The RhoGDI-alpha/JNK signaling pathway plays a significant role in mycophenolic acid-induced apoptosis in an insulin-secreting cell line.
Mycophenolic acid (MPA)-induced beta-cell toxicity is an important factor for islet graft function. The signal transduction mechanisms underlying this process have not been fully explored. Using a proteomics approach, we examined protein expression patterns in MPA-treated RIN-5 cells and found that RhoGDI-alpha expression is altered by MPA-treatment. We examined the relationship between RhoGDI-alpha expression and activated JNK during MPA-induced apoptosis. Cells were treated with N-acetyl-cysteine (NAC), caspase inhibitor, JNK inhibitor, guanosine or GTP for 1 h before being treated with MPA. To investigate the regulatory effects of RhoGDI-alpha on JNK activity, we examined cells showing either elevated or reduced expression of RhoGDI-alpha as a result of transfection with cDNA or siRNA constructs, respectively. MPA significantly increased cell death, caspase-3 expression and JNK activation, but it decreased the expression of a protein spot 25 observed by two-dimensional electrophoresis. This protein 25 was identified as RhoGDI-alpha by mass spectrometry. MPA-induced cell death and down-regulation of RhoGDI-alpha were prevented by guanosine, GTP or a JNK inhibitor. However, MPA-induced cell death was partially restored by treatment with a caspase inhibitor, but not by NAC treatment. RhoGDI-alpha expression was not affected by treatment with NAC or caspase inhibitor. Over-expression of RhoGDI-alpha increased cell viability and decreased activated JNK expression following exposure to MPA, whereas knockdown of RhoGDI-alpha enhanced MPA-induced cell death and increased the activation of JNK. In conclusion, MPA induces significant apoptosis in insulin-secreting cells via down-regulation of RhoGDI-alpha linked with increased JNK expression. This RhoGDI-alpha/JNK pathway might be the focus of therapeutic target for the prevention of MPA-induced islet apoptosis.
Park
Yun-Jong
YJ
The Research Institute for Transplantation, Yonsei University College of Medicine, Seoul, South Korea.
Ahn
Hyung Joon
HJ
Chang
Hye Kyung
HK
Kim
Joon Ye
JY
Huh
Kyu Ha
KH
Kim
Myoung Soo
MS
Kim
Yu Seun
YS
D000704:Analysis of Variance; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001483:Base Sequence; D053148:Caspase 3 / Q000378:metabolism; D002460:Cell Line; D015180:Electrophoresis, Gel, Two-Dimensional; D020730:Guanine Nucleotide Dissociation Inhibitors / Q000378:metabolism*; D050417:Insulin-Secreting Cells / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009173:Mycophenolic Acid / Q000494:pharmacology*; D040901:Proteomics; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors
D020730:Guanine Nucleotide Dissociation Inhibitors; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors; D048031:JNK Mitogen-Activated Protein Kinases; D053148:Caspase 3; D009173:Mycophenolic Acid