Arthur L Horwich MD

Sterling Professor of Genetics and Professor of Pediatrics; and Investigator HHMI

Research Interests

Chaperones in protein folding; ALS (Lou Gehrig's Disease)

Current Projects

Mouse and mammalian cell projects - designed to analyze the order of pathologic events in misfolding induced ALS at a morphology level, the biochemical abnormalities at a genomic/proteomic level, and to try to mimic the progression using cell culture models, including use of ES cells differentiated into motor neurons.

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Research Summary

We are studying protein folding and misfolding in the cell, along two major lines: 1) Examining how “chaperonin” ring structures mediate ATP-dependent folding of a large number of newly-translated proteins. Chaperonins bind polypeptides through exposed hydrophobic surfaces, then carry out productive folding in an encapsulated hydrophilic chamber. We are studying what happens to “substrate” proteins during this process. 2) Studying a misfolding disease, SOD1-linked ALS (Lou Gehrig’s Disease), using mouse and worm models, employing both genetics and biochemistry to understand why SOD misfolding causes motor neuron damage.

Extensive Research Description

Studies of the past decade have shown that many diseases ofneurodegeneration are the result of protein misfolding, and we havebegun to seek an understanding of the mechanism of such degeneration. We have focused on misfolding caused by mutant forms of theanti-oxidant cytosolic enzyme SOD1 (superoxide dismutase), that producean inherited form of ALS (Lou Gehrig’s disease), with progressive,fatal motor neuron dysfunction. We are using both C.elegans and miceexpressing mutant G85R SOD1, a version of SOD1 unable to reach the native state, studying the transgenic animals with genetic, morphologic, and biochemical approaches, toinvestigate how the mutant SOD1 selectively produces motor neuron dysfunction.

Mouse studies.
We have identified that there is a progression of misbehavior of mutant SOD1 protein itself in the spinal cord of transgenic animals. Initially it is soluble and associated to a significant extent with the abundant cytosolic chaperone Hsc70 (by contrast the wild-type protein does not form such association, presumably because it occupies the native state). Subsequently the mutant SOD1 protein begins to form both soluble oligomers (observable by gel filtration chromatography) and insoluble aggregates and at this point an additional molecular chaperone, Hsp110 becomes associated with the soluble form. At this point the animal exhibits mild lower extremity symptoms (weakness or pulling in when held by the tail) which progress over the next two to three months to lethal paralysis, usually commencing in the lower extremities. We are currently seeking to understand ultrastructural correlates of this progression using EM, analyzing motor neuron cell bodies in the spinal cord, sciatic nerve axons, and neuromuscular junctions. Where do abnormalities first present? What is the progression? Can we identify biochemical correlates using immunohistochemistry or laser capture microdissection of motor neurons coupled with genomics (RNA-seq e.g.) or proteomics? At another level, what are the electrophysiologic correlates? Finally, can we modify disease by crossing in various mutant genes?



Cellular models.
We have produced mouse embryo fibroblast lines from our transgenic mice and are studying macromolecular processes in them, inspecting for mutant-specific behavior. We have also produced ES cell lines from the animals and are beginning to differentiate motor neurons from them, with a view to having a supply of motor neurons with which to carry out additional phenotypic studies. In short, can the ALS phenotype or some portion of it be recapitulated in culture?

Overall, we are hoping that between worm, cell culture, and mammalian systems we will be able to deduce the mechanism of SOD1-linked ALS and potentially design preventive steps. Understandings of SOD1-linked disease may extend more generally to other forms of ALS, or possibly to other neurodegenerative diseases.

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Selected Publications