Mitogenic Signaling by Ret/ptc2 Requires Association with Enigma via a LIM Domain

Abstract

PC12-E2 cells, a stable variant subcloned from native cell populations, produce neurites in a rapid, transcription-independent manner upon exposure to nerve growth factor (NGF) or basic fibroblast growth factor (bFGF). They also give a similar morphological response to interleukin-6 (IL-6), which is, however, transcription-dependent and with a slower onset, a phenomenon basically not observed in native PC12 cells. The response profile of PC12-E2 cells to NGF and bFGF is similar to that observed for native PC12 cells pre-exposed (primed) to NGF, and such cells also respond to IL-6 in a fashion indistinguishable from PC12-E2 cells. Mechanistically, NGF and bFGF induce a sustained phosphorylation and activation of ERK1 and ERK2 in both cells, while IL-6 produces only a transient and weak tyrosine phosphorylation. However, it does stimulate a prolonged and biphasic tyrosine phosphorylation and nuclear translocation of Stat3 (signal transducers and activators of transcription 3; at least 24 h) and, to a lesser extent, Stat1. Gel shift and supershift analyses confirm that IL-6 predominantly activates Stat3 (and some Stat1) and stimulates sis-inducible element binding activity. Other members of the same cytokine subfamily, including ciliary neurotrophic factor and leukemia inhibitory factor, also cause a transient initial phase of tyrosine phosphorylation and activation of Stat1 and Stat3 (up to 1 h) but fail to stimulate a second phase of response and do not produce significant neurites. These results suggest that sustained signaling of either STAT or ERK pathways in PC12-E2 cells leads to induction of neuronal differentiation. However, only the latter is effective in native PC12 cells as the activation of Stat3 and Stat1 in native PC12 cells by IL-6 fails to induce neuronal differentiation. Thus, the response of PC12-E2 cells to IL-6 suggests the constitutive expression of a required factor(s) for differentiation, that is induced in native PC12 cells by NGF or bFGF (possibly by ERK activation), but not by IL-6 via Janus kinase/STAT activation. This factor(s), which has a sufficient half-life to allow primed cells to remain responsive to IL-6 for several days, is necessary but not sufficient for differentiation (as measured by neurite proliferation) to occur.

Microarray analysis refines classification of non-medullary thyroid tumours of uncertain malignancy

“…We identified mitogenic signatures involving the cAMP (TSHR) and the mitogenactivated protein kinase (GRB10 and NR2F2) pathways, as well as lymphangiogenesis event (vascular endothelial growth factor-C), as described in the literature (Durick et al, 1996;Du Villard et al, 2000;More et al, 2003;De la Torre et al, 2006). This may be associated with the proliferative and invasive signature recently identified in the relatively indolent FVPTC (Finn et al, 2007) resembling to our T-UM.…”

Intracellular signaling of the Ufo/Axl receptor tyrosine kinase is mediated mainly by a multi-substrate docking-site

“…We conclude, that these observations may result from secondary interactions, such as binding of GRB2 via SHC and/or phosphotyrosine independent binding. The latter has been observed in several cases recently (HolgadoMadruga et al, 1996;Galcheva-Gargova et al, 1996;Wu et al, 1996;Castagnino et al, 1995;Durick et al, 1996;Charest et al, 1996). Other protein domains of the respective receptor substrates could be involved in the interaction as well.…”

Inhibition of activated Ras-induced neuronal differentiation of PC12 cells by the LIM domain of LIM-kinase 1

“…Point mutations of conserved cysteine residues that coordinate zinc ion by glycine residues in LIM1 and LIM2 motif of DK mutant of LIMK1 suggest that the zinc-coordinated structure of either one of the two LIM motifs is required to exhibit inhibitory activity. Based on the signi®cant di erence in amino acid sequences of LIM1 and LIM2 motifs in LIMK1, the respective LIM motifs likely have distinct target binding speci®cities, as previously noted for LIM motifs in zyxin and enigma, which contain multiple LIM motifs with distinct binding speci®cities in the single molecule (Schmeichel and Beckerle, 1994;Wu and Gill, 1994;Durick et al, 1996). Thus, the respective LIM motifs in LIMK1 may function by binding with distinct sites of the single target protein or distinct proteins that are involved in Ras-induced di erentiation signaling.…”