3.4. Effects of MQL on MAP kinase signaling pathways
Next, to investigate MAP kinases upstream of MQL-induced HO-1
expression, we cultured CD4+ T cells with U0126 (ERK
inhibitor), SB20350 (p38 inhibitor), or SP600125 (JNK inhibitor) in the
presence of MQL. As a result, MQL-induced HO-1 expression was most
significantly reduced by U0126 treatment compared with the other
inhibitors (Fig. 4A) . To explore this further, we measured
ERK1/2, p38, and JNK activation in the presence or absence of T cell
stimulation (anti-CD3/CD28 antibodies). Consistent with the previous
finding, MQL dose-dependently increased the phosphorylation of ERK, but
not p38 or JNK (Fig. 4B and C) .
The Ras-Raf-MEK-ERK signaling pathway plays a crucial role in gene
expression related to cell growth, proliferation, differentiation, and
survival30,31. Ras proteins are small GTPases that can
activate Raf-MEK-ERK signaling32. GTP-bound Ras
interacts with the Raf family (A-Raf, B-Raf, and C-Raf), inducing Raf
dimerization and phosphorylation of C-Raf (also called
Raf-1)33,34. It has also been reported that
phosphorylation of C-Raf (Ser338) can induce ERK activation in
lymphocytes such as T cells and B cells35. Therefore,
we hypothesized that C-Raf could be the signal upstream of ERK, and
indeed found that MQL treatment induced phosphorylation of C-Raf
(Ser338) (Fig. 4D) .