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Systems Biology of Stem Cell Fate and Cellular Reprogramming

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Systems Biology of Stem Cell Fate and Cellular Reprogramming REVIEWS STEM CELLS Systems biology of stem cell fate and cellular reprogramming Ben D. MacArthur*‡, Avi Ma’ayan‡ and Ihor R. Lemischka* Abstract | Stem cell differentiation and the maintenance of self-renewal are intrinsically complex processes requiring the coordinated dynamic expression of hundreds of genes and proteins in precise response to external signalling cues. Numerous recent reports have used both experimental and computational techniques to dissect this complexity. These reports suggest that the control of cell fate has both deterministic and stochastic elements: complex underlying regulatory networks define stable molecular ‘attractor’ states towards which individual cells are drawn over time, whereas stochastic fluctuations in gene and protein expression levels drive transitions between coexisting attractors, ensuring robustness at the population level. Inner cell mass Stem cells have a crucial role during development, To begin to deconstruct these intrinsically complex Early cells in the embryo that tissue regeneration and healthy homeostatic cell turn­ regulatory mechanisms it is now common for stem cell generate all lineages of the over. Collectively, all stem cells share the ability to self­ studies to combine low­throughput experimental tech­ mature organism but do not renew and differentiate into various different lineages. niques with an ever­increasing range of different high­ give rise to the placenta. Embryonic stem (ES) cells — which are derived from the throughput experimental techniques. Consequently, stem Blastocyst inner cell mass of the developing blastocyst — are pluri­ cell studies now often produce large amounts of data, and The embryo before potent, whereas stem cells derived from adult tissues are integrating these data into a coherent quantitative picture implantation, which contains at generally only multipotent, maintaining a limited, tissue­ of cell fate control at the systems level is an important least two distinct cell types: the 1–3 trophectoderm and the inner specific, regenerative potential . In culture, ES cells can current research challenge. To address this challenge cell mass. be propagated indefinitely, whereas adult tissue stem cells several groups have begun to apply systems biology are more limited in this capacity. approaches to understanding the regulation of stem cell Owing to their ability to generate tissue de novo fate decisions11,12,17,18. following disease or injury there is widespread hope Instead of focusing on the role of individual genes, of developing stem cell­based therapies for various proteins or pathways in biological phenomena, the aim *Department of Gene and degener ative diseases4. Recent reports have indicated that of systems biology is to characterize the ways in which Cell Medicine and Black Family Stem Cell Institute, differentiated adult cells can easily be reprogrammed to essential molecular parts interact with each other to deter­ 5,6 19–23 Mount Sinai School of an embryonic­like pluripotent state (thus potentially mine the collective dynamics of the system as a whole . Medicine, One Gustave Levy providing a patient­specific source of pluripotent stem However, it is difficult to understand collective behaviour Place, New York, New York cells), as well as be reprogrammed to other adult cell in complex systems using experimental approaches alone. 10029, USA. ‡ types without intermediate reversion to a pluripotent Therefore systems biology approaches often employ high­ Department of Pharmacology 7,8 and Systems Therapeutics, state . These findings have served to intensify interest throughput experimental techniques alongside theo­ Systems Biology Center New in understanding the molecular basis of cell fate regu­ retical and computational methods, which are specifically York (SBCNY), Mount Sinai lation and the potential therapeutic uses of stem cells9. designed to dissect collective phenomena in complex sys­ School of Medicine, However, before such stem cell­based therapies can be tems24–26. Although to date systems biology approaches are One Gustave Levy Place, 27,28 New York, New York 10029, routinely and safely developed, numerous crucial issues mostly successful in lower organisms, such as yeast and 29,30 USA. must be addressed. In particular, although great progress bacteria , the complexity of mammalian stem cell bio logy e-mails: has been made towards understanding the roles of the as well as the experimental reproducibility of many stem [email protected]; homeodomain transcription factors OCT4 (also known cell systems makes mammalian stem cell biology a good [email protected]; as POU5F1) and NANOG, as well as SRY box­containing platform for the development of future systems bio logy [email protected] doi:10.1038/nrm2766 factor 2 (SOX2) in the maintenance of stem cell pluri­ techniques. In the context of stem cell biology, the aim 10–16 Published online potency (BOX 1), the extended molecular mechanisms of systems biology approaches is to characterize the 9 September 2009 of ES cell fate control have yet to be fully determined. molecular components involved in stem cell self­renewal 672 | OCTOBER 2009 | VOLUME 10 www.nature.com/reviews/molcellbio © 2009 Macmillan Publishers Limited. All rights reserved REVIEWS Box 1 | The core embryonic stem cell transcriptional circuit approaches aim to use advances in the understanding of the molecular basis of normal cell fate decisions during development to generate strategies for the experimental Trophectoderm Tcl1 conversion of adult cells from one type to another. Hand1 In this Review we discuss a range of ways in which Tbx3 Eomes high­throughput experimental techniques and compu­ Rest Ectoderm tational methods are being fruitfully combined towards Zic3 Lhx5 the development of stem cell systems biology approaches. We begin by outlining how data from high­throughput Pou5f1 Otx1 Hesx1 experiments can be used to reconstruct accurate stem OCT4 Stat3 Hoxb1 cell regulatory networks. However, as stem cell regulatory Nanog NANOG circuits are typically intricate and contain highly nested Rex1 SOX2 Mesoderm Myf5 feedback loops and feedforward loops that give rise to com­ Sall4 Sox2 plex dynamics, it is difficult to elucidate cell behaviour T Tcf3 from this regulatory circuitry. Therefore, we also discuss Gsc how computational techniques can be used to relate Dax1 Endoderm dynamic cell behaviour to regulatory architecture. In Foxa2 particular, we focus on how cell types can be thought of Self-renewal and attractors pluripotency Gata6 as balanced states or ‘ ’ of underlying regulatory networks and the ways in which stochastic and determin­ Lineage commitment istic mechanisms interact to define cell fate. We conclude with some suggestions of directions for future work in this Maintenance of pluripotency and self-renewal in embryonic stem (ES) cells is controlled area, including ways in which these notions might be used by a complex interplay between signalling from the extracellularNature Revie wsenvironment | Molecular and Cell the Biolog y to better understand cellular reprogramming. dynamics of core transcription factors. Although self-renewal signalling pathways differ 123 between mice and humans , the core transcriptional circuitry seems to be remarkably Dissecting stem cell complexity conserved. In particular, the homeodomain transcription factors OCT4 (also known as POU5F1) and NANOG, as well as SRY box-containing factor 2 (SOX2), form a Molecular biology has entered the high­throughput transcriptional module that has a central role in maintaining ES cell identity both in age. Consequently, it is now typical for stem cell studies mice and humans10,11,13–16 (see the figure). This module is rich in positive feedback and to make use of various disparate high­throughput tech­ feedforward loops (see also BOX 2). In particular, OCT4 and SOX2 form a heterodimer niques to determine the molecular mechanisms of cell that positively regulates the expression of the Pou5f1 (which encodes OCT4), Sox2 and fate specification. These techniques include: micro­ Nanog10,11,124,125. In addition, NANOG also interacts directly with OCT4 (not shown)17 arrays to assess genome­wide mRNA expression; high­ and positively regulates the expression of all three genes11. Thus, these three transcription throughput chromatin immunoprecipitation (ChIP) factors regulate their own and each other’s expression in a highly coordinated manner, such as ChIP-on-chip31, ChIP-seq (ChIP­sequencing)32 and involving positive protein–protein and protein–DNA feedback loop interactions. ChIP-PET (ChIP­paired­end­ditag)10 to assess protein–DNA Furthermore, all three transcription factors co-occupy numerous developmentally interactions; and mass spectrometry proteomics33 and important genes and repress the expression of the genes involved in lineage commitment. 34 These include: Hand1 (heart and neural crest derivatives-expressed 1), eomesodermin phospho proteomics to assess the protein composition (Eomes) (both involved in trophectoderm development); Lhx5 (LIM homeobox 5), of molecular complexes and global changes in post­ Otx1 (orthodenticle homologue 1), Hoxb1 (all involved in ectoderm development); translational modifications. Because high­throughput Myf5 (myogenic factor 5), T (brachyury protein homologue), Gsc (goosecoid) (all involved techniques measure system­wide expression patterns, in mesoderm development); and Foxa2 (forkhead box A2) and Gata6 (GATA-binding rather than focusing on the behaviour of key molecular protein 6) (both
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