Key gravity‐sensitive signaling pathways drive T‐cell activation

Returning astronauts have experienced altered immune function and increased vulnerability to infection during spaceflights dating back to Apollo and Skylab. Lack of immune response in microgravity occurs at the cellular level. We analyzed differential gene expression to find gravity‐dependent genes and pathways. We found inhibited induction of 91 genes in the simulated freefall environment of the random positioning machine. Altered induction of 10 genes regulated by key signaling pathways was verified using real‐time RT‐PCR. We discovered that impaired induction of early genes regulated primarily by transcription factors NF‐κB, CREB, ELK, AP‐1, and STAT after crosslinking the T‐cell receptor contributes to T‐cell dysfunction in altered gravity environments. We have previously shown that PKA and PKC are key early regulators in T‐cell activation. Since the majority of the genes were regulated by NF‐κB, CREB, and AP‐1, we studied the pathways that regulated these transcription factors. We found that the PKA pathway was down‐regulated in vg. In contrast, PI3‐K, PKC, and its upstream regulator pLAT were not significantly down‐regulated by vectorless gravity. Since NF‐κB, AP‐1, and CREB are all regulated by PKA and are transcription factors predicted by microarray analysis to be involved in the altered gene expression in vectorless gravity, the data suggest that PKA is a key player in the loss of T‐cell activation in altered gravity.

[1]  S. Gerondakis,et al.  Rel-dependent induction of A1 transcription is required to protect B cells from antigen receptor ligation-induced apoptosis. , 1999, Genes & development.

[2]  S. Mizel,et al.  In vitro activation and nuclear translocation of NF-kappa B catalyzed by cyclic AMP-dependent protein kinase and protein kinase C , 1989, Molecular and cellular biology.

[3]  F. Birg,et al.  CD28 costimulation regulates long-term expression of the three genes (alpha, beta, gamma) encoding the high-affinity IL2 receptor. , 1995, Research in immunology.

[4]  D. Botstein,et al.  Cluster analysis and display of genome-wide expression patterns. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[5]  S. E. Ritzmann,et al.  Hematology and immunology studies , 1975 .

[6]  William Stafford Noble,et al.  Analysis of strain and regional variation in gene expression in mouse brain , 2001, Genome Biology.

[7]  P. C. Rambaut,et al.  Biochemical responses of the Skylab crewmen: An overview , 1977 .

[8]  Ash A. Alizadeh,et al.  Distinct types of diffuse large B-cell lymphoma identified by gene expression profiling , 2000, Nature.

[9]  M. Arai,et al.  Granulocyte colony-stimulating factor , 2006, Cancer investigation.

[10]  P. Baeuerle,et al.  Tumor necrosis factor β (TNF-β) induces binding of the NF-κB transcription factor to a high-affinity κB element in the TNF-β promoter , 1990 .

[11]  C. Kang,et al.  NF‐κB and AP‐1 regulate activation‐dependent CD137 (4‐1BB) expression in T cells , 2003 .

[12]  B. Bierer,et al.  Differential chemokine expression profiles in human peripheral blood T lymphocytes: dependence on T-cell coreceptor and calcineurin signaling. , 2003, Blood.

[13]  Millie Hughes-Fulford,et al.  Signal Transduction and Mechanical Stress , 2004, Science's STKE.

[14]  B. Schraven,et al.  Integration of receptor-mediated signals in T cells by transmembrane adaptor proteins. , 1999, Immunology today.

[15]  A Cogoli,et al.  Cell sensitivity to gravity. , 1984, Science.

[16]  T. Speed,et al.  Summaries of Affymetrix GeneChip probe level data. , 2003, Nucleic acids research.

[17]  J. Barnett,et al.  Propanil Inhibits Tumor Necrosis Factor-α Production by Reducing Nuclear Levels of the Transcription Factor Nuclear Factor-κB in the Macrophage Cell Line IC-21 , 2001 .

[18]  C. Scheidereit,et al.  Activation of NF‐kappa B in vivo is regulated by multiple phosphorylations. , 1994, The EMBO journal.

[19]  L. Owen-Schaub,et al.  Activation-Dependent Transcriptional Regulation of the Human fas Promoter Requires NF-κB p50-p65 Recruitment , 1999, Molecular and Cellular Biology.

[20]  D. Baltimore,et al.  Activation in vitro of NF-kappa B by phosphorylation of its inhibitor I kappa B. , 1990, Nature.

[21]  Yoshio Masuda,et al.  Evaluation of the three-dimensional clinostat as a simulator of weightlessness , 1997, Planta.

[22]  S. N. Zaloguev,et al.  Preliminary results of Cytos 2 experiment. , 1985, Acta astronautica.

[23]  A. Cogoli,et al.  Early immune response and regulation of IL-2 receptor subunits. , 2005, Cellular signalling.

[24]  H. Erdjument-Bromage,et al.  The transcriptional activity of NF-kappaB is regulated by the IkappaB-associated PKAc subunit through a cyclic AMP-independent mechanism. , 1997, Cell.

[25]  P. Baeuerle,et al.  Tumor necrosis factor beta (TNF-beta) induces binding of the NF-kappa B transcription factor to a high-affinity kappa B element in the TNF-beta promoter. , 1990, Cytokine.

[26]  J. Cazenave,et al.  The kinetics of translocation and cellular quantity of protein kinase C in human leukocytes are modified during spaceflight , 1999, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[27]  R. Huijser,et al.  DESKTOP RPM : NEW SMALL SIZE MICROGRAVITY SIMULATOR FOR THE BIOSCIENCE LABORATORY , 2000 .

[28]  C. Li,et al.  Model-based analysis of oligonucleotide arrays: expression index computation and outlier detection. , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[29]  P. Lelkes,et al.  Growing tissues in microgravity , 1998, Nature Medicine.

[30]  R. Shadduck,et al.  Granulocyte colony stimulating factor. I. Response to acute granulocytopenia. , 1971, Blood.

[31]  M Hughes-Fulford,et al.  Function of the cytoskeleton in gravisensing during spaceflight. , 2003, Advances in space research : the official journal of the Committee on Space Research.

[32]  A. Cogoli,et al.  Activation signals of T lymphocytes in microgravity. , 1996, Journal of biotechnology.

[33]  L. Samelson,et al.  The role of membrane-associated adaptors in T cell receptor signalling. , 2000, Seminars in immunology.

[34]  A. Cogoli,et al.  Simulated microgravity inhibits the genetic expression of interleukin‐2 and its receptor in mitogen‐activated T lymphocytes , 1998, FEBS letters.

[35]  C. Kang,et al.  NF-kappaB and AP-1 regulate activation-dependent CD137 (4-1BB) expression in T cells. , 2003, FEBS letters.

[36]  T. Taniguchi,et al.  The IL-2 receptor complex: its structure, function, and target genes. , 1993, Annual review of immunology.

[37]  Joshua M. Stuart,et al.  MICROARRAY EXPERIMENTS : APPLICATION TO SPORULATION TIME SERIES , 1999 .

[38]  J. Warrington,et al.  Identification and validation of endogenous reference genes for expression profiling of T helper cell differentiation by quantitative real-time RT-PCR. , 2001, Analytical biochemistry.

[39]  D. Baltimore,et al.  Activation in vitro of NF-κB" by phosphorylation of its inhibitor IκB" , 1990, Nature.

[40]  M Hughes-Fulford,et al.  Vibrational force alters mRNA expression in osteoblasts , 1997, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[41]  P. Young,et al.  Herpesvirus Entry Mediator Ligand (HVEM-L), a Novel Ligand for HVEM/TR2, Stimulates Proliferation of T Cells and Inhibits HT29 Cell Growth* , 1998, The Journal of Biological Chemistry.

[42]  A. Cogoli.  Signal transduction in T lymphocytes in microgravity. , 1997, Gravitational and space biology bulletin : publication of the American Society for Gravitational and Space Biology.

[43]  Augusto Cogoli,et al.  Mitogenic signal transduction in Tlymphocytes in microgravity , 1993, Journal of leukocyte biology.

[44]  H. Erdjument-Bromage,et al.  The Transcriptional Activity of NF-κB Is Regulated by the IκB-Associated PKAc Subunit through a Cyclic AMP–Independent Mechanism , 1997, Cell.

[45]  D. A. Wolf,et al.  Gene expression in space , 1999, Nature Medicine.

[46]  Marian L. Lewis,et al.  Spaceflight alters microtubules and increases apoptosis in human lymphocytes (Jurkat) , 1998, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[47]  Xin Chen,et al.  The TRANSFAC system on gene expression regulation , 2001, Nucleic Acids Res..

[48]  A. Cogoli,et al.  Influence of microgravity on mitogen binding and cytoskeleton in Jurkat cells. , 1999, Advances in space research : the official journal of the Committee on Space Research.

[49]  W. R. Hawkins,et al.  Clinical aspects of crew health , 1975 .

[50]  Partha S. Vasisht.  Computational Analysis of Microarray Data , 2003 .

[51]  F. Birg,et al.  CD28 costimulation regulates long-term expression of the three genes (α, β, γ) encoding the high-affinity IL2 receptor , 1995 .