Glial fibrillary acidic protein exhibits altered turnover kinetics in a mouse model of Alexander disease

Abstract

Edited by George N. DeMartinoMutations in the astrocyte-specific intermediate filament glial fibrillary acidic protein (GFAP) lead to the rare and fatal disorder, Alexander disease (AxD). A prominent feature of the disease is aberrant accumulation of GFAP. It has been proposed that this accumulation occurs because of an increase in gene transcription coupled with impaired proteasomal degradation, yet this hypothesis remains untested. We therefore sought to directly investigate GFAP turnover in a mouse model of AxD that is heterozygous for a disease-causing point mutation (Gfap R236H/؉ ) (and thus expresses both wild-type and mutant protein). Stable isotope labeling by amino acids in cell culture, using primary cortical astrocytes, indicated that the in vitro halflives of total GFAP in astrocytes from wild-type and mutant mice were similar at ϳ3-4 days. Surprisingly, results obtained with stable isotope labeling of mammals revealed that, in vivo, the half-life of GFAP in mutant mice (15.4 ؎ 0.5 days) was much shorter than that in wild-type mice (27.5 ؎ 1.6 days). These unexpected in vivo data are most consistent with a model in which synthesis and degradation are both increased. Our work reveals that an AxD-causing mutation alters GFAP turnover kinetics in vivo and provides an essential foundation for future studies aimed at preventing or reducing the accumulation of GFAP. In particular, these data suggest that elimination of GFAP might be possible and occurs more quickly than previously surmised.Alexander disease (AxD) 2 is a rare and often fatal human disease of the central nervous system caused by dominant mutations in the astrocyte intermediate filament glial fibrillary acidic protein (GFAP) (1). Most AxD patients have de novo mutations in GFAP (2), encoding for various missense mutations as well as small in-frame insertions and deletions. The hallmark feature of the pathology is the formation of aggregates, known as Rosenthal fibers (RFs), within the cell bodies and processes of astrocytes, along with variable degrees of white matter deficits. Although the genetic basis for AxD is clear, the mechanisms by which GFAP mutations lead to astrocyte dysfunction and the cascade of secondary effects on other cells in the central nervous system remain unresolved. Previous studies demonstrated that simple overexpression of wild-type GFAP to high levels induces the formation of RFs (3), and indeed increased levels of GFAP are consistently present in autopsy samples from patients with AxD (4 -8). Mouse models engineered to express mutations equivalent to common human mutations (9) illustrate a connection between GFAP levels and severity of disease, which has led to the concept of "GFAP toxicity."The excessive accumulation of GFAP and the formation of RFs in AxD presumably reflect a fundamental alteration in proteostasis. Proteostasis, or protein homeostasis, involves a vast network of pathways that control protein synthesis, folding, trafficking, aggregation, and degradation. Protein aggregation disorders such as Alzhei...

Alexander disease: an astrocytopathy that produces a leukodystrophy

“…In contrast, the half‐life of GFAP in mutant mice in vivo was much shorter than in control mice (∼ 15 days vs. 28 days, respectively). The faster turnover of GFAP in mutant mice in vivo , as the authors propose, may be because of an acceleration of both protein synthesis and protein degradation . It should be noted that the analysis of proteins in vivo was performed at 2 months of age, when astrocytes have already accumulated high levels of GFAP and many RFs.…”

“…Another contributing factor to GFAP protein levels is the turnover rate of the protein. In an interesting study of protein turnover, that of GFAP in the AxD KI mice in vivo and in primary astrocyte cultures from these mice was compared to GFAP turnover in control mice . The half‐life of GFAP protein in vitro was much shorter than in vivo and did not differ between KI and control astrocytes.…”

GFAP at 50

“…Yet at the same time there is an increase in autophagy ( Tang et al., 2008 ) and activation of caspases 3 and 6 ( Chen et al., 2011 , 2013 ). When the knockin mouse model was studied during adulthood using methods of isotope turnover, the half-life of GFAP was observed to be actually decreased ( Moody et al., 2017 ). Furthermore, recent studies on autopsy tissues from three patients found that the mutant form of the protein is not the majority of the GFAP present in the brain, suggesting that it is less stable than the wild type ( Heaven et al., 2019 ).…”

Neurochemical regulation of the expression and function of glial fibrillary acidic protein in astrocytes

“…Alexander disease is a rare, fatal neurologic disorder characterized by white‐matter degeneration and cytoplasmic inclusions in astrocytes known as Rosenthal fibers, a product of heterozygous mutations in the GFAP gene (Moody, Barrett‐Wilt, Sussman, & Messing, ). The cytoplasmic accumulations of GFAP and Rosenthal fibers in astrocytes cause proteasome inhibition, stress kinase activation, mechanistic target of rapamycin activation, loss of glutamate‐ and potassium‐buffering capacity, loss of astrocyte coupling, and changes in cell morphology (Olabarria & Goldman, ).…”