On the sporadic Amyotrophic Lateral Sclerosis-Frontotemporal Dementia (sALS/FTD) clinical spectrum, the
aggregation and accumulation of disease-associated proteins such as TDP-43 is a notable neuropathological
hallmark, yet we know little about why this highly abnormal event might occur. Although disruptions in multiple
cellular processes have been implicated in ALS, 3 critical gaps in knowledge remain: 1) What triggers the
aggregation of wildtype proteins in sporadic disease? Is protein aggregation sufficient to drive pathology? 2)
What drives the cell-specific vulnerabilities and variable clinical manifestation from ALS to FTD? 3) How do
disease-associated alterations in protein homeostasis perturb communication in the tissue microenvironment?
Given that more than 95% of ALS arises sporadically, and that the mechanisms of sporadic disease remain
unknown, we will look beyond individual mutations, and establish a novel conceptual framework that examines
the cellular changes that occur during disease states. We posit that by focusing on why TDP-43 aggregation
occurs, especially in sporadic ALS, we will gain insights into pathogenic mechanisms underlying this
spectrum of disorders. Our central hypothesis is that there are physical changes at the cell and tissue
scale that initiate ALS/FTD. We propose that altered biophysical properties within cells (predominantly
altered molecular crowding), which are linked to mechanical perturbations to the tissue microenvironment
(stiffening, inflammation, edema causing osmotic stress), lead to age-dependent cellular dysfunction by
altering the dynamics of assembly, disassembly and transport of macromolecular protein machines. We will
test this hypothesis in cellular models, animal models, and patient tissue by (1) using novel tools to probe the
intracellular biophysical environment of cells; (2) integrating these findings using novel genomics technologies
applied to mouse models to study (i) how intracellular changes in crowding and extracellular changes in the
tissue microenvironment may drive pathogenesis in vivo, and (ii) how such perturbations disrupt cell-cell
communication in vulnerable regions of tissue; and (3) relating our findings to human disease by re-examining
these findings in the context of a clinically and neuropathologically deeply curated cohort of ALS/FTD patients.
These studies will allow us to address the following questions: 1) Why does abnormal protein aggregation
and accumulation occur in sporadic disease, and how might this contribute to disease pathogenesis; 2) Do
these alterations in protein homeostasis perturb intercellular communication in the tissue microenvironment;
and 3) What drives the cell type vulnerability that makes ALS/FTD unique? The proposed work will accomplish
the following: A) represent the first detailed survey of molecular crowding in neural cells; B) uncover whether a
causal link between intracellular crowding, protein aggregation, and neurodegeneration exists, C) establish
whether the impacts of intracellular crowding show cell type specific signatures including changes in protein-
protein interactions, and D) provide a new framework to explore therapeutic strategies for treating ALS/FTD.
