Immunoglobulin Structure, Stability and Aging

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DESCRIPTION (provided by applicant): Light chain amyloidosis (AL) is the most common systemic protein aggregation disease. It is not only debilitating and often fatal, but also difficult to diagnose, because of the variety of affected internal organs. Its molecular cause is a conformational change in specific immunoglobulin light chains (LCs), due to somatic mutations accumulated during the course of immune responses. Specific mutations cause the LCs to polymerize into fibrils that then deposit as plaques. The disease has no direct and effective treatment and the development of one is complicated by the lack of an animal model. This renewal application seeks to build upon the in vitro fibrillogenesis reactions, cellular models and an inhibitory peptide that were developed in the first grant period. These tools will be used to determine the molecular specificity inherent in LC fibrillogenesis reactions. Since we previously showed that some amyloidogenic LC are not secreted and form intracellular inclusion bodies, our first Aim is to test multiple amyloidogenic LCs and determine which sequences are compatible with secretion and which LCs form inclusion bodies. One type of intracellular aggregate, termed aggresome, is of particular interest, because it is likely fibrillar, based on its sensitivity to the inhibitory peptide. This survey of LCs is important in characterizing the intracellular and extracellular pathways of LC amyloid formation. Aim2 will pursue our observations that some LCs can form two distinct types of fibrils, and on the other hand, that LC fibrils can initiate polymerization of other, more distant proteins. A library of mutants will be used to determine the requirements for formation of fibrils from either the oxidized or the reduced LC. We will next test the hypothesis that the unfolding of proteins with Greek key fold (such as LC, p2m, TTR or SOD1) is constrained and thereby leads to a common conformation that can give rise to similar fibrils. This hypothesis will be tested in two ways: seeding of each of these proteins with fibrils of another, and determining the sensitivity of different fibrils to the inhibitory peptide. The similarity uncovered in vitro will then be applied to the intracellular environment by determining the degree of co-aggregation of Greek key proteins. Aim 3 is to use mice transgenic for one amyloidogenic LC and assess their suitability as an animal model for AL by manipulating them genetically and immunologically so as to form amyloid deposits. If successful, this model will be used to test the efficacy of the peptide that specifically blocks fibrillogenesis of Greek Key proteins as a potential treatment modality. Relevance: This project continues to investigate the molecular mechanisms that underlie light chain amyloidosis, a fatal protein aggregation disease where deposits of antibody light chains cause plaques and lead to dysfunction of vital organs.