Progress 10/01/11 to 09/30/14
Outputs
Target Audience: Metabolic syndrome, a substantial medical and economical burden to the modern society, is associated with primary disturbances in adipose tissue. Proper regulation of adipose tissue during hyperplasia is essential to energy homeostasis. Detailed understanding of adipogenesis thus provides molecular insights in metabolic syndrome stratification and management. By focusing on preadipocyte differentiation, this project reveals the transformation of epigenetic landscape and involving epigenetic factors in a key step of adipogenesis. Results has wide implications to biomedical researchers, healthcare professionals and pharmaceutical companies. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? This project has provided important training opportunities for a post-doc, a graduate student and a few undergraduate students. During the award period, students were trained and vedeloped expertise in the interdisciplinary fields of cell biology, biochemistry to mass spectrometry-based proteomics. In addition, it provides partial summer support for a graduate student. The collaboration among students help synergistic progress on multiple projects. The project also partially provides support for students to attend and present at a professional annual conference. How have the results been disseminated to communities of interest? We have been disseminating the results of our studies through peer-reviewed journal articles, and presentation at professional conferences. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Adipogenesis, the differentiation of fat storage cells, is essential to the development and homeostasis of the adipose tissue. Pharmacological agents that modulate adipogenesis provide valuable therapeutic modalities in treatments of metabolic syndrome. For instance, metformin (a widely prescribed drug to treat type 2 diabetes) can decrease lipogenic gene expression and inhibit the differentiation of preadipocytes. In this project, we hypothesize that epigenetic regulation plays an essential role in adipogenesis and set off to elucidate major changes in epigenetic landscape during adipogenesis. Using a systematic approach enabled by quantitative mass spectrometric approach, we identified substantial changes in multiple histone modifications, especially the acetylation levels of lysine residues. The most prominent change takes place on histone H4 Lys16. The histone acetyltransferase (MYST1) that catalyzes histone H4 K16 acetylation directly interacts with a histone methyltransferase complex (MLL/SET), which is also implicated in adipogenesis. In addition, our study reveals complex changes in methylation on K27/K36, suggesting multiple chromatin subdomains that respond differently during preadipocyte differentiation. In summary, our study, for the first time, describes the transformation of global histone modification landscape during adipocyte differentiation. Key chromatin factors that regulate these changes in histone modifications can serve as molecular targets to modulate adipogenesis and alleviate metabolic syndrome. For instance, our results indicate MYST1 as a novel chromatin factor in adipogenesis. Conceivably, inhibitors that disrupt either its enzymatic activities or its interactions with MLL/SET complex have the potential to treat metabolic syndrome. Furthermore, the high-density, quantitative profiles of histone modifications generated from our study provide useful resources to other researchers, aiming to understand molecular details of adipogenesis and to develop pharmacological agents for the management of metabolic syndrome.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Timothy O. Street, Xiaohui Zeng, Riccardo Pellarin, Massimiliano Bonomi, Andrej Sali, Mark J.S. Kelly, Feixia Chu and David A. Agard. Elucidating the mechanism of substrate recognition by the bacterial Hsp90 molecular chaperone, J. Mol. Biol., 426 (12), 2393-2404.
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Xiaohui Zeng-Elmore, Xiong-Zhuo Gao, Riccardo Pellarin, Dina Schneidman-Duhovny, Xiu-Jun Zhang, Katie A. Kozacka, Yang Tang, Andrej Sali, Robert J. Chalkley, Rick H. Cote and Feixia Chu. Molecular architecture of photoreceptor phosphodiesterase elucidated by chemical cross-linking and integrative modeling, J. Mol. Biol., 426 (22), 3713-3728.
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Medhat Rehan, Teal Furnholm, Ryan H. Finethy, Feixia Chu, Gomaah El-Fadly, and Louis S. Tisa. Copper Tolerance by Frankia involves Surface-Binding and Copper Transport, Appl Microbiol Biotechnol., 98(18), 8005-8015.
- Type:
Journal Articles
Status:
Accepted
Year Published:
2015
Citation:
Ethan Baker, Yang Tang, Feixia Chu and Louis S. Tisa. Molecular Responses of Frankia sp. strain QA3 to Naphthalene Stress, Can. J. Microbiol., accepted.
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Progress 10/01/12 to 09/30/13
Outputs
Target Audience: This project will elucidate the transformation of epigenome during adipocyte differentiation and reveal important chromatin factors in adipocyte homeostasis. Results will be of interest to researchers, healthcare professionals and pharmaceutical companies. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? We are continuing our educational efforts at the University of New Hampshire. Using this project as the platform, both undergraduate and graduate students in my group have been trained in cell culture, cell state manipulation, histone protein purification, cell-cycle arrest and flow cytometry, immunofluorescence, western blotting, mass spectrometric analysis, protein identification and quantification. It not only helps build future workforce and human resources, but also leads to two peer-reviewed journal articles on their side-projects. Protocols we streamlined are applicable to other projects in our group with similar biochemical and proteomic workflow. Nancy Fernandes, a graduate student in the group, was awarded 3-year NSF Graduate Research Fellowship award beginning August 2013. Her dissertation research works in a synergistic manner with this project. How have the results been disseminated to communities of interest? October, 2012, invited talk in Tecan Symposium, titled "From paleoproteomics to structural biology: sample preparation to increase information density for mass spectrometry-based proteomics” (approximate audience of 100-200 people); April, 2013, invited seminar in Chemistry and Chemical Biology Department at Rensselaer Polytechnic Institute, titled “Navigating epigenetic landscape with quantitative mass spectrometry” (approximate audience of 30 people); April, 2013, two poster presentations given by students at the annual meeting of American Society of Biochemistry and Molecular Biology (approximate audience of >10,000 people). What do you plan to do during the next reporting period to accomplish the goals? We expect to gain conclusive results on histone modifications that are prominently altered during preadipocyte differentiation in a couple of months. Results will guide us to focus on a few nutrition factors implicated in regulating chromatin enzymes and test their function in preadipocyte differentiation. External grant applications will be written that leverage the outcome and impacts of this initial funding. Stakeholders will be engaged in conferences, publications and seminars.
Impacts
What was accomplished under these goals?
Metabolic syndrome raises the risk for heart disease and diabetes. Metabolic syndrome is associated with primary disturbances in adipose tissue. Therefore, the regulation of adipocyte development potentially provides a key intervention step to the management of metabolic syndrome. Consistent with this notion, several dietary components and pharmacological agents have shown to regulate adipogenesis and insulin sensitivity. In this propose study, we hypothesize that epigenetic regulation plays an essential role in adipocyte development, and hope to shed light on underlying mechanisms in adipogenesis. Results from our study on this key physiological process at the molecular level will help understand the meritorious functions of some nutritional factors in the metabolic syndrome and reveal new molecular targets for more specific nutritional interventions. This is the second year into our Hatch project, entitled ‘Epigenetic regulation in adipogenesis and its nutritional implications’. Thus far, we have locally established and expanded a preadipocyte cell line, 3T3-L1 mouse embryonic fibroblast (ATCC CL-173). We have streamlined the preadipocyte differentiation protocol using the differentiation cocktail of insulin, dexamethasone and methylisobutylxanthine. We have acquired high-density mass spectrometric data for comparative quantitation of histone modifications in differentiated and undifferentiated cells. Currently, we are analyzing data in replicates to gain statistical distribution of the data. In addition, we have delineated the boundary of experimental variability for each histone modification by analyzing control samples with biological and technical replicates. We expect to soon confidently identify histone posttranslation modifications that mediate the transformation of epigenetic landscape during adipocyte differentiation.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Bin Wu, Alys Peisley, Claire Richards, Hui Yao, Xiaohui Zeng, Cecilie Lin, Feixia Chu, Thomas Walz and Sun Hur. Structural Basis for Viral dsRNA Recognition by MDA5, Cell, 152, 276-89 (2013).
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Poshen B. Chen, Jui-Hung Hung, Taylor L. Hickman, Andrew H. Coles, James F. Carey, Zhiping Weng, Feixia Chu, and Thomas G. Fazzio. Hdac6 is a stem-cell specific modulator of Tip60-p400 function, eLife, 2:01557 (2013).
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Progress 10/01/11 to 09/30/12
Outputs
OUTPUTS: Activities and products: this is the first year into our Hatch project, entitled Epigenetic regulation in adipogenesis and its nutritional implications. Thus far, we have locally established and expanded a preadipocyte cell line, 3T3-L1 mouse embryonic fibroblast (ATCC CL-173), with media supplemented with [12C, 14N]- or [13C, 15N]-labeled amino acid Lysine. A preliminary analysis of histone proteins shows a near complete incorporation of Lysine amino acid in cells metabolically labeled with [13C, 15N]-Lys. We have also tested a couple of 3T3-L1 preadipocyte differentiation protocols, and optimized the procedure using the differentiation cocktail of insulin, dexamethasone and methylisobutylxanthine. We are now poised to scale up the preadipocyte differentiation experiments with isotope-labeled cells to obtain sufficient amount of histone proteins for quantitative analysis of histone modifications with mass spectrometry. To improve sensitivity and through-put, we have further optimized techniques for our mass spectrometric analysis workflow. Specifically, we have identified a resin that can capture hydrophilic peptides, meanwhile effective in separating peptides of various hydrophobicity. It will undoubtedly improve the detection of a few important modified peptides, including histone H3 K4me2/K4me3 peptides, which are the products of MLL3/MLL4 histone methyltransferases. MLL3 and MLL4 play a vital role in adipogenesis, and are likely to increase the K4me2/K4me3 peptide levels during differentiation. Secondly, we have screened and identified conditions for effective in-gel derivatization of free-Lys residues. With current protocol, we can achieve higher sample-handling through-put and lower biochemical variation. We are continuing our educational efforts at the University of New Hampshire. Using this project as the platform, both undergraduate and graduate students in my group have been trained in cell culture, cell state manipulation, histone protein purification, cell-cycle arrest and flow cytometry, immunofluorescence, western blotting, mass spectrometric analysis, protein identification and quantification. It not only helps build future workforce and human resource, but also lead to three peer-reviewed journal articles on their side-projects. Two undergraduate researchers just graduated this past Spring. One of them is currently enrolled as a graduate student at Duke University. The other is pursuing a part-time graduate course in Regulation Sciences at Northeastern University, and working as a full-time research technician in Ragon Institute of Mass General Hospital, MIT, and Harvard. PARTICIPANTS: Feixia Chu, Ph.D. (Principle Investigator); Xiaohui Zeng, Ph.D. (post-doctoral trainee); Sean Closs (undergraduate trainee); Kenan Mazic (undergraduate trainee). TARGET AUDIENCES: The target audience for this project includes research scientists, healthcare professionals and pharmaceutical companies. Results from the project will shed light on important elements in adipose tissue homeostasis and potential targets for therapeutic intervention of complex metabolic and cardiovascular diseases. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
At current stage, our project has already contributed to changes in knowledge. Our improved techniques are widely applicable for the analysis of histone epigenetic states involved in many other cellular processes and dysregulated in multiple diseases. Our publications also contribute to the knowledge expansion. This project has provided great training opportunities to students and a post-doc, and has contributed to the development of human resources. Therefore, it broadly brings changes in conditions in biological/agricultural research at the University of New Hampshire by strengthening competence in mass spectrometry-based proteomics.
Publications
- Brickner DG, Ahmed S, Meldi L, Thompson A, Light W, Young M, Hickman TL, Chu F, Fabre E, Brickner JH; 2012; DNA zip codes control gene positioning and interchromosomal clustering at the nuclear periphery; Developmental Cell, 22:1234-46.
- Hu H, Hu L, Yu Z, Chasse AE, Chu F, Li Z; 2012; An orphan kinesin in trypanosomes cooperates with a kinetoplastid-specific kinesin to maintain cell morphology through regulating subpellicular microtubules; J Cell Sci., 125:4126-4136.
- Wu B, Peisley A, Richards C, Yao H, Zeng X, Lin C, Chu F, Walz T, Hur S; 2012; Structural Basis for Viral dsRNA Recognition by MDA5; Cell (accepted).
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