Using modern microscopy and image analysis methods to study dosage compensation in C. elegans

Condensine sind essentiell für die Faltung von Chromatin und wurden auch mit der Transkriptionsregulation in Verbindung gebracht. Der zugrunde liegende Mechanismus für die Transkriptionsregulation ist jedoch unklar. Condensin DC in C. elegans ist ein gutes Modell zur Erforschung der Transkriptionsregulation durch Condensine, da es spezifisch für die Dosiskompensation der Gene auf dem X Chromosom benutzt wird. Condensin DC bindet an beide X Chromosome in C. elegans Hermaphroditen und reduziert deren Transkription um die Hälfte. In meiner Dissertation habe ich untersucht, welche Rolle ein dynamisches Binden von Condensin DC an Chromatin spielt und wie dies die Transkription während der Embryogenese reguliert. Condensine binden dynamisch an Chromatin, um es zu komprimieren und durch Bildung von Schlaufen die Transkription zu regulieren. Mit Hilfe von „fluorescence recovery after photobleaching“ (FRAP) habe ich in adulten Darmzellen von C. elegans untersucht, welche Faktoren das dynamische Binden von Condensin DC an die X Chromosomen beeinflussen. Meine Daten zeigen, dass sowohl die ATPase-Domäne von Condensin DC, als auch eine nicht-katalytische Aktivität einer Histon-Demethylase die Bindedynamik von Condensin DC beeinflussen und damit Transkription regulieren. Zusätzlich habe ich mit einem Mikroskopieansatz, der auf dem Nachweis von einzelnen RNA Molekülen beruht (smFISH), die Transkription von mehreren Genen untersucht, die durch Condensin DC während der Embryonalentwicklung reguliert werden. Die aus diesen Daten ermittelten Transkriptionskinetiken deuten darauf hin, dass Condensin DC vorrangig die Häufigkeit der Transkriptionsinitiation reguliert. Zusammenfassend liefert meine Forschung neue Einblicke in die Transkriptionsregulation durch Condensine und kann als Basis für detailliertere, mechanistische Studien der Rolle von Condensinen in der Transkriptionsregulation in C. elegans und auch in anderen Organismen dienen.Condensins are essential for chromosome compaction and have been implicated in transcription regulation. The mechanistic foundation of this regulatory function is poorly understood. A clear paradigm to address this question is the X-specific condensin DC in C. elegans, which specifically binds to and transcriptionally represses X chromosomes in XX hermaphrodites by 2-fold. In my thesis, I studied condensin DC binding dynamics to the X chromosome and how condensin DC affects transcription kinetics in single embryos. The binding of condensins to chromatin has been described in recent microscopy-based studies as dynamic in processes including loop formation, chromatin compaction and transcription regulation. To study the dynamics of condensin DC binding, I established fluorescence recovery after photobleaching (FRAP) in C. elegans adult intestinal cells. With this method, I studied how the ATPase domain and different histone modifiers regulate the dynamic binding of condensin DC. I found that the ATPase domain is critical for binding of the complex and that the noncatalytic activity of a histone demethylase increases the dynamics of condensin DC binding, which is crucial for its role in transcription regulation. To further study the mechanism of condensin DC in transcription regulation, I used an imaging approach based on widefield single-molecule RNA fluorescence in situ hybridization (smFISH). I obtained thousands of smFISH images for a set of condensin DC-regulated genes and extracted mature and nascent RNA counts in 3D, which I used to determine transcription burst characteristics throughout embryonic development. My data show that condensin DC regulates the frequency of transcription initiation to down-regulate X-chromosomal genes. Taken together, my results provide new insight into condensin-mediated transcription regulation, which can be used to inform future studies on the mechanism of condensins in transcription regulation in C. elegans and other organisms

Analysis of genomic Regions of IncreaseD Gene Expression (RIDGE)s in immune activation

A RIDGE (Region of IncreaseD Gene Expression), as defined by previous studies, is a consecutive set of active genes on a chromosome that span a region around 110 kbp long. This study investigated RIDGE formation by focusing on the well-defined, immunological important MHC locus. Macrophages were assayed for gene expression levels using the Affymetrix MG-U74Av2 chip are were either 1) uninfected, 2) primed with IFN-g, 3) viral activated with mCMV, or 4) both primed and viral activated. Gene expression data from these conditions was studied using data structures and new software developed for the visualisation and handling of structured functional genomic data. Specifically, the data was used to study RIDGE structures and investigate whether physically linked genes were also functionally related, and exhibited co-expression and potentially co-regulation. A greater number of RIDGEs with a greater number of members than expected by chance were found. Observed RIDGEs featured functional associations between RIDGE members (mainly explored via GO, UniProt, and Ingenuity), shared upstream control elements (via PROMO, TRANSFAC, and ClustalW), and similar gene expression profiles. Furthermore RIDGE formation cannot be explained by sequence duplication events alone. When the analysis was extended to the entire mouse genome, it became apparent that known genomic loci (for example the protocadherin loci) were more likely to contain more and longer RIDGEs. RIDGEs outside such loci tended towards single-gene RIDGEs unaffected by the conditions of study. New RIDGEs were also uncovered in the cascading response to IFNg priming and mCMV infection, as found by investigating an extensive time series during the first 12 hours after treatment. Existing RIDGEs were found to be elongated having more members the further the cascade progress

2010 IMSAloquium, Student Investigation Showcase

IMSA students engage in investigations in nanotechnology, particle physics, law, neonatal medicine, literature, transplantation biology, water purity, the educational achievement gap, neurobiology and memory, ethics, theatre, discrete mathematics, economics, and more.https://digitalcommons.imsa.edu/archives_sir/1002/thumbnail.jp

06. 2010 IMSAloquium Student Investigation Showcase

https://digitalcommons.imsa.edu/class_of_2010/1004/thumbnail.jp

The Chittagong story: studies on the ecology of rat floods and bamboo masting

Rodent population outbreaks due to the 50-year cycle of gregarious flowering and seed masting of Melocanna baccifera were first noted in the Chittagong Hill Tracts (CHT) of Bangladesh during the crop production cycle of 2008. The wave of flowering has steadily moved southward through the region each year, with seed masting still occurring in some areas of the CHT during 2010. Because of a lack of surveillance, it is not yet known whether all Melocanna bamboo forests across the region have now initiated flowering. Ecological surveys carried out during the masting event have provided some preliminary evidence that nearly all rodent species are able to exploit Melocanna bamboo seeds as a food resource, with nearly 30% of the seed fallen in forests damaged by rodents. Breeding potential of the predominant species found, Rattus rattus, appears to confirm that aseasonal breeding occurs due to the abundant supply of bamboo seed during masting events. These preliminary results obtained from ongoing research surveys are discussed in the context of the management response to the regional famine triggered by the severe crop damage caused by rodent population outbreaks

Interpersonal deceit and lie-detection using computer-mediated communication

This thesis examines the use of computer-mediated communication for lie-detection and interpersonal deceit. The literature within the fields of lie-detection and mediated communication are reviewed and it is proposed that there is a lack of knowledge surrounding how people use CMC to deceive one another. Qualitative research was carried out in order to address this shortcoming, exploring the self-reported experiences of chat room users who have been exposed to online deceit. Reports were provided that describe the misrepresentation of age, gender, vocation, affection, and appearance. The importance of stereotypes in driving suspicions is also emphasised within the reports. It is suggested that this key characteristic has more dominance in CMC than it would do face-to-face because of the occlusion of the traditional nonstrategic clues to deceit. Evidence for an alternative set of nonstrategic leakage clues was examined further by conducting a variant of the Guilty-Knowledge test within the context of a CMC based crime. It was found that participants exhibited a response time inhibition effect when presented with 'guilty knowledge' and that this effect was detectable through a standard two-button mouse. The use of such nonstrategic cues to deceit was explored further in a study that examined how CMC might be used to add additional control to a Statement Validity Assessment truth-validation test. It was found that the content analysis technique used by SVA was unable in its present form to correctly distinguish between truthful and fabricated statements of participants interviewed using a CMC chat program. In addition, it was found that the deletion-behaviours of participants fabricating a story within CMC provided no quantitative or qualitative evidence that they were lying

Engineering handbook

2004 handbook for the faculty of Engineerin