Simulated microgravity contributes to autophagy induction by regulating AMP-activated protein kinase.

Exposure to microgravity is supposed to affect almost all biological systems, and we speculated that microgravity is potentially involved in autophagy regulation. A clinostat was used to simulate microgravity, and HEK293 cells that stably express GFP-LC3 were used for sensitive monitoring of autophagy induction. The clinorotation of GFP-LC3 cells resulted in autophagosome formation in the cytoplasm and a change in autophagosomal marker expression. Autophagy induction was accompanied by phosphorylation of AMPK (Thr 172) and by the dephosphorylation of mammalian target of rapamycin. To elucidate the role of AMPK in microgravity-induced autophagy, we suppressed AMPK expression by knockdown via siRNA, which inhibited the induction of autophagy upon exposure to microgravity. In addition, the clinorotation of C2C12 myotube cells resulted in the enlarged and distinctive LC3 spots in the cytoplasm and AMPK activation. These results indicate that simulated microgravity possibly contributes to autophagy induction by regulating AMPK.

[1]  R. Candau,et al.  The role of AMP-activated protein kinase in the coordination of skeletal muscle turnover and energy homeostasis. , 2012, American journal of physiology. Cell physiology.

[2]  R. Candau,et al.  AMPK promotes skeletal muscle autophagy through activation of forkhead FoxO3a and interaction with Ulk1 , 2012, Journal of cellular biochemistry.

[3]  R. Shaw,et al.  The AMPK signalling pathway coordinates cell growth, autophagy and metabolism , 2011, Nature Cell Biology.

[4]  D. Clemmons,et al.  Suppression of AMPK activation via S485 phosphorylation by IGF-I during hyperglycemia is mediated by AKT activation in vascular smooth muscle cells. , 2011, Endocrinology.

[5]  I. Jang,et al.  The expression of damage-regulated autophagy modulator 2 (DRAM2) contributes to autophagy induction , 2012, Molecular Biology Reports.

[6]  C. Kang,et al.  Impact of simulated microgravity on microvascular endothelial cell apoptosis , 2011, European Journal of Applied Physiology.

[7]  B. Viollet,et al.  AMPK and mTOR regulate autophagy through direct phosphorylation of Ulk1 , 2011, Nature Cell Biology.

[8]  A. Zallone,et al.  Microgravity during spaceflight directly affects in vitro osteoclastogenesis and bone resorption , 2009, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[9]  R A Knight,et al.  Guidelines for the use and interpretation of assays for monitoring cell death in higher eukaryotes , 2009, Cell Death and Differentiation.

[10]  Dalia I. Hemdan,et al.  Polyphenols prevent clinorotation-induced expression of atrogenes in mouse C2C12 skeletal myotubes. , 2009, The journal of medical investigation : JMI.

[11]  Ifla Cataloguing Section.  GUIDELINES FOR USE , 2009 .

[12]  John L Cleveland,et al.  Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes , 2008, Autophagy.

[13]  A. Goldberg,et al.  FoxO3 coordinately activates protein degradation by the autophagic/lysosomal and proteasomal pathways in atrophying muscle cells. , 2007, Cell metabolism.

[14]  Armando R. Tovar,et al.  La AMPK como un sensor de energía celular y su función en el organismo , 2007 .

[15]  G. Bjørkøy,et al.  p62/SQSTM1 Binds Directly to Atg8/LC3 to Facilitate Degradation of Ubiquitinated Protein Aggregates by Autophagy* , 2007, Journal of Biological Chemistry.

[16]  U. Raue.  Skeletal muscle gene expression with age , 2007 .

[17]  N. Torres,et al.  [AMPK as a cellular energy sensor and its function in the organism]. , 2007, Revista de investigacion clinica; organo del Hospital de Enfermedades de la Nutricion.

[18]  Guido Kroemer,et al.  Lysosomes and autophagy in cell death control , 2005, Nature Reviews Cancer.

[19]  N. Ishioka,et al.  Clinorotation prevents differentiation of rat myoblastic L6 cells in association with reduced NF-kappa B signaling. , 2005, Biochimica et biophysica acta.

[20]  Marco Sandri,et al.  Foxo Transcription Factors Induce the Atrophy-Related Ubiquitin Ligase Atrogin-1 and Cause Skeletal Muscle Atrophy , 2004, Cell.

[21]  Daniel J Klionsky,et al.  Development by self-digestion: molecular mechanisms and biological functions of autophagy. , 2004, Developmental cell.

[22]  I. Nonaka,et al.  Skeletal muscle gene expression in space‐flown rats , 2004, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[23]  B. Kemp,et al.  Intrasteric control of AMPK via the γ1 subunit AMP allosteric regulatory site , 2004 .

[24]  D. Vertommen,et al.  Identification of Phosphorylation Sites in AMP-activated Protein Kinase (AMPK) for Upstream AMPK Kinases and Study of Their Roles by Site-directed Mutagenesis* , 2003, Journal of Biological Chemistry.

[25]  M. Delp,et al.  SIMULATED MICROGRAVITY ALTERS MESENTERIC RESISTANCE ARTERY VASOCONSTRICTOR RESPONSIVENESS , 2002 .

[26]  G. Schulze-Tanzil,et al.  Simulated microgravity alters differentiation and increases apoptosis in human follicular thyroid carcinoma cells , 2002, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[27]  K. Kita,et al.  EFFECTS OF 3‐D CLINO‐ROTATION ON GENE EXPRESSION IN HUMAN FIBROBLAST CELLS , 2002, Cell biology international.

[28]  E. Wang,et al.  Peptide Elongation Factor eEF1A-2/S1 Expression in Cultured Differentiated Myotubes and Its Protective Effect against Caspase- 3-mediated Apoptosis* , 2002, The Journal of Biological Chemistry.

[29]  池本 円.  Space shuttle flight(STS-90)enhances degradation of rat myosin heavy chain in association with activation of ubiquitin-proteasome pathway , 2002 .

[30]  D J Glass,et al.  Identification of Ubiquitin Ligases Required for Skeletal Muscle Atrophy , 2001, Science.

[31]  Takeshi Noda,et al.  LC3, a mammalian homologue of yeast Apg8p, is localized in autophagosome membranes after processing , 2000, The EMBO journal.

[32]  T. Nagaya,et al.  Rotation in clinostat results in apoptosis of osteoblastic ROS 17/2.8 cells. , 2000, Journal of gravitational physiology : a journal of the International Society for Gravitational Physiology.

[33]  A. Goldberg,et al.  Importance of the ATP-Ubiquitin-Proteasome Pathway in the Degradation of Soluble and Myofibrillar Proteins in Rabbit Muscle Extracts* , 1996, The Journal of Biological Chemistry.

[34]  M. Solomon,et al.  A Predictive Scale for Evaluating Cyclin-dependent Kinase Substrates , 1996, The Journal of Biological Chemistry.

[35]  G. Lopaschuk,et al.  High Rates of Fatty Acid Oxidation during Reperfusion of Ischemic Hearts Are Associated with a Decrease in Malonyl-CoA Levels Due to an Increase in 5′-AMP-activated Protein Kinase Inhibition of Acetyl-CoA Carboxylase (*) , 1995, The Journal of Biological Chemistry.

[36]  D. Hardie,et al.  Role of the AMP-activated protein kinase in the cellular stress response , 1994, Current Biology.

[37]  D. Riley,et al.  Hypogravity‐induced atrophy of rat soleus and extensor digitorum longus muscles , 1987, Muscle & nerve.