Induction of Neurite Outgrowth by Interleukin-6 Is Accompanied by Activation of Stat3 Signaling Pathway in a Variant PC12 Cell (E2) Line

doi:10.1074/jbc.271.22.13023

Induction of Neurite Outgrowth by Interleukin-6 Is Accompanied by Activation of Stat3 Signaling Pathway in a Variant PC12 Cell (E2) Line*

Yvonne Y. Wu and
Ralph A. Bradshaw^‡

From the Department of Biological Chemistry, College of Medicine, University of California, Irvine, California 92717-1700

^‡ To whom correspondence should be addressed. Tel.: 714-824-6236; Fax: 714-824-8036; E-mail: rablab{at}uci.edu

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.

Footnotes

↵* This work was supported by Research Grant AG 09735 from the National Institutes of Health. The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

NGF

nerve growth factor

FGF

fibroblast growth factor

bFGF

basic FGF

CNTF

ciliary neurotrophic factor

DMEM

Dulbecco's modified Eagle's medium

EGF

epidermal growth factor

EMSA

electrophoretic mobility shift assay

IL-6

interleukin-6

JAK

Janus kinase

LIF

leukemia inhibitory factor

LIFR

LIF receptor

STAT

signal transducers and activators of transcription

PMSF

phenylmethylsulfonyl fluoride

SIE

c-sis-inducible element

ERK

extracellular signal-regulated kinase

PAGE

polyacrylamide gel electrophoresis

DTT

dithiothreitol

SIF

c-sis-inducible factor.
- Received January 11, 1996.
- Revision received February 23, 1996.