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The pool of map kinase associated with microtubules is small but constitutively active
M Morishima-Kawashima1, K S Kosik
1Harvard Medical School, Boston, Massachusetts, USA.
Abstract:
Mitogen-activated protein kinase (MAPK) is activated by many kinds of stimuli and plays an important role in integrating signal transduction cascades. MAPK is present abundantly in brain, where we have studied its association with microtubules. Immunofluorescence of primary hippocampal neurons revealed that MAPK staining co-localized with microtubules and biochemical analyses showed that MAPK co-purified with microtubules. Approximately 4% of MAPK in cytosolic extracts was associated with microtubules, where it was associated with both tubulin and microtubule-associated proteins (MAPs) fractions. Further fractionation of MAPs suggested that a portion of MAPK is associated with MAP2. An association with MAP2 was also demonstrated by co-immunoprecipitation and in vitro binding experiments. A similar association was shown for the juvenile MAP2 isoform, MAP2C. The pool of MAPK associated with microtubules had a higher activity relative to the nonassociated pool in both brain and proliferating PC12 cells. Although MAPK was activated by nerve growth factor in PC12 cells, the activity of microtubule-associated MAPK did not further increase. These results raise the possibility that microtubule-associated MAPK operates through constitutive phosphorylation activity to regulate microtubule function in neurons.
Insights
Mitogen-activated protein kinase (MAPK) associates with brain microtubules and MAP2. This microtubule-bound MAPK exhibits higher activity, suggesting a role in neuronal function regulation.
Area of Science:
- Neuroscience
- Cell Biology
- Biochemistry
Background:
- Mitogen-activated protein kinase (MAPK) is crucial for signal transduction.
- MAPK is abundant in the brain, and its interaction with neuronal structures is of interest.
Purpose of the Study:
- To investigate the association of MAPK with microtubules in neurons.
- To determine the functional significance of MAPK-microtubule interactions.
Main Methods:
- Immunofluorescence in primary hippocampal neurons.
- Biochemical co-purification and fractionation assays.
- Co-immunoprecipitation and in vitro binding experiments.
Main Results:
- MAPK co-localizes and co-purifies with microtubules in brain neurons.
- A fraction of MAPK binds to tubulin and microtubule-associated proteins (MAPs), including MAP2 and MAP2C.
- Microtubule-associated MAPK shows higher activity compared to non-associated MAPK in brain and PC12 cells.
Conclusions:
- MAPK associates with neuronal microtubules and MAP2.
- Microtubule-bound MAPK possesses elevated activity, potentially regulating microtubule function via constitutive phosphorylation in neurons.