Mutant PFN1 causes ALS phenotypes and progressive motor neuron degeneration in mice by a gain of toxicity

Abstract

Mutations in the profilin 1 (PFN1) gene cause amyotrophic lateral sclerosis (ALS), a neurodegenerative disease caused by the loss of motor neurons leading to paralysis and eventually death. PFN1 is a small actin-binding protein that promotes formin-based actin polymerization and regulates numerous cellular functions, but how the mutations in PFN1 cause ALS is unclear. To investigate this problem, we have generated transgenic mice expressing either the ALSassociated mutant (C71G) or wild-type protein. Here, we report that mice expressing the mutant, but not the wild-type, protein had relentless progression of motor neuron loss with concomitant progressive muscle weakness ending in paralysis and death. Furthermore, mutant, but not wild-type, PFN1 forms insoluble aggregates, disrupts cytoskeletal structure, and elevates ubiquitin and p62/ SQSTM levels in motor neurons. Unexpectedly, the acceleration of motor neuron degeneration precedes the accumulation of mutant PFN1 aggregates. These results suggest that although mutant PFN1 aggregation may contribute to neurodegeneration, it does not trigger its onset. Importantly, these experiments establish a progressive disease model that can contribute toward identifying the mechanisms of ALS pathogenesis and the development of therapeutic treatments.A LS is a neurodegenerative disease that causes a relentless progressive loss of motor neurons, leading to progressive weakness that ends in paralysis and death (1). Mutations in the PFN1 gene have been identified as a genetic cause for ALS (2, 3). PFN1 is a major regulator of actin polymerization through its ability to bind actin-ADP monomers and promote the conversion of actin-ADP to actin-ATP, through transporting the actin-ATP monomers and through its interactions within formins present at the growing end of actin filaments. Additionally PFN1 binds to phosphoinositides and a large network of proteins with poly-Lproline stretches. Through these binding interactions, PFN1 regulates several cellular functions including actin dynamics, membrane trafficking, neuronal synaptic structure and activity, small GTPase signaling, and others (4). Despite our understanding of its function, how PFN1 mutations cause motor neuron degeneration remains elusive. Some evidence implicates a loss of function in the mutants. The ALS-associated mutations cause structural instability and accumulate in cells at lower levels than the wild-type protein (5). Mutant PFN1 binds less efficiently to actin than the wild-type protein, suggesting that the mutations compromise PFN1 function (3). The severest mutations also are incapable of compensating for loss-of-function PFN1 mutation in yeast (6). Other evidence supports a gain of function. Expression of mutant, but not wild-type, PFN1 inhibits filamentous actin formation and impairs growth cone function and neurite growth (3). Furthermore, PFN1 mutants have been shown to alter stress granule dynamics in cultured mammalian cells and form cellular aggregates that may contain other proteins contributing to path...

Profilin2a‐phosphorylation as a regulatory mechanism for actin dynamics

“…53 Eight different mutations of profilin1 have been associated with familiar forms of the motoneuron disease ALS. [54][55][56] Mutated sites localize close to the actin or PLP binding sites comparable to the phospho-sites in pro-filin2a (Table S1). Similar to our profilin2a mutant S129D, the profilin1 G118V displayed a marked decrease in G-actin binding.…”

ALS-causing mutations in profilin-1 alter its conformational dynamics: A computational approach to explain propensity for aggregation

“…Eight mutations in PFN1 were found to be associated with fALS 1–5 . All of the PFN1 mutations, except E117G, are specific to a subset of fALS patients 6–8 , as are the SOD1 and TDP-43 mutations.…”

ATP emerged to induce protein folding, inhibit aggregation and increase stability

“…1A). C71G-PFN1 characteristic of severe aggregation causes ALS by gain of toxicity (20). Previously we showed that C71G-PFN1 co-existed in equilibrium between the unfolded and folded states (21), as characterized by two set of HSQC peaks ( Fig.…”