Sostdc1 Plays an Essential Role in Mammalian Tooth Patterning : Insight into the Rodent Dental Evolution

This thesis work focuses on the role of TGF-beta family antagonists during the development of mouse dentition. Tooth develops through an interaction between the dental epithelium and underlying neural crest derived mesenchyme. The reciprocal signaling between these tissues is mediated by soluble signaling molecules and the balance between activatory and inhibitory signals appears to be essential for the pattern formation. We showed the importance of Sostdc1 in the regulation of tooth shape and number. The absence of Sostdc1 altered the molar cusp patterning and led to supernumerary tooth formation both in the molar and incisor region. We showed that initially, Sostdc1 expression is in the mesenchyme, suggesting that dental mesenchyme may limit supernumerary tooth induction. We tested this in wild-type incisors by minimizing the amount of mesenchymal tissue surrounding the incisor tooth germs prior to culture in vitro. The cultured teeth phenocopied the extra incisor phenotype of the Sostdc1-deficient mice. Furthermore, we showed that minimizing the amount of dental mesenchyme in cultured Sostdc1-deficient incisors caused the formation of additional de novo incisors that resembled the successional incisor development resulting from activated Wnt signaling. Sostdc1 seemed to be able to inhibit both mesenchymal BMP4 and epithelial canonical Wnt signaling, which thus allows Sostdc1 to restrict the enamel knot size and regulate the tooth shape and number. Our work emphasizes the dual role for the tooth mesenchyme as a suppressor as well as an activator during tooth development. We found that the placode, forming the thick mouse incisor, is prone to disintegration during initiation of tooth development. The balance between two mesenchymal TGF-beta family signals, BMP4 and Activin is essential in this regulation. The inhibition of BMP4 or increase in Activin signaling led to the splitting of the large incisor placode into two smaller placodes resulting in thin incisors. These two signals appeared to have different effects on tooth epithelium and the analysis of the double null mutant mice lacking Sostdc1 and Follistatin indicated that these TGF-beta inhibitors regulate the mutual balance of BMP and Activin in vivo. In addition, this work provides an alternative explanation for the issue of incisor identity published in Science by Tucker et al. in 1998 and proposes that the molar like morphology that can be obtained by inhibiting BMP signaling is due to partial splitting of the incisor placodes and not due to change in tooth identity from the incisor to the molar. This thesis work presents possible molecular mechanisms that may have modified the mouse dental pattern during evolution leading to the typical rodent dentition of modern mouse. The rodent dentition is specialized for gnawing and consists of two large continuously growing incisors and toothless diastema region separating the molars and incisors. The ancestors of rodents had higher number of more slender incisors together with canines and premolars. Additionally, murine rodents, which include the mouse, have lost their ability for tooth replacement. This work has revealed that the inhibitory molecules appear to play a role in the tooth number suppression by delineating the spatial and temporal action of the inductive signals. The results suggest that Sostdc1 plays an essential role in several stages of tooth development through the regulation of both the BMP and Wnt pathway. The work shows a dormant sequential tooth forming potential present in wild type mouse incisor region and gives a new perspective on tooth suppression by dental mesenchyme. It reveals as well a novel mechanism to create a large mouse incisor through the regulation of mesenchymal balance between inductive and inhibitory signals.

Substrate specificity plays an important role in uncoupling the catalytic and scaffolding activities of rat testis DNA topoisomerase IIalpha

Topoisomerase II (topo II) is a dyadic enzyme found in all eukaryotic cells. Topo II is involved in a number of cellular processes related to DNA metabolism, including DNA replication, recombination and the maintenance of genomic stability. We discovered a correlation between the development of postnatal testis and increased binding of topo IIalpha to the chromatin fraction. We used this observation to characterize DNA-binding specificity and catalytic properties of purified testis topo IIalpha. The results indicate that topo IIalpha binds a substrate containing the preferred site with greater affinity and, consequently, catalyzes the conversion of form I to form IV DNA more efficiently in contrast to substrates lacking such a site. Interestingly, topo IIalpha displayed high-affinity and cooperativity in binding to the scaffold associated region. In contrast to the preferred site, however, high-affinity binding of topo IIalpha to the scaffold-associated region failed to result in enhanced catalytic activity. Intriguingly, competition assays involving scaffold-associated region revealed an additional DNA-binding site within the dyadic topo IIalpha. These results implicate a dual role for topo IIalpha in vivo consistent with the notion that its sequestration to the chromatin might play a role in chromosome condensation and decondensation during spermatogenesis.

The prankster’s passion: Using theatrical terms to unpack players motivations in popular social performance

Amongst social players, the prank, as a social performance form, holds a lot of potential to impact on personal, relational and social status within a group or between one group and another group. More than simply showing off, a prank in the strictest definition of the term, is a social performance in which one player, a prankster, deploys mischief, trickery or deceit, to cause a moment of anxiety, fear or anger about a happening for another spectator-become-collaborating-player, a prankee – to enhance social bonds, entertain, or comment on a social, cultural or political phenomenon. During a prank, the prankster’s ability to be creative, clever or culturally astute, and the prankee’s ability to be duped, be a good sport, play along, or even play/pay the prankster back, both become fodder for other spectators and society to scrutinize. In Australia, pranking traditions are popular with many social groups, from the community-building pranks of footballers, bucks parties and ‘drop bear’ tales told to tourists, to the more controversial pranks of radio shock jocks, activists and artists. In this paper, I consider whether theatrical terms – theoretical terms from the stage such as actor, acting, objective, arc, performance, audience and emotion, such as those offered by Joseph Roach – are useful in understanding the passion some social players show for pranksterism. Are theatrical terms such as Roach’s as useful as analysts of social self-performance such as Erving Goffman suggest they are? Do they assist in understanding the personal actions, reactions and emotions of prankster and prankee? Do they assist in understanding the power relations between prankster and prankee? Do they assist in understanding the relation between the prank – be it an everyday prank amongst families, friends and coworkers, an entertainment program prank of the sort seen on Prank Patrol, Punked or Scare Tactics, or an activist pranks perpetrated by a guerrilla artist, ‘jammers’ or ‘hackers’ intent on turning dominant social systems back on themselves – the social players, and the public sphere in which the prank takes place? I reflect on how reading pranks as performances, by players, for highly participatory audiences, helps understand why they are so prevalent, and so recurrent across times, cultures and contexts, and also so controversial when not performed well enough – or when performed too well – prompting outrage from the prankster, prankee or society as passionate as any debate about a performance by players in a theatre.

Unraveling the “passion orchestra” in academia

This paper disentangles how organization members' “passion orchestra” is related to their entrepreneurial intentions in the particularly relevant context of academia. Drawing on passion literature and identity theory, we propose and test a model linking two central parts of researchers' “passion orchestra”, namely entrepreneurial and obsessive scientific passion, directly and indirectly, to spin-off and start-up intentions. While spin-off intentions refer to intentions to found a firm based upon research results, start-up intentions denote intentions to start any type of company. Using a sample of 2308 researchers from 24 European universities, our findings reveal that higher levels of entrepreneurial passion are associated with both stronger spin-off and start-up intentions. Further, obsessive scientific passion is positively associated with researchers' intentions to create a spin-off, and negatively with their propensity to establish a start-up. Entrepreneurial self-efficacy and affective organizational commitment mediate these effects. Finally, the two types of passion show characteristic interactions. Obsessive scientific passion moderates the entrepreneurial passion–intentions relationship such that it strengthens spin-off intentions. Our results highlight that recasting the individual driven by a singular passion to one with a “passion orchestra” provides a more holistic understanding of the new venture creation process. Implications for research and practice are discussed.

The Multifunctional PE_PGRS11 Protein from Mycobacterium tuberculosis Plays a Role in Regulating Resistance to Oxidative Stress

Mycobacterium tuberculosis utilizes unique strategies to survive amid the hostile environment of infected host cells. Infection-specific expression of a unique mycobacterial cell surface antigen that could modulate key signaling cascades can act as a key survival strategy in curtailing host effector responses like oxidative stress. We demonstrate here that hypothetical PE_PGRS11 ORF encodes a functional phosphoglycerate mutase. The transcriptional analysis revealed that PE_PGRS11 is a hypoxia-responsive gene, and enforced expression of PE_PGRS11 by recombinant adenovirus or Mycobacterium smegmatis imparted resistance to alveolar epithelial cells against oxidative stress. PE_PGRS11-induced resistance to oxidative stress necessitated the modulation of genetic signatures like induced expression of Bcl2 or COX-2. This modulation of specific antiapoptotic molecular signatures involved recognition of PE_PGRS11 by TLR2 and subsequent activation of the PI3K-ERK1/ 2-NF-kappa B signaling axis. Furthermore, PE_PGRS11 markedly diminished H2O2-induced p38 MAPK activation. Interestingly, PE_PGRS11 protein was exposed at the mycobacterial cell surface and was involved in survival of mycobacteria under oxidative stress. Furthermore, PE_PGRS11 displayed differential B cell responses during tuberculosis infection. Taken together, our investigation identified PE_PGRS11 as an in vivo expressed immunodominant antigen that plays a crucial role in modulating cellular life span restrictions imposed during oxidative stress by triggering TLR2-dependent expression of COX-2 and Bcl2. These observations clearly provide a mechanistic basis for the rescue of pathogenic Mycobacterium-infected lung epithelial cells from oxidative stress.

Väkivalta, antisemitismi ja konflikti Mel Gibsonin elokuvassa The Passion of The Christ

Tutkielman tarkoituksena on tutkia Mel Gibsonin elokuvaa The Passion of the Christ. Tarkastelussa mukana on myös Gibsonin muu tuotanto. Tutkielma edustaa poikkitieteellistä lähestymistapaa käyttäen eksegetiikan apuvälineitä kuten lähdekritiikkiä, mutta myös elokuvatutkimuksen välineitä. Tutkimuskysymyksiä nousee lähinnä kaksi, joista seuraa kolmas kysymys: 1. Onko elokuva juutalaisvastainen? Tarkastelen lähdekritiikin avulla mitä evankeliumeja Gibson on käyttänyt elokuvassaan. Mitä muita lähteitä hän on käyttänyt? Mikä on Gibsonin omaa ilmaisua? 2. Miksi elokuva on väkivaltainen? Mitä väkivalta palvelee elokuvassa? 3. Millainen on elokuvamaailman konflikti? Pohdin elokuvamaailman sisälle rakennettua konfliktia, mutta viittaan sillä myös konfliktiin, jonka elokuva itsessään synnytti. Kysymysten ratkaisu vaatii elokuvassa käytettyjen lähteiden tutkimista, mutta myös kysymysten tarkastelua osana laajempaa kokonaisuutta, jossa on mukana koko Gibsonin elokuvatuotanto. On myös selvää, ettei Gibson ole yhtä kuin hänen elokuvansa, mutta toisaalta hänen elokuviaan ei voi tarkastella irrotettuna ohjaajasta itsestään. Ensimmäisessä luvussa tarkastelen elokuvaa ilmiönä ja elokuvasta käytyä ennakkokeskustelua. Luvussa kaksi tarkastelen Gibsonin taustaa. Millaisista lähtökohdista Gibson lähti tekemään elokuvaa? Luvussa kolme esittelen käsikirjoituksesta alkavan elokuvan yleisen tuotantoprosessin. Tutkielman päälähteenä olen käyttänyt Brentanon kirjaa The Dolorous Passion of Our Lord Jesus Christ. Kirja pohjautuu 1800-luvulla eläneen katolisen nunnan, Anne Catherine Emmerichin näkyihin. Luvussa neljä tarkastelen Gibsonin muuta tuotantoa, ja tuon keskusteluun mukaan Scorsesen elokuvan Jeesuksesta. Gibsonin muu tuotanto on jäänyt tutkijoilta liian vähälle huomiolle. Elokuva The Passion of the Christ on nähtävä osana Gibsonin muuta tuotantoa. Näin ollen elokuvaa The Passion of the Christ voidaan ymmärtää paremmin. Luvussa viisi käyn elokuvan The Passion of the Christ läpi kappale kerrallaan tutkimalla, mitä lähteitä Gibson on käyttänyt elokuvassaan. Mitä hän on ottanut evankeliumeista, mitä Emmerichiltä ja mikä on hänen omaa ilmaisuaan? Luvussa kuusi käyn läpi elokuvan vastaanottoa niin raamatuntutkijoiden kuin suuren yleisön parissa. Tutkielmassa todetaan, ettei Gibson ole antisemitisti, vaan ksenofobinen rasisti. Hänen elokuvansa ovat ksenofobisesti rasistisia. Gibsonin kaikista elokuvista on löydettävissä itseään toistavia piirteitä, joissa esiintyy muukalaiskammoa ja väkivaltaa. Gibsonin nimittäminen antisemitistiksi ei tekisi Gibsonille oikeutta. Juutalaiset ovat vain osa laajempaa kokonaisuutta. Väkivalta palvelee kaikissa elokuvissa uuden, Jumalan valtakunnan syntymistä. Konflikti syntyy uuden ja vanhan valtakunnan kansalaisten välillä. Uhrien veren kautta syntyy Jumalan valtakunta. Johtopäätöksillä on merkitystä niin Gibsonin kuin hänen elokuviensa ymmärtämiselle. Elokuvan The Passion of the Christ tulevissa tutkimuksissa on otettava huomioon, ei vain Gibsonin tausta ja lähteet, vaan myös Gibsonin muu elokuvatuotanto.

UDP-glucose 4, 6-dehydratase Activity Plays an Important Role in Maintaining Cell Wall Integrity and Virulence of Candida albicans

Candida albicans, a human fungal pathogen, undergoes morphogenetic changes that are associated with virulence. We report here that GAL102 in C. albicans encodes a homolog of dTDP-glucose 4,6-dehydratase, an enzyme that affects cell wall properties as well as virulence of many pathogenic bacteria. We found that GAL102 deletion leads to greater sensitivity to antifungal drugs and cell wall destabilizing agents like Calcofluor white and Congo red. The mutant also formed biofilms consisting mainly of hyphal cells that show less turgor. The NMR analysis of cell wall mannans of gal102 deletion strain revealed that a major constituent of mannan is missing and the phosphomannan component known to affect virulence is greatly reduced. We also observed that there was a substantial reduction in the expression of genes involved in biofilm formation but increase in the expression of genes encoding glycosylphosphatidylinositol-anchored proteins in the mutant. These, along with altered mannosylation of cell wall proteins together might be responsible for multiple phenotypes displayed by the mutant. Finally, the mutant was unable to grow in the presence of resident peritoneal macrophages and elicited a weak pro-inflammatory cytokine response in vitro. Similarly, this mutant elicited a poor serum pro-inflammatory cytokine response as judged by IFN gamma and TNF alpha levels and showed reduced virulence in a mouse model of systemic candidiasis. Importantly, an Ala substitution for a conserved Lys residue in the active site motif YXXXK, that abrogates the enzyme activity also showed reduced virulence and increased filamentation similar to the gal102 deletion strain. Since inactivating the enzyme encoded by GAL102 makes the cells sensitive to antifungal drugs and reduces its virulence, it can serve as a potential drug target in combination therapies for C. albicans and related pathogens.

Autophosphorylation of Ser(428) of EhC2PK Plays a Critical Role in Regulating Erythrophagocytosis in the Parasite Entamoeba histolytica

The protozoan parasite Entamoeba histolytica can invade both intestinal and extra intestinal tissues resulting in amoebiasis. During the process of invasion E. histolytica ingests red blood and host cells using phagocytic processes. Though phagocytosis is considered to be a key virulence determinant, the mechanism is not very well understood in E. histolytica. We have recently demonstrated that a novel C2 domain-containing protein kinase, EhC2PK is involved in the initiation of erythrophagocytosis. Because cells overexpressing the kinase-dead mutant of EhC2PK displayed a reduction in erythrophagocytosis, it appears that kinase activity is necessary for initiation. Biochemical analysis showed that EhC2PK is an unusual Mn2+-dependent serine kinase. It has a trans-autophosphorylated site at Ser(428) as revealed by mass spectrometric and biochemical analysis. The autophosphorylation defective mutants (S428A, KD Delta C) showed a reduction in auto and substrate phosphorylation. Time kinetics of in vitro kinase activity suggested two phases, an initial short slow phase followed by a rapid phase for wild type protein, whereas mutations in the autophosphorylation sites that cause defect (S428A) or conferred phosphomimetic property (S428E) displayed no distinct phases, suggesting that autophosphorylation may be controlling kinase activity through an autocatalytic mechanism. A reduction and delay in erythrophagocytosis was observed in E. histolytica cells overexpressing S428A and KD Delta C proteins. These results indicate that enrichment of EhC2PK at the site of phagocytosis enhances the rate of trans-autophosphorylation, thereby increasing kinase activity and regulating the initiation of erythrophagocytosis in E. histolytica.

SUB1 Plays a Negative Role during Starvation Induced Sporulation Program in Saccharomyces cerevisiae

Saccharomyces cerevisiae Sub1 is involved in several cellular processes such as, transcription initiation, elongation, mRNA processing and DNA repair. It has also been reported to provide cellular resistance during conditions of oxidative DNA damage and osmotic stress. Here, we report a novel role of SUB1 during starvation stress-induced sporulation, which leads to meiosis and spore formation in diploid yeast cells. Deletion of SUB1 gene significantly increased sporulation efficiency as compared to the wild-type cells in S288c genetic background. Whereas, the sporulation functions of the sub1(Y66A) missense mutant were similar to Sub1. SUB1 transcript and protein levels are downregulated during sporulation, in highly synchronized and sporulation proficient wild-type SK1 cells. The changes in Sub1 levels during sporulation cascade correlate with the induction of middle sporulation gene expression. Deletion of SUB1 increased middle sporulation gene transcript levels with no effect on their induction kinetics. In wild-type cells, Sub1 associates with chromatin at these loci in a temporal pattern that correlates with their enhanced gene expression seen in sub1. cells. We show that SUB1 genetically interacts with HOS2, which led us to speculate that Sub1 might function with Set3 repressor complex during sporulation. Positive Cofactor 4, human homolog of Sub1, complemented the sub1. sporulation phenotype, suggesting conservation of function. Taken together, our results suggest that SUB1 acts as a negative regulator of sporulation.

Expression of VEGFR-2 on HaCaT cells is regulated by VEGF and plays an active role in mediating VEGF induced effects

Vascular endothelial growth factor (VEGF) and its receptor VEGFR-2 play important roles in mitogenesis and chemotaxis of endothelial cells. In normal human skin, VEGF is expressed and secreted by epidermal keratinocytes. Emerging data suggest that keratin

Insulin-like growth factor-binding protein-3 plays an important role in regulating pharyngeal skeleton and inner ear formation and differentiation

Insulin-like growth factor-binding protein (IGFBP)-3 is the major insulin-like growth factor (IGF) carrier protein in the bloodstream. IGFBP-3 prolongs the half-life of circulating IGFs and prevents their potential hypo-glycemic effect. IGFBP-3 is also expressed in many peripheral tissues in fetal and adult stages. In vitro, IGFBP-3 can inhibit or potentiate IGF actions and even possesses IGF-independent activities, suggesting that local IGFBP-3 may also have paracrine/autocrine function(s). The in vivo function of IGFBP-3, however, is unclear. In this study, we elucidate the developmental role of IGFBP-3 using the zebrafish model. IGFBP-3 mRNA expression is first detected in the migrating cranial neural crest cells and subsequently in pharyngeal arches in zebrafish embryos. IGFBP-3 mRNA is also persistently expressed in the developing inner ears. To determine the role of IGFBP-3 in these tissues, we ablated the IGFBP-3 gene product using morpholino-modified antisense oligonucleotides (MOs). The IGFBP-3 knocked down embryos had delayed pharyngeal skeleton morphogenesis and greatly reduced pharyngeal cartilage differentiation. Knockdown of IGFBP-3 also significantly decreased inner ear size and disrupted hair cell differentiation and semicircular canal formation. Furthermore, reintroduction of a MO-resistant form of IGFBP-3 "rescued" the MO-induced defects. These findings suggest that IGFBP-3 plays an important role in regulating pharyngeal cartilage and inner car development and growth in zebrafish.

'Politics, Passion, Prejudice: Alice Childress's Wedding Band: A Love/Hate Story in Black and White'

Cashman, N. (2009). 'Politics, Passion, Prejudice: Alice Childress's Wedding Band: A Love/Hate Story in Black and White', Journal of American Studies, 43, 3, pp. 407?423 Sponsorship: APRS