2011 — 2016 |
Larschan, Erica Nicole |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Establishing Coordinate Gene Regulation During Drosophila Dosage Compensation
DESCRIPTION (provided by applicant): All organisms must regulate their genes precisely for normal development and to prevent disease states. However, significant gene copy number variation exists across genomes 1;therefore, coordinate regulation is required to equalize transcription levels 2 3. Our long-term goal is to describe the molecular mechanisms used to target genes for coordinate regulation, the essential initial step in their regulation. Dosage compensation is one of the best model systems for studying this process because all of the genes on a single chromosome are specifically identified and co-regulated. Drosophila, like mammals 4, increase the transcript levels of a large number of diversely regulated genes along the length of the single male X-chromosome precisely two-fold relative to each female X-chromosome 5. The objective of this application is to understand how dosage compensation in Drosophila is established, the critical first step in the regulatory process. The Drosophila Male Specific Lethal (MSL) complex is central to dosage compensation;it first identifies the X chromosome using a combination of cis-acting DNA sequences 6 and co-transcriptional recruitment by its roX (RNA on X) non-coding RNA components7 and then spreads into the bodies of active genes 8. However, we do not know how the MSL complex specifically identifies the MSL Recognition Element (MRE) sequences on the male X because known MSL components are insufficient for direct recognition of MREs in vitro 9. We used an innovative genetic screen for new regulators of dosage compensation that function in both males and females and thereby identified the essential CLAMP zinc-finger protein. Guided by strong preliminary data, we propose the following novel mechanism for identifying genes for coordinate regulation: CLAMP and the MSL complex associate inter-dependently, thereby generating a positive feedback amplification system that creates X- specificity from a two-fold X-enrichment of MRE sequences. The rationale for this work is that determining how the MSL complex specifically targets the X-chromosome will yield key insight into how genes are identified for coordinately regulation within sub-nuclear domains. We will test our novel mechanism using three specific aims: 1) We will define DNA sequence requirements for CLAMP binding in vivo and in vitro;2) We will identify CLAMP interacting proteins that mediate its interaction with the MSL complex. At the same time, we will define new interaction partners of a previously unstudied essential transcriptional regulator;3) We will establish the mechanism by which CLAMP and the MSL complex function inter-dependently at high affinity sites. Our proposed research is significant because we expect to describe for the first time the previously unknown mechanism that allows MSL complex to identify its high affinity sites, thereby defining the critical first step in establishing coordinate gene regulation. Defining the novel mechanism by which CLAMP and the MSL complex function together to generate a domain of enhanced transcription is likely to provide key insight into how genes are identified for coordinate regulation across species. PUBLIC HEALTH RELEVANCE: Our proposed research is relevant to public health because loss of precise regulation of genes underlies a large number of diseases including autism 10 and Chron's disease 11. Our research addresses the critical initial step in the coordinate regulation of genes, identifying target genes for subsequent regulation. Our findings are therefore relevant to the part of NIH's mission that pertains to developing fundamental knowledge that will provide tools for mitigating diseases.
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1 |
2011 |
Larschan, Erica Nicole |
P20Activity Code Description: To support planning for new programs, expansion or modification of existing resources, and feasibility studies to explore various approaches to the development of interdisciplinary programs that offer potential solutions to problems of special significance to the mission of the NIH. These exploratory studies may lead to specialized or comprehensive centers. |
Establishing Sub-Nuclear Domains of Coordinate Gene Regulation @ University of Rhode Island
This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Coordinate gene regulation is a fundamental biological process essential to all cells from the germ line to the immune system. Regulation is disrupted in disorders as diverse as autism, Crohn's disease, and Down's syndrome. X-chromosome dosage compensation is one of the best models for studying this process because thousands of genes are co-regulated. Drosophila, like mammals, increase the transcript levels from a large number of diverse genes along the single male X-chromosome precisely two-fold relative to each female X-chromosome and all other chromosomes. The Drosophila Male Specific Lethal (MSL) complex is central to dosage compensation. It has long been proposed that MSL complex first identifies high affinity sites on the male X-chromosome and then spreads along its length. However, there is no temporal data to support this hypothesis due to limitations in obtaining sufficient material for biochemistry. Therefore, we will develop an innovative cell induction system to generate sufficient material to perform chromatin immunoprecipitation followed by NextGen sequencing (ChIPseq) at many time points. We expect that this pilot project will generate the first system to define how sub-nuclear domains are formed in real-time across species and provide a strong foundation for an NIH R01.
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0.966 |
2018 — 2021 |
Larschan, Erica Nicole |
R35Activity Code Description: To provide long term support to an experienced investigator with an outstanding record of research productivity. This support is intended to encourage investigators to embark on long-term projects of unusual potential. |
Establishment of Active Chromatin Domains
Chromatin domains form within the nucleus during early development to precisely co-regulate nearby genes during the maternal to zygotic transition. Prior to zygotic genome activation, the entire genome is very open and only a few key early transcription factors are bound. Yet, many essential active and repressive chromatin domains are present right after zygotic genome activation. Little is understood about how embryos rapidly establish the critical active chromatin domains that are essential for the function of the zygotic genome. Here are several key questions about active chromatin domains that we will address in this proposal: 1) How are different effector complexes specifically targeted to active chromatin domains by similar cis-elements such that the proper domain forms at the correct genomic location? 2) How does competition between transcription factors that recognize similar cis-elements regulate the formation of active chromatin domains during development? 3) How is the three-dimensional architecture of active chromatin domains established? There are many essential chromatin domains that mediate coordinate gene activation throughout the genome including the rDNA locus, the histone locus and the dosage compensated male X-chromosome, all of which must be properly activated in the developing embryo. My research program has initially focused on two of these essential chromatin domains of coordinate gene activation in Drosophila: 1) the dosage compensated X-chromosome that balances gene dosage between sexes and 2) the histone locus body (HLB) that coordinately regulates histone gene expression. Drosophila is an ideal organism with which to study the formation of chromatin domains early in development due to their rapid and synchronized early development and the large number of genetic and biochemical tools and genomic data sets available. While many cis and trans acting factors that regulate chromatin domains have been identified, little is known about the molecular mechanisms that drive their formation at specific genomic loci during early development. Defining how the chromatin domains on the active male X-chromosome and the HLB are established and maintained over developmental time will reveal key principles by which active chromatin domains form. Using steady-state measurements, we have recently discovered that a single transcription factor, CLAMP, regulates formation of both the dosage compensated X-chromosome and the HLB active chromatin domains, providing us an entry point for revealing new insights into the dynamic process of chromatin domain formation. The significance of our work is that we will define how active chromatin domains form over developmental time and will seek to identify common underlying mechanisms that drive formation of two very different domains at specific genomic loci.
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1 |
2020 — 2021 |
Johnson, Mark Aikens (co-PI) [⬀] Johnson, Mark Aikens (co-PI) [⬀] Johnson, Mark Aikens (co-PI) [⬀] Johnson, Mark Aikens (co-PI) [⬀] Johnson, Mark Aikens (co-PI) [⬀] Johnson, Mark Aikens (co-PI) [⬀] Johnson, Mark Aikens (co-PI) [⬀] Larschan, Erica Nicole Mowry, Kimberly L. [⬀] |
T32Activity Code Description: To enable institutions to make National Research Service Awards to individuals selected by them for predoctoral and postdoctoral research training in specified shortage areas. |
Interdisciplinary and Inclusive Predoctoral Training in Molecular, Cellular, and Biochemical Sciences
PROJECT SUMMARY Solving the complex problems in human health and modern biology represents a major challenge for those who will lead biomedical research in the near and long-term future. The core disciplines of our training program?molecular biology, cell biology, and biochemistry?have led the way in development of innovations that are driving life sciences research and applications today. It is imperative that US life scientist training programs evolve to meet the demand for a diverse group of leaders who are trained in rigorous and transparent implementation and reporting of quantitative analysis of biological data. We recognize that this demand will require a change in training culture that focuses on a high standard of professional development for trainers and trainees. The objectives of this predoctoral training program are to: (1) Build and sustain an equitable and inclusive training environment for an increasingly diverse group of PhD students. (2) Integrate training in the design and implementation of rigorous and transparently reported experimentation throughout the program. (3) Integrate training in quantitative and computational approaches throughout training program. (4) Integrate career exploration and student professional development throughout the program. Faculty trainers in the Molecular Biology, Cell Biology, and Biochemistry Graduate Program (MCBGP) are accomplished scientists who are drawn from 11 Departments at Brown University and the Warren Alpert Medical School. The mission of the MCBGP is to train the next generation of leaders in biomedical research to probe the molecular mechanisms of cellular and biochemical processes by building and sustaining an equitable and inclusive training environment in which a diverse group of PhD students will successfully gain quantitative, conceptual, technical, and professional skills that will allow them to conduct the rigorous and reproducible research that interdisciplinary life science demands. The MCBGP admits 9-14 students per year based on their research and academic potential. During the first or second year of graduate study, trainees will be selected from the ~20 eligible MCB graduate students for appointment to the training grant on the basis of their potential for success in research. Each year, 4 first-year and 4 second-year predoctoral students will be supported; funds to support 8 trainees per year are requested.
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