Sensitivity Analysis of Intracellular Signaling Pathway Kinetics Predicts Targets for Stem Cell Fate Control

Directing stem cell fate requires knowledge of how signaling networks integrate temporally and spatially segregated stimuli. We developed and validated a computational model of signal transducer and activator of transcription-3 (Stat3) pathway kinetics, a signaling network involved in embryonic stem cell (ESC) self-renewal. Our analysis identified novel pathway responses; for example, overexpression of the receptor glycoprotein-130 results in reduced pathway activation and increased ESC differentiation. We used a systematic in silico screen to identify novel targets and protein interactions involved in Stat3 activation. Our analysis demonstrates that signaling activation and desensitization (the inability to respond to ligand restimulation) is regulated by balancing the activation state of a distributed set of parameters including nuclear export of Stat3, nuclear phosphatase activity, inhibition by suppressor of cytokine signaling, and receptor trafficking. This knowledge was used to devise a temporally modulated ligand delivery strategy that maximizes signaling activation and leads to enhanced ESC self-renewal.

[1]  Kwang-Hyun Cho,et al.  In silico identification of the key components and steps in IFN‐γ induced JAK‐STAT signaling pathway , 2005, FEBS letters.

[2]  Kevin M. McBride,et al.  STAT3 nuclear import is independent of tyrosine phosphorylation and mediated by importin-alpha3. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[3]  U. Grohmann,et al.  Cutting Edge: Silencing Suppressor of Cytokine Signaling 3 Expression in Dendritic Cells Turns CD28-Ig from Immune Adjuvant to Suppressant1 , 2005, The Journal of Immunology.

[4]  J. Schneider-Mergener,et al.  The tyrosine 974 within the LIF-R-chain of the gp130/LIF-R heteromeric receptor complex mediates negative regulation of LIF signalling. , 2005, Cellular signalling.

[5]  P. Zandstra,et al.  Spatial Organization of Embryonic Stem Cell Responsiveness to Autocrine Gp130 Ligands Reveals an Autoregulatory Stem Cell Niche , 2006, Stem cells.

[6]  M. Katsuki,et al.  STAT3 activation is sufficient to maintain an undifferentiated state of mouse embryonic stem cells , 1999, The EMBO journal.

[7]  Bernd Giese,et al.  Long Term Association of the Cytokine Receptor gp130 and the Janus Kinase Jak1 Revealed by FRAP Analysis* , 2003, Journal of Biological Chemistry.

[8]  D. Lauffenburger,et al.  A ligand-receptor signaling threshold model of stem cell differentiation control: a biologically conserved mechanism applicable to hematopoiesis. , 2000, Blood.

[9]  D. Lauffenburger,et al.  A Computational Study of Feedback Effects on Signal Dynamics in a Mitogen‐Activated Protein Kinase (MAPK) Pathway Model , 2001, Biotechnology progress.

[10]  Stefan Rose-John,et al.  Dynamics of the gp130 cytokine complex: A model for assembly on the cellular membrane , 2005, Protein science : a publication of the Protein Society.

[11]  Radu Dobrin,et al.  Dissecting self-renewal in stem cells with RNA interference , 2006, Nature.

[12]  H. Volk,et al.  Preassociation of nonactivated STAT3 molecules demonstrated in living cells using bioluminescence resonance energy transfer: a new model of STAT activation? , 2004, Journal of leukocyte biology.

[13]  D. Aaronson,et al.  A Road Map for Those Who Don't Know JAK-STAT , 2002, Science.

[14]  P. Heinrich,et al.  Principles of interleukin (IL)-6-type cytokine signalling and its regulation. , 2003, The Biochemical journal.

[15]  S. Bhattacharya,et al.  Regulation of Stat3 nuclear export. , 2003, The Journal of clinical investigation.

[16]  Donald Metcalf,et al.  Myeloid leukaemia inhibitory factor maintains the developmental potential of embryonic stem cells , 1988, Nature.

[17]  J. Timmer,et al.  Identification of nucleocytoplasmic cycling as a remote sensor in cellular signaling by databased modeling , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[18]  P. Heinrich,et al.  Different protein turnover of interleukin-6-type cytokine signalling components. , 1999, European journal of biochemistry.

[19]  P. D’Eustachio,et al.  Essential role of STAT3 for embryonic stem cell pluripotency. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[20]  Nicholas A. Peppas,et al.  Receptors: models for binding, trafficking, and signaling , 1996 .

[21]  J. Johnston,et al.  The role of the inhibitors of interleukin-6 signal transduction SHP2 and SOCS3 for desensitization of interleukin-6 signalling. , 2004, The Biochemical journal.

[22]  D. Hilton,et al.  Kinetic analyses of the binding of leukemia inhibitory factor to receptor on cells and membranes and in detergent solution. , 1992, The Journal of biological chemistry.

[23]  E. Gilles,et al.  Computational modeling of the dynamics of the MAP kinase cascade activated by surface and internalized EGF receptors , 2002, Nature Biotechnology.

[24]  D. Loebel,et al.  Lineage choice and differentiation in mouse embryos and embryonic stem cells. , 2003, Developmental biology.

[25]  D. Hilton,et al.  Distribution and comparison of receptors for leukemia inhibitory factor on murine hemopoietic and hepatic cells , 1991, Journal of cellular physiology.

[26]  Hugh D. Spence,et al.  Minimum information requested in the annotation of biochemical models (MIRIAM) , 2005, Nature Biotechnology.

[27]  D. Lauffenburger,et al.  Ligand/Receptor Signaling Threshold (LIST) Model Accounts for gp130‐Mediated Embryonic Stem Cell Self‐Renewal Responses to LIF and HIL‐6 , 2002, Stem cells.

[28]  P. Zandstra,et al.  LIF‐mediated control of embryonic stem cell self‐renewal emerges due to an autoregulatory loop , 2007, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[29]  Xinmin Cao,et al.  Interdomain Interaction of Stat3 Regulates Its Src Homology 2 Domain-mediated Receptor Binding Activity* , 2002, The Journal of Biological Chemistry.

[30]  H. Niwa,et al.  Efficient selection for high-expression transfectants with a novel eukaryotic vector. , 1991, Gene.

[31]  A. Smith,et al.  Self-renewal of pluripotent embryonic stem cells is mediated via activation of STAT3. , 1998, Genes & development.

[32]  D A Lauffenburger,et al.  Analysis of receptor internalization as a mechanism for modulating signal transduction. , 1998, Journal of theoretical biology.

[33]  Jing Ma,et al.  A Novel Sequence in the Coiled-coil Domain of Stat3 Essential for Its Nuclear Translocation* , 2003, Journal of Biological Chemistry.

[34]  C. Schindler,et al.  STATs Dimerize in the Absence of Phosphorylation* , 2003, Journal of Biological Chemistry.

[35]  D. Lauffenburger,et al.  Receptors: Models for Binding, Trafficking, and Signaling , 1993 .

[36]  F. Horn,et al.  Analysis of Stat3 (signal transducer and activator of transcription 3) dimerization by fluorescence resonance energy transfer in living cells. , 2004, The Biochemical journal.

[37]  Ana D. Lopez,et al.  Maintenance of Pluripotency in Human Embryonic Stem Cells Is STAT3 Independent , 2004, Stem cells.

[38]  H. Kitano,et al.  A comprehensive pathway map of epidermal growth factor receptor signaling , 2005, Molecular systems biology.

[39]  T. Hirano,et al.  IL-6 signal transduction and its physiological roles: the signal orchestration model. , 2003, Reviews of physiology, biochemistry and pharmacology.

[40]  Roland Lang,et al.  SOCS3 regulates the plasticity of gp130 signaling , 2003, Nature Immunology.

[41]  J. Roder,et al.  Derivation of completely cell culture-derived mice from early-passage embryonic stem cells. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[42]  Jason A. Papin,et al.  The JAK-STAT signaling network in the human B-cell: an extreme signaling pathway analysis. , 2004, Biophysical journal.

[43]  P. Heinrich,et al.  Dimerization of the cytokine receptors gp130 and LIFR analysed in single cells , 2005, Journal of Cell Science.

[44]  Satoru Shiono,et al.  Control mechanism of JAK/STAT signal transduction pathway , 2003, FEBS letters.

[45]  Yamamura Ken-ichi,et al.  Efficient selection for high-expression transfectants with a novel eukaryotic vector , 1991 .

[46]  R. Heinrich,et al.  Control of MAPK signalling: from complexity to what really matters , 2005, Oncogene.

[47]  Warren S Alexander,et al.  Absence of Suppressor of Cytokine Signalling 3 Reduces Self‐Renewal and Promotes Differentiation in Murine Embryonic Stem Cells , 2006, Stem cells.

[48]  Mamoru Tamura,et al.  Molecular dynamics of STAT3 on IL-6 signaling pathway in living cells. , 2004, Biochemical and biophysical research communications.