontrolnim elementima SOX3 promotora tek nakon indukcije ovog gena retinoičnom kiselinom. Funkcionalne analize su pokazale da mutacije Pbx/Meis vezivnog mesta dovode do pada aktivnosti SOX3 promotora u RA indukovanim NT2/D1 ćelijama, dok povećana ekspresija PBX1 i MEIS1 proteina dovodi do aktivacije SOX3 gena kako u indukovananim tako i u neindukovanim ćelijama.
Transkripcioni faktor TGIF ostvaruje funkciju transkripcionog represora interakcijom sa konsenzusnim vezivnim mestom u promotoru SOX3 gena. Mutacija ovog mesta dovodi do povećane aktivnosti SOX3 promotora, dok povećana ekspresija transkripcionog faktora TGIF značajno smanjuje ekspresiju SOX3 gena u neindukovanim i RA indukovanim NT2/D1 ćelijama.
Prikazani rezultati po prvi put uspostavljaju funkcionalnu vezu izmedu članova TALE familije transkripcionih faktora (PBX1, MEIS1 i TGIF) i ekspresije humanog SOX3 gena. Povezivanje ove dve familije trankripcionih faktora ima poseban značaj kada se ima u vidu njihova uloga tokom razvića nervnog sistema kičmenjaka.Sox3/SOX3 is considered to be one of the earliest neural markers in vertebrates, playing the role in specifying neuronal fate. The characterization of SOX3 promoter was previously reported and it was demonstrated that general transcription factors NF-Y,
and USF are involved in transcriptional regulation of the SOX3 promoter. In this thesis first evidence is presented that TALE transcription factors PBX1, MEIS1 and TGIF are involved in the regulation of human SOX3 gene expression.
PBX1 and MEIS1 proteins are involved in the up-regulation of human SOX3 gene expression in NT2O1 cells by direct interaction with the consensus Pbx/Meis binding site that is conserved in all analyzed mammalian orthologue promoters. PBX1 is present in the protein complex formed on this site with nuclear proteins from uninduced cells, while both, PBX1 and MEIS1 proteins were detected in the complex created with extract from retinoic acid (RA) induced NT2/D1 cells. Interestingly, MEIS1 is the only transcription factor demonstrated so far to interact with SOX3 promoter upon RA induced activation of the SOX3 gene expression. By functional analysis it was demonstrated that mutations of the Pbx1/Meis1 binding sites resulted in profound reduction of the SOX3 promoter responsiveness to RA. Finally, it was demonstrated that overexpressed Pbx1 and Mets1 increased endogenous SOX3 protein expression in both uninduced and RA induced NT2/D1 cells.
Transcription factor TGIF is involved in the down-regulation of human SOX3 gene by direct interaction with the binding site within SOX3 promoter. Functional analysis showed that mutation of the TGIF binding site increased SOX3 expression while overexpressed TGIF decreased SOX3 expression in both uninduced and RA induced NT2/D1 cells.
By data presented here, for the first time, a functional link was established
between TALE proteins, PBX1, MEIS1 and TGIF and expression of human SOX3 gene. This link is of particular interest since both TALE family members and members of SOX super-family are recognized as important developmental regulators
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
The role of TALE transcription factors in the retinoic acid induced activation of SOX3 gene
No full textSox3/SOX3 gen je jedan od najranijih markera neuralnog razvića kičmenjaka, koji ima ključnu ulogu u održavanju populacije ćelija neuralnih progenitora. Naša ranija istraživanja dovela su do prve karakterizacije promotora humanog SOX3 gena i identifikacije kontrolnih elemenata preko kojih su opšti transkripcioni faktori NF-Y, Sp1 i USF uključeni u regulaciju ekspresije ovog gena. Rezultati predstavljeni u ovom radu po prvi put pokazuju da su TALE transkripcioni faktori PBX1, MEIS1 i TGIF uključeni u regulaciju ekspresije humanog SOX3 gena.
PBX1 i MEIS1 proteini direktno interaguju sa konsenzusnim mestom za vezivanje Pbx/Meis heterodimera koje je visoko evolutivno očuvano u bazalnom promotoru Sox3 gena sisara. PBX1 je prisutan u proteinskim kompleksima formiranim na ovom mestu sa jedarnim proteinima iz neindukovanih ćelija, dok su i PBX1 i MEIS1 prisutni u kompleksima formiranim sa jedarnim proteinima iz NT2/D1 ćelija indukovanih retinoičnom kiselinom. Zanimljivo je da je MEIS1 jedini do sada identifikovani faktor koji interaguje sa kontrolnim elementima SOX3 promotora tek nakon indukcije ovog gena retinoičnom kiselinom. Funkcionalne analize su pokazale da mutacije Pbx/Meis vezivnog mesta dovode do pada aktivnosti SOX3 promotora u RA indukovanim NT2/D1 ćelijama, dok povećana ekspresija PBX1 i MEIS1 proteina dovodi do aktivacije SOX3 gena kako u indukovananim tako i u neindukovanim ćelijama.
Transkripcioni faktor TGIF ostvaruje funkciju transkripcionog represora interakcijom sa konsenzusnim vezivnim mestom u promotoru SOX3 gena. Mutacija ovog mesta dovodi do povećane aktivnosti SOX3 promotora, dok povećana ekspresija transkripcionog faktora TGIF značajno smanjuje ekspresiju SOX3 gena u neindukovanim i RA indukovanim NT2/D1 ćelijama.
Prikazani rezultati po prvi put uspostavljaju funkcionalnu vezu izmedu članova TALE familije transkripcionih faktora (PBX1, MEIS1 i TGIF) i ekspresije humanog SOX3 gena. Povezivanje ove dve familije trankripcionih faktora ima poseban značaj kada se ima u vidu njihova uloga tokom razvića nervnog sistema kičmenjaka.Sox3/SOX3 is considered to be one of the earliest neural markers in vertebrates, playing the role in specifying neuronal fate. The characterization of SOX3 promoter was previously reported and it was demonstrated that general transcription factors NF-Y,
and USF are involved in transcriptional regulation of the SOX3 promoter. In this thesis first evidence is presented that TALE transcription factors PBX1, MEIS1 and TGIF are involved in the regulation of human SOX3 gene expression.
PBX1 and MEIS1 proteins are involved in the up-regulation of human SOX3 gene expression in NT2O1 cells by direct interaction with the consensus Pbx/Meis binding site that is conserved in all analyzed mammalian orthologue promoters. PBX1 is present in the protein complex formed on this site with nuclear proteins from uninduced cells, while both, PBX1 and MEIS1 proteins were detected in the complex created with extract from retinoic acid (RA) induced NT2/D1 cells. Interestingly, MEIS1 is the only transcription factor demonstrated so far to interact with SOX3 promoter upon RA induced activation of the SOX3 gene expression. By functional analysis it was demonstrated that mutations of the Pbx1/Meis1 binding sites resulted in profound reduction of the SOX3 promoter responsiveness to RA. Finally, it was demonstrated that overexpressed Pbx1 and Mets1 increased endogenous SOX3 protein expression in both uninduced and RA induced NT2/D1 cells.
Transcription factor TGIF is involved in the down-regulation of human SOX3 gene by direct interaction with the binding site within SOX3 promoter. Functional analysis showed that mutation of the TGIF binding site increased SOX3 expression while overexpressed TGIF decreased SOX3 expression in both uninduced and RA induced NT2/D1 cells.
By data presented here, for the first time, a functional link was established
between TALE proteins, PBX1, MEIS1 and TGIF and expression of human SOX3 gene. This link is of particular interest since both TALE family members and members of SOX super-family are recognized as important developmental regulators
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Epigenetic mechanisms regulating the expression of human SOX3 gene and pluripotency factors (SOX2, OCT4, i NANOG) during the early stages of NTera2/D1 cells neural differentiation
No full textključne aktere u procesima održavanja pluripotentnosti, ćelijskog opredeljivanja i diferencijacije. Precizno koordinisana aktivnost ovih gena neophodna je za pravilan tok humane neuralne diferencijacije. U ovoj doktorskoj disertaciji analizirani su ekspresioni profili i epigenetički mehanizmi regulacije ekspresije pomenutih gena u ranim fazama neuralne diferencijacije NTeraT2/D1 ćelija indukovane retinoičnom kiselinom.
U ovom model sistemu in vitro neuralne diferencijacije detektovane su dinamične izmene u ekspresionim profilima gena SOX1 i SOX3 i gena faktora pluripotentnosti (SOX2, OCT4 i NANOG) tokom ranih faza neuralne diferencijacije. Pokazano je da se ekspresija gena SOX1 i SOX3 aktivira odmah nakon indukcije neuralne diferencijacije ovih ćelija, čime je potvrđena njihova uloga ranih neuralnih markera. Ekspresija gena SOX2, OCT4 i NANOG, koji formiraju jezgro regulatorne mreže odgovorne za održavanje pluripotentnosti, smanjuje se tokom inicijacije neuralne diferencijacije i izlaska NTera2/D1 ćelija iz stanja pluripotentnosti na osnovu čega je zaključeno da signali koji indukuju neuralnu diferencijaciju već u početnim stadijumima dovode do intenzivnih promena u „molekularnom miljeu“ ćelije.
U analizi epigenetičkih mehanizama uključenih u regulaciju ekspresije ispitivanih gena praćeni su metilacija DNK i dinamika izmene histonskih posttranslacionih modifikacija na njihovim regulatornim regionima, sa posebnim osvrtom na regulaciju ekspresije gena SOX3. Metodom metilacija-specifičnog PCR-a pokazano je da su promotori gena SOX1 i SOX3 hipometilovani kako u nediferenciranim NTera2/D1 ćelijama, tako i u ćelijama čija je neuralna diferencijacija indukovana retinoičnom kiselinom. Metodom bisulfitnog pirosekvenciranja DNK, koja predstavlja zlatni standard u merenju stepena metilacije, potvrđena je hipometilacija jednog od dva CpG ostrva u okviru promotora gena SOX3 čime je potvrđeno da metilacija DNK nije mehanizam regulacije ekspresije ovog gena u ranim fazama neuralne diferencijacije NTera2/D1 ćelija.
Metodom hromatinske imunoprecipitacije analizirani su profili odabranih histonskih posttranslacionih modifikacija na regulatornim regionima gena SOX3 i SOX1 kao i gena faktora pluripotentnosti (SOX2, OCT4 i NANOG). Pokazano je da su izmene u depoziciji aktivirajućih histonskih markera H3K4me3 i H2B-acetil na regulatornim regionima gena SOX3 u korelaciji sa indukcijom ekspresije ovog gena u ranim fazama neuralne diferencijacije NTera2/D1 ćelija. Inicijacija neuralne diferencijacije praćena je smanjenjem prisustva H3K4me3 i H2B-acetil markera
na promotoru gena SOX2, dok nijedna od analiziranih histonskih modifikacija nije u korelaciji sa detektovanim ekspresionim profilom gena SOX1...genes SOX1 and SOX3, together with pluripotency factors (SOX2, OCT4 and NANOG), have key roles in pluripotency maintenace, cell specification and differentiation and they act in a coordinate manner to regulate process of human neurogenesis. This doctoral disertation provides an insight into the expression profiles and epigenetic regulation of these genes during the early phases of retinoic acid-induced neural differentiation of NTera2/D1 cells.
In this in vitro model system of human neurogenesis we have demontrated that expression profiles of SOX3 and SOX1 genes and pluripotency factors (SOX2, OCT4 i NANOG) are highly dynamic. It is shown that retinoic acid activates expression of SOX1 and SOX3, confirming their roles as early neural markers in vertebrates. Expression of SOX2, OCT4 and NANOG, which constitute core pluripotency regulatory circuitry, is reduced during the course of neural differentiation, suggesting that response to differentitation-inducing stimuli is followed by fast and intensive changes in cell transcriptome.
For the analyses of epigenetic mechanisms acting on regulatory regions of SOX1 and SOX3 genes and pluripotency factors, DNA methylation and histone modifications profiles were examined, with special emphasis on SOX3 gene regulation. Using methylation-specific PCR it is shown that SOX1 and SOX3 gene promoters are hypomethylated in both undifferentiated NTera2/D1 cells and cells induced with retinoic acid. Bisulphite sequencing of the CpG island II in the promoter of SOX3 gene further confirmed that DNA methylation is not a mechanism governing regulation of SOX3 activity during the early stages of NTera2/D1 cell differentiation.
Selected histone posttranslational modifications profiles on the regulatory regions of SOX1 and SOX3 genes and pluripotency factors were analyzed using chromatin immunoprecipitation. Changes in abundance of active H3K4me3 and H2B-acetyl marks on SOX3 regulatory regions are correlated with transcriptional activation of this gene. Furthermore, it is demonstrated that initiation of neural differentiation is accompanied by the decrease in H3K4me3 and H2B-acetyl marks on the promoter of SOX2 gene, while none of the analyzed histone modifications was correlated with previously detected SOX1 expression profile. In silico analyses demonstrated that histone profiles on the promoter regions of SOX1 and SOX3 genes in H1 human embryonal stem cells corresponds to bivalent genes with low transcriptional activity, while SOX2 gene promoter is enriched with activating histone marks. Obtained results have demonstrated important differences in the epigenetic mechanisms regulating the expression of SOXB1 family members, despite the existence of their functional redundancy..
Epigenetic mechanisms regulating the expression of human SOX3 gene and pluripotency factors (SOX2, OCT4, i NANOG) during the early stages of NTera2/D1 cells neural differentiation
Full text linkključne aktere u procesima održavanja pluripotentnosti, ćelijskog opredeljivanja i diferencijacije. Precizno koordinisana aktivnost ovih gena neophodna je za pravilan tok humane neuralne diferencijacije. U ovoj doktorskoj disertaciji analizirani su ekspresioni profili i epigenetički mehanizmi regulacije ekspresije pomenutih gena u ranim fazama neuralne diferencijacije NTeraT2/D1 ćelija indukovane retinoičnom kiselinom.
U ovom model sistemu in vitro neuralne diferencijacije detektovane su dinamične izmene u ekspresionim profilima gena SOX1 i SOX3 i gena faktora pluripotentnosti (SOX2, OCT4 i NANOG) tokom ranih faza neuralne diferencijacije. Pokazano je da se ekspresija gena SOX1 i SOX3 aktivira odmah nakon indukcije neuralne diferencijacije ovih ćelija, čime je potvrđena njihova uloga ranih neuralnih markera. Ekspresija gena SOX2, OCT4 i NANOG, koji formiraju jezgro regulatorne mreže odgovorne za održavanje pluripotentnosti, smanjuje se tokom inicijacije neuralne diferencijacije i izlaska NTera2/D1 ćelija iz stanja pluripotentnosti na osnovu čega je zaključeno da signali koji indukuju neuralnu diferencijaciju već u početnim stadijumima dovode do intenzivnih promena u „molekularnom miljeu“ ćelije.
U analizi epigenetičkih mehanizama uključenih u regulaciju ekspresije ispitivanih gena praćeni su metilacija DNK i dinamika izmene histonskih posttranslacionih modifikacija na njihovim regulatornim regionima, sa posebnim osvrtom na regulaciju ekspresije gena SOX3. Metodom metilacija-specifičnog PCR-a pokazano je da su promotori gena SOX1 i SOX3 hipometilovani kako u nediferenciranim NTera2/D1 ćelijama, tako i u ćelijama čija je neuralna diferencijacija indukovana retinoičnom kiselinom. Metodom bisulfitnog pirosekvenciranja DNK, koja predstavlja zlatni standard u merenju stepena metilacije, potvrđena je hipometilacija jednog od dva CpG ostrva u okviru promotora gena SOX3 čime je potvrđeno da metilacija DNK nije mehanizam regulacije ekspresije ovog gena u ranim fazama neuralne diferencijacije NTera2/D1 ćelija.
Metodom hromatinske imunoprecipitacije analizirani su profili odabranih histonskih posttranslacionih modifikacija na regulatornim regionima gena SOX3 i SOX1 kao i gena faktora pluripotentnosti (SOX2, OCT4 i NANOG). Pokazano je da su izmene u depoziciji aktivirajućih histonskih markera H3K4me3 i H2B-acetil na regulatornim regionima gena SOX3 u korelaciji sa indukcijom ekspresije ovog gena u ranim fazama neuralne diferencijacije NTera2/D1 ćelija. Inicijacija neuralne diferencijacije praćena je smanjenjem prisustva H3K4me3 i H2B-acetil markera
na promotoru gena SOX2, dok nijedna od analiziranih histonskih modifikacija nije u korelaciji sa detektovanim ekspresionim profilom gena SOX1...genes SOX1 and SOX3, together with pluripotency factors (SOX2, OCT4 and NANOG), have key roles in pluripotency maintenace, cell specification and differentiation and they act in a coordinate manner to regulate process of human neurogenesis. This doctoral disertation provides an insight into the expression profiles and epigenetic regulation of these genes during the early phases of retinoic acid-induced neural differentiation of NTera2/D1 cells.
In this in vitro model system of human neurogenesis we have demontrated that expression profiles of SOX3 and SOX1 genes and pluripotency factors (SOX2, OCT4 i NANOG) are highly dynamic. It is shown that retinoic acid activates expression of SOX1 and SOX3, confirming their roles as early neural markers in vertebrates. Expression of SOX2, OCT4 and NANOG, which constitute core pluripotency regulatory circuitry, is reduced during the course of neural differentiation, suggesting that response to differentitation-inducing stimuli is followed by fast and intensive changes in cell transcriptome.
For the analyses of epigenetic mechanisms acting on regulatory regions of SOX1 and SOX3 genes and pluripotency factors, DNA methylation and histone modifications profiles were examined, with special emphasis on SOX3 gene regulation. Using methylation-specific PCR it is shown that SOX1 and SOX3 gene promoters are hypomethylated in both undifferentiated NTera2/D1 cells and cells induced with retinoic acid. Bisulphite sequencing of the CpG island II in the promoter of SOX3 gene further confirmed that DNA methylation is not a mechanism governing regulation of SOX3 activity during the early stages of NTera2/D1 cell differentiation.
Selected histone posttranslational modifications profiles on the regulatory regions of SOX1 and SOX3 genes and pluripotency factors were analyzed using chromatin immunoprecipitation. Changes in abundance of active H3K4me3 and H2B-acetyl marks on SOX3 regulatory regions are correlated with transcriptional activation of this gene. Furthermore, it is demonstrated that initiation of neural differentiation is accompanied by the decrease in H3K4me3 and H2B-acetyl marks on the promoter of SOX2 gene, while none of the analyzed histone modifications was correlated with previously detected SOX1 expression profile. In silico analyses demonstrated that histone profiles on the promoter regions of SOX1 and SOX3 genes in H1 human embryonal stem cells corresponds to bivalent genes with low transcriptional activity, while SOX2 gene promoter is enriched with activating histone marks. Obtained results have demonstrated important differences in the epigenetic mechanisms regulating the expression of SOXB1 family members, despite the existence of their functional redundancy..
Epigenetic mechanisms regulating the expression of human SOX3 gene and pluripotency factors (SOX2, OCT4, i NANOG) during the early stages of NTera2/D1 cells neural differentiation
No full textključne aktere u procesima održavanja pluripotentnosti, ćelijskog opredeljivanja i diferencijacije. Precizno koordinisana aktivnost ovih gena neophodna je za pravilan tok humane neuralne diferencijacije. U ovoj doktorskoj disertaciji analizirani su ekspresioni profili i epigenetički mehanizmi regulacije ekspresije pomenutih gena u ranim fazama neuralne diferencijacije NTeraT2/D1 ćelija indukovane retinoičnom kiselinom.U ovom model sistemu in vitro neuralne diferencijacije detektovane su dinamične izmene u ekspresionim profilima gena SOX1 i SOX3 i gena faktora pluripotentnosti (SOX2, OCT4 i NANOG) tokom ranih faza neuralne diferencijacije. Pokazano je da se ekspresija gena SOX1 i SOX3 aktivira odmah nakon indukcije neuralne diferencijacije ovih ćelija, čime je potvrđena njihova uloga ranih neuralnih markera. Ekspresija gena SOX2, OCT4 i NANOG, koji formiraju jezgro regulatorne mreže odgovorne za održavanje pluripotentnosti, smanjuje se tokom inicijacije neuralne diferencijacije i izlaska NTera2/D1 ćelija iz stanja pluripotentnosti na osnovu čega je zaključeno da signali koji indukuju neuralnu diferencijaciju već u početnim stadijumima dovode do intenzivnih promena u „molekularnom miljeu“ ćelije.U analizi epigenetičkih mehanizama uključenih u regulaciju ekspresije ispitivanih gena praćeni su metilacija DNK i dinamika izmene histonskih posttranslacionih modifikacija na njihovim regulatornim regionima, sa posebnim osvrtom na regulaciju ekspresije gena SOX3. Metodom metilacija-specifičnog PCR-a pokazano je da su promotori gena SOX1 i SOX3 hipometilovani kako u nediferenciranim NTera2/D1 ćelijama, tako i u ćelijama čija je neuralna diferencijacija indukovana retinoičnom kiselinom. Metodom bisulfitnog pirosekvenciranja DNK, koja predstavlja zlatni standard u merenju stepena metilacije, potvrđena je hipometilacija jednog od dva CpG ostrva u okviru promotora gena SOX3 čime je potvrđeno da metilacija DNK nije mehanizam regulacije ekspresije ovog gena u ranim fazama neuralne diferencijacije NTera2/D1 ćelija.Metodom hromatinske imunoprecipitacije analizirani su profili odabranih histonskih posttranslacionih modifikacija na regulatornim regionima gena SOX3 i SOX1 kao i gena faktora pluripotentnosti (SOX2, OCT4 i NANOG). Pokazano je da su izmene u depoziciji aktivirajućih histonskih markera H3K4me3 i H2B-acetil na regulatornim regionima gena SOX3 u korelaciji sa indukcijom ekspresije ovog gena u ranim fazama neuralne diferencijacije NTera2/D1 ćelija. Inicijacija neuralne diferencijacije praćena je smanjenjem prisustva H3K4me3 i H2B-acetil markerana promotoru gena SOX2, dok nijedna od analiziranih histonskih modifikacija nije u korelaciji sa detektovanim ekspresionim profilom gena SOX1...genes SOX1 and SOX3, together with pluripotency factors (SOX2, OCT4 and NANOG), have key roles in pluripotency maintenace, cell specification and differentiation and they act in a coordinate manner to regulate process of human neurogenesis. This doctoral disertation provides an insight into the expression profiles and epigenetic regulation of these genes during the early phases of retinoic acid-induced neural differentiation of NTera2/D1 cells.In this in vitro model system of human neurogenesis we have demontrated that expression profiles of SOX3 and SOX1 genes and pluripotency factors (SOX2, OCT4 i NANOG) are highly dynamic. It is shown that retinoic acid activates expression of SOX1 and SOX3, confirming their roles as early neural markers in vertebrates. Expression of SOX2, OCT4 and NANOG, which constitute core pluripotency regulatory circuitry, is reduced during the course of neural differentiation, suggesting that response to differentitation-inducing stimuli is followed by fast and intensive changes in cell transcriptome.For the analyses of epigenetic mechanisms acting on regulatory regions of SOX1 and SOX3 genes and pluripotency factors, DNA methylation and histone modifications profiles were examined, with special emphasis on SOX3 gene regulation. Using methylation-specific PCR it is shown that SOX1 and SOX3 gene promoters are hypomethylated in both undifferentiated NTera2/D1 cells and cells induced with retinoic acid. Bisulphite sequencing of the CpG island II in the promoter of SOX3 gene further confirmed that DNA methylation is not a mechanism governing regulation of SOX3 activity during the early stages of NTera2/D1 cell differentiation.Selected histone posttranslational modifications profiles on the regulatory regions of SOX1 and SOX3 genes and pluripotency factors were analyzed using chromatin immunoprecipitation. Changes in abundance of active H3K4me3 and H2B-acetyl marks on SOX3 regulatory regions are correlated with transcriptional activation of this gene. Furthermore, it is demonstrated that initiation of neural differentiation is accompanied by the decrease in H3K4me3 and H2B-acetyl marks on the promoter of SOX2 gene, while none of the analyzed histone modifications was correlated with previously detected SOX1 expression profile. In silico analyses demonstrated that histone profiles on the promoter regions of SOX1 and SOX3 genes in H1 human embryonal stem cells corresponds to bivalent genes with low transcriptional activity, while SOX2 gene promoter is enriched with activating histone marks. Obtained results have demonstrated important differences in the epigenetic mechanisms regulating the expression of SOXB1 family members, despite the existence of their functional redundancy..
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
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