WorldCat Identities

Krasnow, Mark 1956-

Works: 16 works in 16 publications in 1 language and 19 library holdings
Roles: Thesis advisor
Publication Timeline
Publications about  Mark Krasnow Publications about Mark Krasnow
Publications by  Mark Krasnow Publications by Mark Krasnow
Most widely held works by Mark Krasnow
Branching morphogenesis by Mark Krasnow ( Visual )
1 edition published in 2003 in English and held by 2 WorldCat member libraries worldwide
Study of the genetic programs that control development of branched networks of tubes in the Drosophilia
Roles for hedgehog signaling in zebrafish development by John Kenneth Mich ( )
1 edition published in 2010 in English and held by 2 WorldCat member libraries worldwide
The zebrafish has emerged as a powerful model organism for the experimental interrogation of the processes that bring about vertebrate development. In this thesis we study functions of Hedgehog (Hh) signaling, a process that is frequently dysregulated in cancer, that occur during the formation of the zebrafish embryo. First, we investigate a possible link between Hh signaling and the directed migration of germ cells to the presumptive gonad in the early embryo and find that Hh-targeting drugs can perturb germ cell migration, but Hh signaling itself does not regulate this process. Second, we study the formation of motoneurons in the developing ventral spinal cord and find that Hh signaling and retinoic acid signaling work together to induce these cells by the same downstream output of Gli transcription factor activity. Third, we generated transgenic zebrafish that reliably report Hh signaling events at multiple levels of the pathway and we apply these tools in initial studies of zebrafish regeneration. In summary, these studies further our knowledge of the roles for Hh signaling in vertebrate development and will lead insight into the pathology of Hh-related disorders
Depot-specific gene expression programs of adipocytes physiological and developmental implications by Heather Jean Clemons ( )
1 edition published in 2010 in English and held by 2 WorldCat member libraries worldwide
Adipose tissue is found in diverse depots throughout the human body. The diversity of physiological specialization of these fat depots is reflected in the diverse depot-specific alterations seen in lipodystrophies and links between specific patterns of fat distribution and susceptibility to diseases, including Type II Diabetes and coronary artery disease. Previous studies, although focused on only 2 major fat depots (omental and abdominal subcutaneous), have identified numerous differences in metabolic, endocrine and developmental programs. We carried out a more extensive and higher resolution investigation of the anatomic specialization of white adipose tissue, based on 59 samples collected from 7 anatomically diverse fat depots, from 56 individuals. Using DNA microarrays we measured relative abundance of ~25,000 distinct mRNAs in each adipose tissue sample and in adipocytes and stromal-vascular cells isolated from each sample, and compared their gene expression patterns with ~120 samples representing diverse non-adipose tissues. Adipocytes from different regions of the body displayed distinct gene expression profiles. Characteristic patterns of expression of HOX genes distinguished adipocyte samples by site of origin; these patterns were recapitulated when adipocyte precursors from each site were cultured and differentiated ex vivo, suggesting that these genes may have a role in programming and/or maintaining depot-specific differentiation of adipocytes. Expression in adipocytes of 300 genes with major roles in energy metabolism showed both depot-dependent and inter-individual variation. Some of the patterns suggested important differences among anatomic depots in metabolic roles. For example, a set of genes involved in lipid uptake and storage and hormone-regulated lipolysis were generally most highly expressed in breast, pericolonic and omental adipocytes whereas genes involved in glycogen metabolism and de novo fatty acid synthesis were generally most highly expressed in breast and abdominal subcutaneous adipocytes, suggesting that these depots play a role as glucose buffers. Dozens of genes known or predicted to encode secreted molecular signals were highly expressed in adipocytes compared to either adipose stromal-vascular cells or other organs; many of these were differentially expressed among adipocyte depots. Some of these genes appear to be good candidates for encoding novel adipokines. In summary, we found extensive variation among adipose depots, as well as inter-individual variation, in global gene expression. Differences in genes linked to development, metabolism and cell-cell communication suggest new hypotheses relevant to depot-specific and general aspects of adipocyte biology
The ciliated cell transcriptome by Ramona Amy Hoh ( )
1 edition published in 2010 in English and held by 2 WorldCat member libraries worldwide
Multiciliated cells of the respiratory epithelium are unique in that they generate hundreds of modified centrioles called basal bodies per cell. Each basal body anchors a motile cilium at the cell apical surface, and coordinated beating of motile cilia is vital for protecting from airway infection and for respiratory function. We used mice expressing GFP from the promoter of a ciliated cell-specific gene, FOXJ1, to obtain sorted populations of ciliating cells for transcriptional analysis. In addition to successfully identifying candidates found in other proteomics and genomics studies of motile and nonmotile cilia, approximately half of the significantly upregulated genes identified here have not yet been linked to cilia, and of those a third of are currently uncharacterized. We identified several genes associated with human diseases. These include FTO, which has been linked to human obesity, and DYX1C1, which is a candidate gene for developmental dyslexia. Interestingly, FTO localizes to cilia and Dyx1c1-GFP localizes to cilia and centrosomes, establishing novel links between cilia and two genetic diseases with poorly understood cellular and molecular etiology. Finally, we identified a number of transcription factors that are differentially expressed in ciliating mouse tracheal epithelial cells, including the proto-oncogene c-myb. We show that C-myb is expressed specifically in ciliating cells, and that this expression is temporally restricted to early in the differentiation process. These results suggest a role for the leukemogenic transcription factor C-myb in ciliated cell differentiation
Catalytic promiscuity in the alkaline phosphatase superfamily identifying specificity determinants in the diesterase family leads to a hypothesis of a "generalist" core bimetallo site found throughout the superfamily by Helen Irene Wiersma-Koch ( )
1 edition published in 2013 in English and held by 1 WorldCat member library worldwide
Catalytic promiscuity, the property of enzymes possessing low levels of activity toward non-cognate reactions, can be exploited as a functional tool to investigate conserved and non-conserved mechanism of enzyme specificity and catalysis between members in the same superfamily. Members of the main branch of the Alkaline Phosphatase (AP) superfamily have a structurally conserved core and active site bimetallo site. Other active site features that confer specificity for a given reaction differ between the families. Families within this superfamily catalyze a wide range of reactions, and enzymes within different families show catalytic promiscuity toward reactions catalyzed by other families within this branch. Structural comparisons and phylogenetic analysis suggests that all of the families in the superfamily arose from a common ancestor whose active site consisted of the bimetallo site alone, absent of peripheral, specificity determining features. Experimental data with a member of the nucleotide pyrophosphatase/phosphodiesterase (NPP) family suggests that a "minimal" mutant version of this enzyme that lacks peripheral, specificity determining features, has equal activity toward the two major reactions catalyzed by the AP superfamily, phosphate monoester and phosphate diester hydrolysis reactions. Together, the structural comparisons, phylogenic analysis, and experimental results lead to the hypothesis that the common ancestor of the AP superfamily is a "generalist." By using a variety of techniques, we provide support for a mechanism of evolution in which a "generalist" enzyme may give rise to multiple enzymes specific for, related, but individual reactions. Support for this mechanism, first proposed in 1976, has, until, now been limited. We suggest that this "generalist" mechanism is a valid mechanism for the evolution of ancient enzymes, present in the early evolution of life, into diverse superfamilies in which each family possesses specificity for one given reaction
Characterization of novel RNA-protein regulatory interactions in Saccharomyces cerevisiae by Nikoleta Georgieva Tsvetanova ( )
1 edition published in 2012 in English and held by 1 WorldCat member library worldwide
The dynamic processes of a living cell depend on the coordinated temporal and spatial regulation of the many steps of gene expression. Transcription regulation is one control point of gene expression, and a gene can also be regulated post-transcriptionally, by RNA-binding proteins (RBPs). The biological significance of post-transcriptional regulation is especially evident in cases, where RBP binding controls the temporal precision of suppression and activation of important cellular stress responses. We developed a proteome-wide experimental approach for in vitro identification of novel RBPs and RNA-protein interactions in Saccharomyces cerevisiae. We found 12 novel RNA-binding proteins, the majority of which, surprisingly, are currently annotated as enzymes with roles in metabolic processes. We next used this proteomic approach to screen for proteins specifically interacting with the HAC1 RNA, which mediates activation of the yeast unfolded protein response (UPR). We found that HAC1 associated reproducibly with four small yeast GTPases, three of which are of the Ypt family of ras-GTPases. We further characterized one of them, the yeast Rab1 homolog Ypt1, and showed that Ypt1 interacted with unspliced HAC1 RNA only in the absence of ER stress. Selective Ypt1 depletion increased HAC1 RNA stability and expression, and also affected timely recovery from UPR. By developing and applying a novel proteomic approach for studying RNA-protein interactions, we established Ypt1 as an important regulator of HAC1 expression and UPR signaling. This unexpected protein-RNA interaction provides a biochemical mechanism for coordinating the key cellular processes of vesicle trafficking and ER homeostasis
C-myb controls the initiation of ciliogenesis in the developing mouse airway epithelium by Fraser Elisabeth Tan ( )
1 edition published in 2011 in English and held by 1 WorldCat member library worldwide
In the mammalian lung, multiciliated cells lining the conducting airways play a crucial role in mucociliary clearance, the primary defense mechanism of the lung against foreign particles and bacterial infection. The cilia of the multiciliated cells beat in metachronic waves to propel mucus up from the deep distal branches of the lung and out of the trachea, to be either expectorated or swallowed. Breakdown of the mucociliary escalator, due to ciliary dysfunction as occurs in primary ciliary dyskinesia, or due to changes in osmotic balance, as occurs in cystic fibrosis, result in chronic airway bacterial infections. Viral infection in healthy individuals also causes loss of cilia, and this correlates with high rates of bacterial infections that follow many viral infections. Despite the importance of these cells, we know very little about how they are specified during development, nor about how multiple cilia are generated. Each cilium is a membrane encased microtubule based structure that protrudes from the apical surface of the cell. Most cells bear a solitary, or primary, cilium, while only a few specialized cell types can generate hundreds of cilia each. Each cilia is nucleated by a single centriole, and centriole number is normally tightly regulated. How these multiciliated cells circumvent this regulation to generate hundreds of centrioles is unknown. One the developing multiciliated cell has generated its centrioles, those centrioles migrate to and dock with the apical membrane and generate their ciliary axonemes. While many proteins involved in ciliary function and intraflagellar transport have been identified, little is known about how these process are controlled from a molecular level. Very few transcription factors are known to play a role in ciliogenesis. The most well-studied one is Foxj1, which is required for motility in single cilia, and is also critical for proper centriole migration in multiciliated cells. However, ectopic expression of Foxj1 cannot confer a ciliated fate on a non-ciliated cell, and cells null for foxj1 still generate centrioles, so there must be other transcription factors that initiate ciliogenesis and act in the early steps of the process, such as centriole generation. Here, we have identified the first transcription factor that acts upstream of Foxj1 during ciliogenesis. We screened through a subset of the known and predicted transcription factors in the mouse genome for those expressed in the developing lung in a pattern similar to that of Foxj1. We isolated a single candidate, the myeloblastosis oncogene (c-myb). We showed that c-Myb was expressed in post-mitotic cells in the developing lung epithelium, many of which co-expressed Foxj1, indicating that c-Myb is expressed in ciliating cells. Analysis of the timing of c-Myb expression during development as compared to Foxj1 and to maturing ciliary axonemes revealed that c-Myb was expressed during the early steps of ciliogenesis and was downregulated as the cells matured. Specific removal of c-Myb from the developing airway epithelium abrogated Foxj1 expression and centriole generation at E15.5. c-myb null epithelia did not have mature ciliated cells at E17.5, although Foxj1 expression and centriole generation appear to have recovered by this time point. This is the first identification of a transcription factor that regulates the initiation of ciliogenesis in the developing mouse lung. Finally, we propose a modular model of transcriptional control of cilia, in which cells deploy specific transcriptional modules to build the particular kind of cilia they require
Genetic probing of the origin of alveolar myofibroblasts and bronchopulmonary dysplasia, a disease of alveolar development by Krystal Renee St. Julien ( )
1 edition published in 2013 in English and held by 1 WorldCat member library worldwide
Alveolarization, the final stage of lung development, promotes gas-exchange by dividing the sac-like terminal airspaces of the lung into thin-walled pockets. Myofibroblasts are found at sites of septation, and mouse mutants in PDGFR alpha signaling lack myofibroblasts and air sacs fail to septate, suggesting that myofibroblasts play a role in septation. Further, a similar arrest in alveolarization is observed in diseases like bronchopulmonary dysplasia (BPD), a disease of premature infants that twin studies implicate may have important genetic contributions. In this thesis, I explore the developmental origins of myofibroblasts in mouse and the genetic basis of BPD. I first describe a novel protocol developed to extract DNA from dried blood spots taken from newborn infants that can be used in genotyping and genome-wide association studies (GWAS). I then describe an agnostic exploration of the genetic basis of BPD. Finally, I describe the use of genetic lineage tracing in mice to identify airway smooth muscle cells as a source of ductal myofibroblasts during lung development
Dissecting the developmental origins of organ-specific vascular beds by Patrick Evan Bogard ( )
1 edition published in 2012 in English and held by 1 WorldCat member library worldwide
One of the challenges facing modern medicine is to understand mammalian organ formation. Each organ has a specialized function and form, yet they are built from similar parts. One important part of each organ is the circulatory system, composed of arteries, veins, and capillaries, which is responsible for delivery of oxygen and nutrients and for the removal of carbon dioxide and metabolic waste. For centuries scientists have been trying to understand how the circulatory system functions and more recently how it forms. In this thesis, I focus on the development of the mouse pulmonary vasculature, the circulatory system that is responsible for transporting blood from the heart to the lungs where oxygenation occurs, and back to the heart where the oxygenated blood is pumped throughout the body. The lung is a complex organ made up of multiple hollow airway tubes that must form in coordination so that proper gas exchange can occur. In the first chapter I review the function and development of the circulatory system. In chapter two I present the experiments completed to determine the cellular origins of the blood vessels of the mouse lung and heart. Currently there are two major models for how the pulmonary arteries of the mouse form, vasculogenesis of lung mesenchymal endothelial progenitors and sprouting angiogenesis from vessels outside the lung. Using fate mapping I directly test the dominant model, vasculogenesis, and show that there are no endothelial progenitor cells resident in the lung mesenchyme. To determine if a vessel outside the lung is forming the pulmonary arteries I performed a clonal analysis labeling single endothelial cells before pulmonary artery formation and followed the fates of their daughter cells. I found a surprising multipotent progenitor population of endothelial cells, the primitive plexus of the lung, that gives rise to pulmonary arterial, venous, and plexus endothelial cells. I showed that the transition from plexus to arterial fate occurs in a leakproof manner in the lung by performing intracardiac perfusion experiments of fluorescently labeled dextrans and lectins. I also demonstrated a similar mechanism of leakproof plexus remodeling occurs in the heart during the development of the coronary arteries and veins. In chapter three I discuss experiments done to understand the molecular mechanisms controlling pulmonary artery formation. I describe an in silico in situ hybridization screen I performed to identify candidate molecules. I describe the expression pattern of BMP pathway activity in the lung as being restricted to the pulmonary arteries, a subset of the plexus and budding airways, using a transgenic mouse line that is only expressed in cells actively undergoing BMP signaling. I analyzed mice homozygously deficient for the BMP antagonists Gremlin and BMPER, and showed that there is no abnormal pulmonary artery formation or patterning phenotype. I also describe experiments in which I have deleted the BMP Receptor 2 from endothelial cells and again there is no abnormal pulmonary artery formation or patterning phenotype. I describe the expression pattern of VEGF in the lung as being mesenchymal during early lung formation, and show that mesenchymal deletions of VEGF do not result in abnormal pulmonary artery formation or patterning phenotypes. I found that multiple single gene manipulations do not result in abnormal phenotypes suggesting that pulmonary artery formation and patterning is robust and likely involves the interaction of multiple signaling pathways. This work describes a common mechanism of organ specific vascular bed formation that has not been previously described at a cellular and molecular level. I hope these studies provide a foundation for understanding how the vascular system of many organs form during organogenesis
Molecular and cellular mechanisms of tracheal invasion of polarized muscle membrane networks in Drosophila by Soren Joseph Peterson ( )
1 edition published in 2011 in English and held by 1 WorldCat member library worldwide
From a structural standpoint, one of the most characteristic general design elements of the mammalian organism is its tubular nature. The lung and circulatory system shuttle oxygen and nutrients to target tissues and allow for the excretion of waste products. In some ways, the vasculature lies at the center of human physiology--its passageways provide the infrastructure for maintaining homeostasis. Despite the importance of tubular networks in human health and disease, we have a poor understanding of many aspects of the genetic, molecular, and cellular programs controlling the development of these complex structures. The Drosophila melanogaster tracheal system, an elaborate network of hollow epithelial tubes, transports gases to and from target tissues. The tracheal system, with its simple structure, tractable genetics, and substantial experimental toolkit has emerged as an excellent model system for studying questions with relevance to more complex tubular systems. During development, tracheal branches ramify on the surface of target tissues, providing oxygen to every cell in the body. However, in a phenomenon unique to the Drosophila flight muscle, trachea are also present within plasma membrane invaginations deep below the muscle's outer extremities and have been described to surround every mitochondria of the flight muscle, thus coupling oxygen delivery directly to aerobic respiration at the mitochondrion. Although the presence of trachea within flight muscle membrane invaginations has been described for over 150 years, the developmental progression and cellular and molecular basis of this subcellular targeting process is unknown. In Chapter 2, we show that tracheal branches invade the developing flight muscle Transverse (T)-tubule plasma membrane invagination system during a brief period of pupal development. Branchless (Bnl) FGF, a fibroblast growth factor that functions as a chemoattractant, is required in the flight muscle, and its cognate receptor, Breathless (Btl) FGFR, is required in trachea for the tracheal invasion process to occur. Whereas Bnl FGF is localized to all flight muscle plasma membranes prior to tracheal invasion, during invasion Bnl FGF localizes preferentially to the T-tubule and is excluded from the surrounding plasma membrane. In addition to Bnl FGF, core polarity regulators commonly found on basolateral membranes in epithelial cells also preferentially localize to the T-tubule network during tracheal invasion. We find that depletion of AP-1[gamma], targeting machinery required for basolateral secretion in Drosophila epithelia, can also reroute Bnl FGF secretion to the outer plasma membrane and away from T-tubule openings, shifting trachea to the plasma membrane and away from T-tubules. We propose that (1) polarized secretion of Bnl FGF to the T-tubule guides tracheal branches into the T-tubule network and that (2) polarized Bnl FGF secretion is established via the redeployment of ancestral basolateral secretion pathways to the T-tubules, a membrane domain having molecular signatures of epithelial basolateral domains. To our knowledge, compartmentalized secretion of Bnl FGF to flight muscle T-tubule membranes is the first example of polarized subcellular secretion of a growth factor with functional consequences for the development of another tissue. In Chapter 3, we examine the molecular machinery involved in maintaining the polarized secretion of Bnl FGF during tracheal invasion. We find a host of secretory machinery, including several Rabs, Myosin V, an actin nucleator, and others, to be involved in the secretion of Bnl-FGF to the T-tubule during tracheal invasion. From these data we propose a molecular model to explain polarized Bnl FGF secretion at the T-tubule. Why is the flight muscle the only tissue invaded by trachea? In Chapter 4, we find that the flight muscle is structurally adapted to allow its T-tubule plasma membrane invagination network to be co-opted by migrating tracheal processes. In contrast to other muscles, the flight muscle T-tubule network forms large holes on the muscle surface as part of its development. The tracheal invasion process in the flight muscle is therefore the consequence not only of a polarized secretion process as discussed in Chapter 2, but also a structurally distinct stage of flight muscle development. In the final chapter, we search for the fine tracheal tubes reported to target and form contacts on the flight muscle mitochondria. We find an extracellular protein-based lattice of the appropriate diameter and localization to potentially represent the reported tracheal extensions to the mitochondria and propose several models of protein lattice formation. We hope that insight derived from this work spurs future investigators in a host of areas. The exquisite example of targeting to specific membrane domains dependent on a subcellular chemoattractant gradient demonstrated by the invading tracheal branch may provide insight into a number of developmental scenarios where fine targeting of different cells is required. We also hope that our finding regarding polarized secretion in muscle inspires new ways to think about the assembly of muscle membranes during development and disease states
Tracheal progenitor outgrowth from the niche during Drosophila melanogaster metamorphosis by Feng Chen ( )
1 edition published in 2013 in English and held by 1 WorldCat member library worldwide
While there has been great progress in identifying stem and progenitor cells and the signals that control their proliferation and differentiation, how stem/progenitor cells exit their niche and how they form new tissue is not well understood. Unlike in the embryo, tissue formation in an adult animal faces new challenges such as longer distances to migrate and a complex milieu of differentiated tissue to migrate around and/or coordinate with. To restore function to a tissue, stem/progenitor cells must also integrate into healthy tissue and coordinate growth with decaying tissue. In this thesis, I examine how tracheal progenitor cells exit their niche to form tracheal tissue during Drosophila melanogaster metamorphosis. Tracheal progenitors exit the niche in two waves. During the first wave, progenitors migrate onto the basal surface of larval tracheal branches destined for destruction, and track along the decaying branches. Progenitor outgrowth requires the embryonic tracheal branch inducer, Breathless FGFR, and surprisingly, the Branchless FGF ligand is expressed in larval branches along which progenitors crawl. In this way, outgrowth is coordinated with tissue decay. Progenitors that remain within the niche during the first wave exit later. However, instead of moving onto the basal surface of larval branches, progenitors exiting the niche during this second wave move along the apical surface, displacing larval cells, and repopulating the tracheal branch. This latter process does not resemble branching morphogenesis in the embryo and does not require Bnl/Btl FGF signaling, demonstrating that progenitor outgrowth does not always require embryonic guidance cues
Studying the functions of Drosophila myosin VI through identification of multiple cargo-binding proteins by Megan Amanda Hartman ( )
1 edition published in 2011 in English and held by 1 WorldCat member library worldwide
Molecular motors utilize energy from ATP hydrolysis to perform mechanical work, and examples include the myosins, a superfamily of proteins that utilizes actin filaments as a track for processive movement. There are many classes of myosins, and they share common elements necessary to regulate actin binding and force generation based on the identity of the nucleotide bound. Their diversity is most evident in their C-terminal tails; in the case of the unconventional myosins that do not form bipolar thick filaments, these regions mediate their associations with specific binding partners. Interactions between myosins and cognate adaptors allow for their recruitment onto cargoes, and it is these cargoes that define the functions of myosins in a cellular environment. Numerous studies have suggested developmental and cellular roles for myosins, but in general, few binding proteins have been discovered for most family members. In the case of mammalian myosin VI, yeast two-hybrid screening has revealed several adaptors that link this motor to vesicles, endosomes, and the Golgi, where it participates in a number of trafficking events. In contrast, very little is known about the proteins that bind to Drosophila myosin VI, despite the necessity of this protein for many essential processes during fly development. To better understand the specific pathways to which myosin VI contributes in flies, we set out to identify its cargoes. We used a combination of affinity chromatography and mass spectrometry in a proteomics-based screen to discover candidates, given the many advantages of this method over yeast two-hybrid and other approaches. Upon obtaining data indicating that over 1000 proteins could potentially associate with myosin VI, we were next charged with determining which interactions were specific and direct, and we chose to screen through select candidates with an in vitro assay. We identified a number of novel cargoes for myosin VI, including those associated with the Golgi, protein trafficking, microtubules, and others of unknown function. Next, we attempted to perform another screen to identify binding partners that might mediate the function of myosin VI in dorsal closure during embryogenesis. Because this motor is very concentrated at the leading edge of cells during this process, we developed an antibody to myosin VI for immunolocalization studies. We then examined embryos mutant for or depleted of myosin VI cargoes and assessed any effect on myosin VI localization. Although some showed slight differences in the staining pattern, we chose a different assay to test for a shared function between myosin VI and one of its novel cargoes. After commencing this project, we became aware of data indicating that myosin VI and Cornetto, one of the novel binding proteins we discovered, participate in Hedgehog secretion. Without further information about the assay used or the results obtained, we used cell culture to validate the original screen data and found that both proteins are indeed required for the timely secretion of exogenously expressed Hedgehog (Hh) from S2R+ cells. We then began examining embryos for defects associated with reduced Hedgehog secretion and found that a small percentage of flies expressing Cornetto RNAi or a dominant negative myosin VI truncation in Hh-producing cells indeed have the types of segmentation problems found in mild hedgehog mutants. Thus, we found a shared function for myosin VI and one of its binding proteins, which not only validates our screen data but also provides important information not previously available about their roles in development. In a second project, I turned my attention to another function for myosin VI to analyze its binding protein CG3529, which is orthologous to Tom1 family proteins involved in membrane trafficking. Based on information available about orthologs, I reasoned that CG3529 might mediate the involvement of myosin VI in asymmetric Notch signaling during pupal development, a role identified for this motor in another large-scale RNAi screen. Despite finding no evidence linking CG3529 to asymmetric trafficking of Notch in the dorsal epidermis as I had hypothesized, it was apparent that CG3529 depletion does affect mechanosensory bristle length, which had been previously noted for myosin VI as well. I then performed experiments to help determine by what pathway these proteins contributed to this process, but the data I collected did not indicate a shared function or binding interactions between CG3529 and the proteins to which its orthologs had been linked. Although it is still not clear what the function of this myosin VI adaptor is, I suspect that it is involved in protein trafficking at or near the endoplasmic reticulum, given that CG3529 appears to localize to this compartment. Beyond the proteins whose functions we were able to address, we obtained many other candidates that could potentially link myosin VI to a variety of cellular compartments and signaling pathways. The data presented here should be useful in future work to analyze the roles of myosin VI in specific systems upon utilization of information available about its novel binding proteins
Hedgehog signaling in urogenital regeneration and neoplasia by Yeok Loo Agnes Lim ( )
1 edition published in 2014 in English and held by 1 WorldCat member library worldwide
The Hedgehog (Hh) signaling pathway functions in embryonic development to control cell growth, cell fate and pattern many aspects of the vertebrate body plan. The role of the pathway in adult tissue homeostasis, regeneration and cancer, however, is still emerging, especially in the context of epithelial/stromal interactions. Previous work on the adult bladder demonstrated the importance of stromal Hh pathway response in mediating homeostatic response to injury by eliciting production of stromal factors that act on the epithelium; in this dissertation I investigate the role of epithelial/stromal interactions involving the Hh pathway in prostate regeneration and bladder cancer. In the adult prostate, Hh pathway response occurs in a subset of stromal cells. Prostate regeneration involves the formation of new prostate branches, and I showed that stromal cells at the tips of nascent buds lacked Hh pathway response. Decreased pathway activity in stromal cells leads to increased branching, and this correlates with increased secretion of hepatocyte growth factor (Hgf), which acts on the epithelium to induce prostate branching. In regions that are not budding, stromal Hh pathway response down-regulates expression of Hgf. Hence, the spatial organization of stromal Hh pathway response in combination with the negative regulation of secreted hepatocyte growth factor (Hgf) levels by Hh pathway activity, together modulate branching morphogenesis during prostate regeneration. I also examined the role of Hh pathway activity in development of invasive urothelial carcinoma and find, despite its initial presence in the cancer cell of origin, that Sonic hedgehog (Shh) expression is lost during progression. Genetic blockade of stromal response to Shh furthermore dramatically accelerates progression. This cancer-protective effect is associated with Shh-induced stromal expression of BMP signals, which stimulate urothelial differentiation, and progression is reduced by pharmacological activation of BMP pathway activity. Taken together, the results of this dissertation highlight the importance, in regeneration and cancer, of epithelial/stromal interactions in which the Hh pathway is a key mediator. In both models examined, Hh ligand produced by the epithelium stiumates pathway response in the stroma, which regulates the secretion of factors that act on the epithelium. In a broader context, this work provides a conceptual framework for understanding epithelial/stromal interactions involving Hh signaling in other organ systems, a more detailed understanding of which will enable better therapeutic strategies for the treatment of disease
Schwann cell migration and myelination in zebrafish peripheral nerves by Julie Rebecca Perlin ( )
1 edition published in 2012 in English and held by 1 WorldCat member library worldwide
The nervous system connects cells throughout the body to coordinate actions and transmit signals. Critical to the nervous system are the neurons that extend axons to their targets and the glial cells that interact with these axons. Oligodendrocytes in the central nervous system and Schwann cells in the peripheral nervous system wrap axons to make the myelin sheath. Myelin is critical for enhancing the speed of action potentials as well as providing support to axons. Myelin was a critical adaptation that has allowed vertebrates to increase in size, precision of movement, and complexity. Insults to myelin cause devastating diseases, and a better understanding of the normal development of myelinating glia is necessary for improving treatment of human neuropathies. In this dissertation I investigate both well-known and novel signaling pathways and their roles in Schwann cell migration and myelination in zebrafish peripheral nerves. Before making myelin, Schwann cells must migrate along and cover the surface of a bundle of axons. Here, I demonstrate that Neuregulin 1 type III (Nrg 1 type III) is required in neurons to signal through ErbB receptors in Schwann cells for Schwann cell migration along the posterior lateral line nerve, a mechanosensory nerve. Further, ectopic expression of this signal in all neurons is sufficient to attract peripheral Schwann cells into the spinal cord. There appears to be distinct regulation of Schwann cell migration in different nerves of the peripheral nervous system, as migration of Schwann cells in motor nerves requires ErbB receptors but not the Nrg 1 type III ligand. Nrg1 type III does, however, control myelination in both sensory and motor nerves. Schwann cell migration is not only important to properly localize Schwann cells, but also to localize the lateral line nerve itself, which begins in the epidermis but then transitions across a basement membrane to the subepidermal space. As they migrate, Schwann cells degrade the basement membrane beneath the skin, which allows the nerve to transition out of the epidermis, and then rebuild the basement membrane after the nerve has been repositioned and protected from the disorganization that otherwise would take place if it remained in the epidermis. Finally, through analysis of a mutation that was found in a forward genetic screen for genes essential in myelination, I also identify a novel regulator of Schwann cell myelination. Together this work elucidates new roles of known genes in Schwann cell and nerve development and also identifies a novel gene required for peripheral myelination
Understanding the molecular mechanisms of cardiomyopathy-causing mutations in sarcomeric proteins by Ruth Sommese ( )
1 edition published in 2013 in English and held by 1 WorldCat member library worldwide
Heart disease is one of the leading causes of morbidity and mortality in the developed world. As such, significant amount of resources and energy are funneled into developing treatments and therapeutics. While genetically caused cardiovascular diseases manifest at the tissue level, the fundamental mechanism that triggers the secondary effects and tissue remodeling generally occurs at the protein level. It is therefore critical to understand the molecular changes on the basic biochemical and biophysical level and connect these to the cellular and developmental disease processes. One such cardiovascular disorder is hypertrophic cardiomyopathy or HCM, which has been estimated to affect approximately 1 in 500 individuals. It disproportionately affects young adults and is the leading cause of sudden cardiac death. Since the first case of genetically linked HCM was identified in the early 1990s, a significant amount of work has been done to understand the molecular mechanism of the disease. Mutations have been identified in the proteins comprising the contractile apparatus of the muscle, the sarcomere. One such protein is [beta]-cardiac myosin, which has been recognized as a significant culprit of genetically linked HCM (~30-50%). Many studies have been performed to understand how single point mutations can alter the enzymatic and mechanical properties of this motor protein, but there is no clear consensus about the molecular mechanism. This has been due mainly to the lack of available human [beta]-cardiac myosin and the use of non-human and non-[beta]-cardiac myosin. Non-human and non-[beta]-cardiac myosin from common animal models differ by >30 residues, and as clearly evident from the disease and from previous studies on myosin, a single amino acids mutation can significantly alter and disrupt myosin function. During my thesis work, I have performed the first biochemical and biophysical work in the field looking at cardiomyopathy mutations using human [beta]-cardiac myosin. I have shown that HCM-causing [beta]-cardiac myosin mutations result in a gain of function, increasing the power or work output of the myosin. I have also examined HCM-causing mutations in troponin T, a component of the thin filament regulatory unit, using human [beta]-cardiac myosin. In the muscle, the interaction of myosin and actin is regulated by calcium through the thin filament proteins, namely the troponin complex and tropomyosin. HCM-causing mutations in troponin T increase calcium sensitivity or the number of force-producing heads that can interact with actin, thereby also increasing force or power output in the muscle. My work not only sheds light on fundamental properties of thick and thin filament function in the human sarcomere and presents the first studies of HCM-causing mutants in a human background, it also establishes a new approach to the problem of cardiomyopathy that will be critical in truly understanding and targeting the disease
Alveolar stem cells in lung development, maintenance, and cancer by Mark Krasnow ( Visual )
1 edition published in 2012 in English and held by 0 WorldCat member libraries worldwide
(CIT): NIH CRM/SCIG Stem Cell Seminar Series (Extramural) presents Mark Krasnow. Mr. Krasnow is elucidating the genetic programs that control embryonic development of the Drosophila respiratory system and mammalian lung. He is especially interested in the developmental patterning mechanisms and the cell and molecular biology of these processes, and in using the information to understand lung disease and to regenerate a lung
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