Michael S. Wolfe, PH.D.
|Title||Professor of Neurology|
|Institution||Brigham and Women's Hospital|
|Address||Brigham & Women's Hospital|
Ctr for Neurologic Disease
77 Avenue Louis Pasteur
Boston MA 02115
The Wolfe lab studies intramembrane proteases that play critical roles both in normal biology and in human disease. The last place in the cell to expect hydrolysis is within the hydrophobic environment of the lipid bilayer. Nevertheless, a number of multi-pass membrane proteins appear to carry out this seemingly paradoxical process (Wolfe and Kopan, Science, 2004; see publications list for references in this section). Such proteases cut within the transmembrane region of their respective substrates, and consistent with this observation, these proteases contain putative catalytic residues located within transmembrane domains.
The specific focus of the lab has been on the chemistry and biology of gamma-secretase. This protease is critical to the pathogenesis of Alzheimer's disease (Esler and Wolfe, Science, 2001) and to cell differentiation during embryonic development. Small organic inhibitors were developed and used as tools to characterize and identify gamma-secretase (Wolfe et al., J Med Chem, 1998; Wolfe et al., Biochemistry, 1999a). Findings from the lab implicate a multi-pass membrane protein called presenilin as the catalytic component of a larger gamma-secretase complex (Wolfe et al., Nature, 1999; Esler et al., Nat Cell Biol, 2000). Missense mutations in presenilin cause hereditary Alzheimer's disease, and these mutations specifically affect gamma-secretase activity.
The lab found that presenilin and a presenilin-associated protein called nicastrin copurify with gamma-secretase activity from an immobilized inhibitor, evidence that nicastrin is also a member of the protease complex (Esler et al., Proc Natl Acad Sci USA, 2002). Moreover, a gamma-secretase substrate also copurified, suggesting an initial substrate docking site on the protease complex distinct from the active site. Helical peptides designed to interact with this docking site can potently inhibit gamma-secretase activity both in cell-free and cell-based assays (Das et al., J Am Chem Soc, 2003; Kornilova et al., Proc Natl Acad Sci USA, 2005). Most recently, the lab determined the stoichiometry of the active gamma-secretase complex, which had been unknown (and, with respect to presenilin, controversial). The four essential components (presenilin, nicastrin, Aph-1 and Pen-2) are each represented only once per complex (Sato et al., J Biol Chem, 2007).
The lab has also discovered a nucleotide binding site on the gamma-secretase complex. Small organic molecules that interact with this site can selectively block gamma-secretase proteolysis of the amyloid beta-protein precursor (APP), critical to the pathogenesis of Alzheimer’s disease, without affecting proteolysis of an alternative substrate, the Notch receptor. Notch signaling, critical in many cell differentiation events, requires proteolysis by gamma-secretase (De Strooper et al., Nature, 1999), and blocking Notch signaling with gamma-secretase inhibitors causes severe toxicity in mice. The finding that compounds can selectively block the cleavage of APP without affecting that of Notch (Fraering et al., J Biol Chem, 2005) has revived this protease as a therapeutic target.
The Wolfe lab has also been actively investigating the structure, mechanism, and inhibition of other intramembrane proteases, such as the serine protease Rhomboid (Urban and Wolfe, Proc Natl Acad Sci USA, 2005) and the presenilin homolog signal peptide peptidase (Sato et al., Biochemistry, 2006; Narayanan et al., J Biol Chem, 2007), both of which are highly conserved across evolution and play critical roles in biology. The goal is to establish common biochemical principles and strategies for designing inhibitors for this new family of membrane-embedded enzymes.
The lab has also begun to combine chemistry and biology toward the study of another factor critical to the pathogenesis of dementias: the microtubule-associated protein tau. Filaments of tau are a common feature in a variety of different neurodegenerative diseases, including Alzheimer’s disease. Mutations in the gene encoding this protein are associated with dominant, familial forms of frontotemporal dementia, and many of these mutations alter pre-mRNA splicing to increase inclusion of exon 10. The Wolfe lab has recently validated the in vivo existence of a hypothetical stem loop at the end of exon 10, where many of the dementia-associated mutations occur (Donahue et al., J Biol Chem, 2006). These mutations destabilize this RNA stem loop structure, allowing more ready access to splicing factors. High-throughput screening has led to identification of small molecules that interact with and stabilize this structure (Donahue et al., J Biomol Screen, 2007), and efforts are ongoing to improve the potency and selectivity of these agents to provide new tools for chemical biology as well as new prototype therapeutics. In addition, the lab has studied alternative splicing of the beta-site APP-cleaving enzyme 1 (BACE1; beta-secretase), determining that alternative splice isoforms are catalytically inactive and that shunting BACE1 down these alternative pathways with antisense oligonucleotides effectively lowers amyloid beta-protein production in cells (Mowrer and Wolfe, J Biol Chem, 2008). Thus, modulation of BACE1 alternative splicing represents a new strategy for developing disease-modifying therapies for Alzheimer’s disease.
Dr. Wolfe is highly committed to graduate-level teaching at HMS. In 2003, he co-directed and taught a new quarter course, Neurobiology 300, “Biochemistry and Biology of Neurodegenerative Diseases”, a 2-hour-per-week class with some didactic presentation but primarily discussion of seminal and current research articles in the field. In 2005 and 2007, he directed and taught this course in its entirety. Each time it has been offered, the course has been very popular, received high praise from the students. Dr. Wolfe has also been involved in leading a discussion section of BCMP 201, “Proteins: Structure, Function, and Catalysis” from 2002-5; in 2006-8, he presented lectures on enzyme mechanisms and inhibition for this course. In 2004, he has also led a discussion section for Med Sci 300, the Conduct of Science, an ethics course required of all DMS graduate students.
Construction of a Tau Minigene and Preliminary Analysis of Its Splicing
Summer, 06/11/07 - 08/03/07
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