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Preferential involvement of MEK1 in the tumor necrosis factor-alpha-induced activation of p42mapk/erk2 in mouse
B W Winston1, L K Remigio, D W Riches
1Department of Pediatrics, National Jewish Center for Immunology and Respiratory Medicine, Denver, Colorado 80206, USA.
Abstract:
The pleiotropic cytokine tumor necrosis factor-alpha (TNF alpha) controls the expression of multiple gene products in macrophages and plays an important role in host defense. TNF alpha is recognized by the receptors, CD120a (p55) and CD120b (p75). Ligation of CD120a (p55) by TNF alpha or by anti-receptor agonistic antibodies initiates signal transduction leading to the activation of mitogen-activated protein kinases (MAPKs) (p42mapk/erk2 and p44mapk/erk1). Phosphorylation and activation of MAPK are mediated by MAPK kinase (MEK), a family of Thr/Tyr kinases. In this study, we investigated the preferential involvement of the MEK isoforms MEK1 and MEK2 in the activation of p42mapk/erk2 in mouse macrophages stimulated with TNF alpha. Exposure of macrophages to TNF alpha stimulated a time-dependent increase in the activity of MEK1 as measured by an in vitro kinase assay using kinase-inactive p42mapk/erk2 (rMAPKkd) as substrate in the presence of gamma-[32P]ATP. Maximal activation of MEK1 was detected at 10 min poststimulation and coincided with maximal transphosphorylation of Tyr and Thr residues of rMAPKkd. By contrast, there was no evidence of MEK2 activation in macrophages in response to TNF alpha. These data suggest that MEK1 is the preferred substrate for MEK kinase, the upstream kinase implicated in activation of the MAPK pathway in macrophages by TNF alpha.
Insights
Tumor necrosis factor-alpha (TNF alpha) activates mitogen-activated protein kinases (MAPKs) in macrophages. This study reveals MEK1, not MEK2, is preferentially involved in TNF alpha-induced p42mapk/erk2 activation.
Area of Science:
- Immunology
- Cell Signaling
- Molecular Biology
Background:
- Tumor necrosis factor-alpha (TNF alpha) is a pleiotropic cytokine crucial for host defense, regulating gene expression in macrophages.
- TNF alpha signals through receptors CD120a (p55) and CD120b (p75), initiating signal transduction pathways.
- Mitogen-activated protein kinases (MAPKs), including p42mapk/erk2 and p44mapk/erk1, are key downstream effectors activated by TNF alpha signaling.
Purpose of the Study:
- To investigate the specific involvement of MEK isoforms, MEK1 and MEK2, in the activation of p42mapk/erk2.
- To elucidate the preferential role of MEK1 versus MEK2 in TNF alpha-mediated MAPK pathway activation in macrophages.
Main Methods:
- Primary mouse macrophages were stimulated with TNF alpha.
- In vitro kinase assays were performed using kinase-inactive p42mapk/erk2 (rMAPKkd) as a substrate to measure MEK1 and MEK2 activity.
- Time-course analysis was conducted to determine the kinetics of MEK activation post-TNF alpha stimulation.
Main Results:
- TNF alpha stimulation induced a time-dependent increase in MEK1 activity in macrophages.
- Maximal MEK1 activation occurred at 10 minutes post-stimulation, coinciding with maximal phosphorylation of rMAPKkd.
- No significant activation of MEK2 was observed in response to TNF alpha stimulation.
Conclusions:
- MEK1 is preferentially involved in the activation of p42mapk/erk2 in mouse macrophages stimulated by TNF alpha.
- These findings suggest MEK1 is the preferred substrate for MEK kinase, an upstream activator in the TNF alpha-induced MAPK pathway.
- This highlights a specific isoform preference in the MAPK signaling cascade initiated by TNF alpha in macrophages.