Thursday, 6 August 2009

What Makes Stem Cells Tick?

Researchers identify phosphorylated signalling proteins in human embryonic stem cells Thursday, 06 August 2009 Investigators at the Burnham Institute for Medical Research (Burnham) and The Scripps Research Institute (TSRI) have made the first comparative, large-scale phosphoproteomic analysis of human embryonic stem cells (hESCs) and their differentiated derivatives. The data may help stem cell researchers understand the mechanisms that determine whether stem cells divide or differentiate, what types of cells they become and how to control those complex mechanisms to facilitate development of new therapies. The study was published in the August 6 issue of the journal Cell Stem Cell. Protein phosphorylation, the biochemical process that modifies protein activities by adding a phosphate molecule, is central to cell signalling. Using sophisticated phosphoproteomic analyses, the team of Sheng Ding, Ph.D., associate professor at TSRI, Evan Y. Snyder, M.D., Ph.D., professor and director of Burnham's Stem Cell and Regenerative Biology program, and Laurence M. Brill, Ph.D., senior scientist at Burnham's Proteomics Facility, catalogued 2,546 phosphorylation sites on 1,602 phosphoproteins. Prior to this research, protein phosphorylation in hESCs was poorly understood. Identification of these phosphorylation sites provides insights into known and novel hESC signalling pathways and highlights signalling mechanisms that influence self-renewal and differentiation. "This research will be a big boost for stem cell scientists," said Dr. Brill. "The protein phosphorylation sites identified in this study are freely available to the broader research community, and researchers can use these data to study the cells in greater depth and determine how phosphorylation events determine a cell's fate." The team performed large-scale, phosphoproteomic analyses of hESCs and their differentiated derivatives using multi-dimensional liquid chromatography and tandem mass spectrometry. The researchers then used the phosphoproteomic data as a predictive tool to target a sample of the signalling pathways that were revealed by the phosphorylated proteins in hESCs, with follow-up experiments to confirm the relevance of these phosphoproteins and pathways to the cells. The study showed that many transcription regulators such as epigenetic and transcription factors, as well as a large number of kinases are phosphorylated in hESCs, suggesting that these proteins may play a key role in determining stem cell fate. Proteins in the JNK signalling pathway were also found to be phosphorylated in undifferentiated hESCs, which suggested that inhibition of JNK signalling may lead to differentiation, a result that was confirmed in hESC cultures. These methods were extremely useful to discover novel proteins relevant to the human embryonic stem cells. For example, the team found that phosphoproteins in receptor tyrosine kinase (RTK) signalling pathways were numerous in undifferentiated hESCs. Follow-up studies used this unexpected finding to show that multiple RTKs can support hESCs in their undifferentiated state. This research shows that phosphoproteomic data can be a powerful tool to broaden understanding of hESCs and how their ultimate fate is determined. With this knowledge, stem cell researchers may be able to develop more focused methods to control hESC differentiation and move closer to clinical therapies. The protein phosphorylation data is available on the Cell Stem Cell website, as well as on the PRIDE website. Another group from Utrecht, The Netherlands, has performed a similar study of phosphoproteins in hESCs (see the references). About Burnham Institute for Medical Research Burnham Institute for Medical Research is dedicated to revealing the fundamental molecular causes of disease and devising the innovative therapies of tomorrow. Burnham, with operations in California and Florida, is one of the fastest-growing research institutes in the country. The Institute ranks among the top-four institutions nationally for NIH grant funding and among the top-25 organizations worldwide for its research impact. Burnham utilizes a unique, collaborative approach to medical research and has established major research programs in cancer, neurodegeneration, diabetes, infectious and inflammatory and childhood diseases. The Institute is known for its world-class capabilities in stem cell research and drug discovery technologies. Burnham is a non-profit, public benefit corporation. About The Scripps Research Institute The Scripps Research Institute is one of the world's largest independent, non-profit biomedical research organizations, at the forefront of basic biomedical science that seeks to comprehend the most fundamental processes of life. Scripps Research is internationally recognized for its discoveries in immunology, molecular and cellular biology, chemistry, neurosciences, autoimmune, cardiovascular, and infectious diseases, and synthetic vaccine development. Established in its current configuration in 1961, it employs approximately 3,000 scientists, postdoctoral fellows, scientific and other technicians, doctoral degree graduate students, and administrative and technical support personnel. Scripps Research is headquartered in La Jolla, California. It also includes Scripps Florida, whose researchers focus on basic biomedical science, drug discovery, and technology development. Scripps Florida is located in Jupiter, Florida. References: Phosphoproteomic Analysis of Human Embryonic Stem Cells Laurence M. Brill, Wen Xiong, Ki-Bum Lee, Scott B. Ficarro, Andrew Crain , Yue Xu, Alexey Terskikh, Evan Y. Snyder, and Sheng Ding Cell Stem Cell, Volume 5, Issue 2, 204-213, 7 August 2009, doi:10.1016/j.stem.2009.06.002 Phosphorylation Dynamics during Early Differentiation of Human Embryonic Stem Cells Dennis Van Hoof, Javier Muñoz , Stefan R. Braam, Martijn W.H. Pinkse , Rune Linding , Albert J.R. Heck, Christine L. Mummery and Jeroen Krijgsveld Cell Stem Cell, Volume 5, Issue 2, 214-226, 7 August 2009, doi:10.1016/j.stem.2009.05.021 Unraveling the Human Embryonic Stem Cell Phosphoproteome Andrew P. Hutchins and Paul Robson Cell Stem Cell, Volume 5, Issue 2, 126-128, 7 August 2009, doi:10.1016/j.stem.2009.07.007 ......... ZenMaster

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