Cox, Sarah Elizabeth; (2003) Protein kinases and protein phosphatases in the central nervous system-Identification, characterisation and functional correlates. Doctoral thesis (Ph.D.), University College London (United Kingdom).

Text Protein_kinases_and_protein_ph.pdf Download (18MB)

Abstract

Protein phosphorylation is an important mechanism in the regulation of many brain cell functions. The precise molecular mechanisms can be elucidated by identification and characterisation of the protein kinases and phosphatases involved. Astrocytes play a vital role in the physiology and metabolic activity of the vertebrate brain and the present study sought to explore the roles of some specific protein kinases and phosphatases in CNS astrocytes and rat brain. The four main protein phosphatases (PP1, PP2A, PP2B, PP2C) were identified in astrocytes and their relative activities measured and compared to other cell types. There was an approximately equal expression of PP1 and PP2A in astrocytes whereas hepatocytes contain more PP2A and human lung mast cells more PP1. Total PP activity increased as astrocytes developed in culture with a peak at day 10. However, there was no change in the relative activities of each PP over this period. Inhibition of PP1 and PP2A by okadaic acid had profound effects on phospholipid associated signal transduction pathways causing stimulation of arachidonic acid release and inhibition of noradrenaline-stimulated inositol phosphate production. There were subtle differences in the properties of AMP-activated protein kinase (AMPK) from rat brain compared to liver. Brain AMPK had a lower specific activity but was more sensitive to activation by AMP and less sensitive to inhibition by phosphocreatine. Levels of AMPK activity were similar in different brain regions and showed no changes during the first 21 days of postnatal development. The activity of ACC, a major substrate of AMPK, was highest in cortex and lowest in cerebellum and during postnatal development peaked at day 14. The activities of PP2C and PP2A, responsible for dephosphorylation of AMPK and ACC, respectively, did not change over 21 days of postnatal brain development. AMPK activity was identified in astrocytes but ACC activity was undetectable. A possible alternative AMPK target in astrocytes, glial fibrillary acidic protein (GFAP), was investigated as a possible substrate for AMPK. The activity ratio of cAMP-dependent protein kinase (PKA) was higher in cerebellum than the other brain regions examined and this correlated with a greater proportion of the type I isoform of PKA. Type I PKA from cerebellum exhibited significant cAMP- independent activity which was not observed in type I PKA from rat heart. A tendency for the PKA activity ratio to decrease through 21 days post-natal brain development and through 21 days astrocyte growth in culture, correlated with a tendency for a decrease in the ratio of type I: type II PKA activity.

Download activity - last month

Export as CSVXMLJSON

Download activity - last 12 months

Export as JSONXMLCSV

Downloads by country - last 12 months

Export as JSONCSVXML

Archive Staff Only

View Item