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Regulation of amyloid precursor protein catabolism in vitro : the role of mitogen-activited protein kinase and protein kinase C

University of British Columbia

Secretory processing of the amyloid precursor protein (APP) generating soluble APP fragments occurs via two routes broadly categorized as non-amyloidogenic (yielding a truncated APP derivative known as APPS) or potentially amyloidogenic (yielding intact Aβ). These catabolic pathways are mutually exclusive and subject to regulation by various first messengers. In continuous cell lines, nonamyloidogenic processing of APP is increased by cholinergic receptor stimulation and its downstream effector protein kinase C (PKC). However, at the time our first set of experiments were initiated, mechanisms underlying APP catabolism in central neurons were largely unknown. Therefore, we examined whether or not first or second messengers of cholinergic neurotransmission increase APPS in primary cultures of rat cortical neurons. Using western blot analysis to measure secreted proteins, we determined that although activation of PKC by phorbol esters increased production of APPS, activation of muscarinic receptors by oxotremorine-M or carbachol did not. One explanation for this apparent discrepancy is that cholinergic agonists and phorbol esters activate downstream effectors differentially. One such effector may be mitogen-activated protein kinase (MAPK) also known as extracellular signal-regulated protein kinase (ERK). APP has been shown to be regulated by a variety of first messengers which also regulate this signaling pathway. Specifically, we hypothesized that regulation of APP processing may involve the sequential activation of the enzymes MAPK kinase (MEK) and ERK. We provide evidence that the MAPK pathway is critically involved in regulation of APP processing by nerve growth factor (NGF), phorbol esters and A^-methyl-D-aspartate (NMDA). Western blot analysis of APPS demonstrated that the MEK inhibitor PD 98059 antagonized NGF stimulation of APPS production in a neuronal cell line. Moreover, inhibition of MEK blocked phorbol ester regulation of APPS in cortical neurons and Aβ release in cell lines. Finally, overexpression of a kinase-inactive MEK mutant inhibited both phorbol ester and NMDA receptor stimulation of APPS. Taken together, these data indicate that the MAPK pathway may be critically involved in regulating APP processing. As signal transduction is intimately associated with amyloid burden in Alzheimer's disease, understanding the critical effector systems responsible for this mismetabolism of APP is necessary in order for effective treatment strategies to be generated.

Thesis/Dissertation

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2009-05-28

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http://hdl.handle.net/2429/8415

Neuroscience

University of British Columbia

UBCV

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