MCDB Home > Faculty > John Lew > Current Research

Our laboratory has had a long standing interest in protein kinases and signal transduction, and recently has focused on the molecular mechanisms of Alzheimer's Disease and related dementias. We employ a multidisciplinary approach combining the methods of protein purification, molecular cloning, genetic engineering, protein chemistry, and kinetic and biophysical analysis to investigate the cellular mechanisms that lead to dementia, on both a molecular biological and detailed structure/mechanistic level. The research in our laboratory falls into two main projects:

Novel pathways of neuronal signal transduction: regulation of cdk5/p35

The cyclin-dependent kinases (cdks) function specifically in the regulation of cellular division. In mammals, cdc2/cyclin B is critical for G₂/M-phase transition, while cdk2/cyclin E and cdk2/cyclin A control passage through G₁/S-phase and S-phase of the cell cycle, respectively. The discovery of a novel member of the cdk family (cdk5/p35) has challenged this dogma by its demonstrated specific expression in post-mitotic neurons. In these cells, cdk5/p35 is essential for normal cytoskeletal dynamics, neurite outgrowth, and brain cortex development. However, endogenous proteolysis of its regulatory subunit, p35, to a p25 species is associated with neuronal apoptosis and is correlated with the progression of Alzheimer's Disease.

What are the physiological signals that trigger aberrant conversion of p35 to p25, and what is the mechanism by which cdk5/p25, as opposed to cdk5/p35, may induce apoptosis and neural degeneration? The goal of pursuing these questions is to understand the molecular basis of Alzheimer's Disease and related dementias, and to identify potential strategies for therapeutic intervention. Our laboratory is using protein purification and molecular cloning techniques to identify and characterize differences in the actions of cdk5/p35 and cdk5/p25, including identification of upstream signaling components as well as differences in their mechanism of action with respect to downstream physiological targets. The elucidation of novel regulators and their mechanisms of action will have a broad impact on our understanding of neuronal signal transduction and the relationship between normal cellular function and the molecular events that may trigger the onset of neurological disease.

Structure, function & mechanism of action of tau

Alzheimer's Disease is characterized by the appearance of the extracellular beta-amyloid senile plaques and the intracellular paired helical filaments (PHF). The major component of the PHFs is the microtubule associated protein, tau, which is abnormally hyperphosphorylated in Alzheimer's Disease but not in normal brain tissue. The abnormal hyperphosphorylation of PHF-tau is catalyzed to a large extent by cdk5/p25, as well as by other kinases; hence, a possible mechanism for the deleterious consequences of endogenous p25. In addition, genetic analysis has shown that naturally occurring mutations in the tau protein is the cause other dementias (eg. FTDP-17) that are highly similar to Alzheimer's Disease.

What are the structural and functional consequences of these mutations, and of phosphorylation of tau by cdk5/p25 and other kinases that potentially induce formation of PHFs? Furthermore, what is the molecular mechanism of the resulting cytotoxicity? Our laboratory is using a variety of structural and biophysical techniques, such as fluorescence spectroscopy, circular dichroism, NMR, and stopped-flow kinetic analysis, to understand the the role of structural determinants in tau that are crucial to promote either normal microtubule dynamics and cell function or, alternatively, aggregration into PHFs leading to neural degeneration and apoptosis. Our overall goal is to elucidate on a structural and mechanistic level the role of tau in neuropathology versus normal cell function, with the prospect of identifying relevant strategies for therapeutic intervention.