Sphingosine 1-phosphate induces cytoskeletal reorganization in C2C12 myoblasts: physiological relevance for stress fibres in the modulation of ion current through stretch-activated channels.
Sphingosine 1-phosphate (S1P) is a bioactive lipid that is abundantly present in the serum and mediates multiple biological responses. With the aim of extending our knowledge on the role played by S1P in the regulation of cytoskeletal reorganization, native as well as C2C12 myoblasts stably transfected with green fluorescent protein (GFP)-tagged alpha- and beta-actin constructs were stimulated with S1P (1 microM) and observed under confocal and multiphoton microscopes. The addition of S1P induced the appearance of actin stress fibres and focal adhesions through Rho- and phospholipase D (PLD)-mediated pathways. The cytoskeletal response was dependent on the extracellular action of S1P through its specific surface receptors, since the intracellular delivery of the sphingolipid by microinjection was unable to modify the actin cytoskeletal assembly. Interestingly, it was revealed by whole-cell patch-clamp that S1P-induced stress fibre formation was associated with increased ion currents and conductance through stretch-activated channels (SACs), thereby suggesting a possible regulatory role for organized actin in channel sensitivity. Experiments aimed at stretching the plasma membrane of C2C12 cells, using the cantilever of an atomic force microscope, indicated that there was a Ca2+ influx through putative SACs. In conclusion, the present data suggest novel mechanisms of S1P signalling involving actin cytoskeletal reorganization and Ca2+ elevation through SACs that might influence myoblastic functions.
Formigli
Lucia
L
Department of Anatomy, Histology and Forensic Medicine, Interuniversitary Institute of Miology (IIM), 85 50134 Florence, Italy.
Meacci
Elisabetta
E
Sassoli
Chiasa
C
Chellini
Flaminia
F
Giannini
Rosalba
R
Quercioli
Franco
F
Tiribilli
Bruno
B
Squecco
Roberta
R
Bruni
Paola
P
Francini
Fabio
F
Zecchi-Orlandini
Sandra
S
D000199:Actins / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism*; D004594:Electrophysiology; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000378:metabolism; D017136:Ion Transport; D007477:Ions; D008246:Lysophospholipids / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D018625:Microscopy, Atomic Force; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D032446:Myoblasts / Q000378:metabolism; D018408:Patch-Clamp Techniques; D010739:Phospholipase D / Q000737:chemistry / Q000378:metabolism; D015398:Signal Transduction; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000502:physiology; D022502:Stress Fibers / Q000378:metabolism; D014162:Transfection; D020741:rho GTP-Binding Proteins / Q000378:metabolism
D000199:Actins; D007477:Ions; D008246:Lysophospholipids; D049452:Green Fluorescent Proteins; C060506:sphingosine 1-phosphate; D010739:Phospholipase D; D020741:rho GTP-Binding Proteins; D013110:Sphingosine; D002118:Calcium