Guidelines for DNA recombination and repair studies: Cellular assays of DNA repair pathways

Understanding the plasticity of genomes has been greatly aided by assays for recombination, repair and mutagenesis. These assays have been developed in microbial systems that provide the advantages of genetic and molecular reporters that can readily be manipulated. Cellular assays comprise genetic, molecular, and cytological reporters. The assays are powerful tools but each comes with its particular advantages and limitations. Here the most commonly used assays are reviewed, discussed, and presented as the guidelines for future studies

Epigenetic control of neural crest development and evolution

170 pagesEmbryogenesis is characterized by the formation of a complex multicellular organism from a single pluripotent cell. My thesis research has explored this phenomenon through the lens of an embryonic cell population called the neural crest. Neural crest cells are a vertebrate-specific cell type that gives rise to a diverse cell lineage in the developing embryo. The neural crest is postulated to serve as a key driver of vertebrate evolution, contributing to a number of structures in the adult body plan, including the peripheral nervous system, the pigmentation of the skin, and the craniofacial skeleton. Neural crest cell fate commitment requires not only shifts in gene expression, but also an extensive remodeling of the epigenomic landscape. Even so, we still have a superficial understanding of the epigenetic mechanisms which promote neural crest identity. To examine the cis-regulatory landscape of neural crest development, I have coupled both classical embryology approaches and genomic techniques to explore cell state changes underlying neural crest identity. To determine how chromatin states are reorganized during neural crest formation, I examined the function of pioneer factor TFAP2A at discrete stages of development. Through this work, I characterized a mechanism by which TFAP2A remodels the epigenome to allow for progressive cell fate commitment within the neural crest lineage. Next, I aimed to understand the epigenetic basis of neural crest developmental plasticity by performing a cis-regulatory comparison between two neural crest subpopulations, the cranial and trunk neural crest. This allowed me to identify TGF-_ signaling as a potent regulator of cranial neural crest identity. Lastly, I sought to determine how the cis-regulatory landscape is modulated in neural crest cells of different species to give rise to evolutionary novelty. As the craniofacial skeleton is largely derived from the neural crest, I investigated the epigenetic mechanisms driving interspecific variation in beak morphology during avian embryonic development. By performing single-cell ATAC-seq in neural crest cells isolated from three avian species, chicken, quail and duck, I identified conserved cis-regulatory regions displaying species-specific accessibility and discovered a potential role for homeodomain transcription factors in modulating the neural crest epigenetic landscape across species. Together, these studies highlight how the epigenome shapes various aspects of cellular identity throughout development and evolution.2025-01-1

Managing Boundaries: The Role of Narratives at a 9-1-1 Call Center

Dispatchers and calltakers who work at 9-1-1 call centers are confronted with memories of emergencies they must address at work even though they are not physically present at the event. The language they use to talk about their work thus always references a potentially traumatic experience processed second-hand. These telecommunicators use personal messaging through the dispatch platform, verbal communication, and texting in cellphones to tell stories about their work and manage emergency response. Often two to three mediums are used in order to communicate different aspects of the same narrative. Through storytelling, dispatchers manage an environment influenced by social hierarchies, workplace command structures, gender dynamics, and the emotional stress of the calls they must process. The fragmented experiences of dispatchers are reflected in the disjointed methods and narrative structures of their storytelling. This study offers an approach to multi-modal communication and presents an analysis of an occupational folk group not previously studied by folklorists.2015-07-1

Network architecture and regulatory logic in neural crest development

The neural crest is an ectodermal cell population that gives rise to over 30 cell types during vertebrate embryogenesis. These stem cells are formed at the border of the developing central nervous system and undergo extensive migration before differentiating into components of multiple tissues and organs. Neural crest formation and differentiation is a multistep process, as these cells transition through sequential regulatory states before adopting their adult phenotype. Such changes are governed by a complex gene regulatory network (GRN) that integrates environmental and cell‐intrinsic inputs to regulate cell identity. Studies of neural crest cells in a variety of vertebrate models have elucidated the function and regulation of dozens of the molecular players that are part of this network. The neural crest GRN has served as a platform to explore the molecular control of multipotency, cell differentiation, and the evolution of vertebrates. In this review, we employ this genetic program as a stepping‐stone to explore the architecture and the regulatory principles of developmental GRNs. We also discuss how modern genomic approaches can further expand our understanding of genetic networks in this system and others

Cutting Edge: Self-Antigen Controls the Balance between Effector and Regulatory T Cells in Peripheral Tissues

Immune homeostasis in peripheral tissues is achieved by maintaining a balance between pathogenic effector T cells (Teff) and protective Foxp3(+) regulatory T cells (Treg). Using a mouse model of an inducible tissue-antigen we demonstrate that antigen (Ag) persistence is a major determinant of the relative frequencies of Teff and Treg cells. Encounter of transferred naïve CD4(+) T cells with transiently expressed tissue-Ag leads to generation of cytokine-producing Teff cells and peripheral Treg cells. Persistent expression of Ag, a mimic of self Ag, leads to functional inactivation and loss of the Teff cells with preservation of Treg in the target tissue. The inactivation of Teff cells by persistent Ag is associated with reduced ERK phosphorylation (pERK), whereas Treg cells show less reduction in pERK and are relatively resistant to ERK inhibition. Our studies reveal a crucial role for Ag in maintaining appropriate ratios of Ag-specific Teff to Treg cells in tissues