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Imaging Intracellular Ca2+ Signals in Striatal Astrocytes from Adult Mice Using Genetically-encoded Calcium Indicators
Published on: November 19, 2014
Astrocytes express Mxi2, a splice isoform of p38MAPK
Guillermo Bodega1, Sergio Ciordia, Isabel Suárez
1Departamento de Biología Celular y Genética, Facultad de Biología, Universidad de Alcalá, Alcalá de Henares, 28871 Madrid, Spain. guillermo.bodega@uah.es
Abstract:
Mitogen-activated protein kinases (MAPKs) are a superfamily of cytoplasmic serine/threonine kinases that transduce many types of extracellular stimuli into cellular responses. p38MAPK is a member of this family with its active form in a diphosphorylated state (p38MAPKdiP). Two strong anti-p38MAPKdiP immunoreactive bands (apparent molecular weight 38 and 34 kDa) were detected by Western blotting in cultured astrocytes. Using a specific antibody and employing immunoprecipitation procedures and SELDI-TOF analysis, the 34 kDa band was found to correspond to Mxi2, a splice variant of p38MAPK; cultured astrocytes therefore express Mxi2. Separate protein extractions of different subcellular fractions, and fluorescent immunovisualisation employing confocal microscopy, showed Mxi2 to have a non-nuclear, cytosolic distribution in the studied cells. ERK1/2, protein whose intracellular distribution is influenced by Mxi2, showed the same cytoplasmic pattern than Mxi2.
Insights
Cultured astrocytes express Mxi2, a p38MAPK splice variant, primarily in the cytoplasm. This finding reveals Mxi2
Area of Science:
- Cellular Biology
- Molecular Biology
- Signal Transduction
Background:
- Mitogen-activated protein kinases (MAPKs) are crucial signaling molecules.
- p38MAPK, a key MAPK, is activated through diphosphorylation (p38MAPKdiP).
- Understanding MAPK variants like Mxi2 is vital for cellular response research.
Purpose of the Study:
- To identify the p38MAPK splice variant expressed in cultured astrocytes.
- To determine the subcellular localization of Mxi2 in these cells.
- To investigate the relationship between Mxi2 and ERK1/2 distribution.
Main Methods:
- Western blotting to detect p38MAPKdiP.
- Immunoprecipitation and SELDI-TOF analysis to identify protein bands.
- Confocal microscopy for subcellular localization studies.
Main Results:
- Two p38MAPKdiP immunoreactive bands (38 and 34 kDa) were detected in astrocytes.
- The 34 kDa band was identified as Mxi2, a p38MAPK splice variant.
- Mxi2 was localized to the cytosol, influencing ERK1/2 cytoplasmic distribution.
Conclusions:
- Cultured astrocytes express the p38MAPK splice variant Mxi2.
- Mxi2 predominantly resides in the cytosol of astrocytes.
- Mxi2's cytosolic localization impacts ERK1/2 distribution within the cell.
