Glial fibrillary acidic protein: from intermediate filament assembly and gliosis to neurobiomarker

Abstract

Glial fibrillary acidic protein (GFAP) is an intermediate filament (IF) III protein uniquely found in astrocytes in the central nervous system (CNS), non-myelinating Schwann cells in the peripheral nervous system (PNS), and enteric glial cells. GFAP mRNA expression is regulated by several nuclear-receptor hormones, growth factors, and lipopolysaccharides (LPSs). GFAP is also subject to numerous post-translational modifications (PTMs), while GFAP mutations result in protein deposits known as Rosenthal fibers in Alexander disease. GFAP gene activation and protein induction appear to play a critical role in astroglial cell activation (astrogliosis) following CNS injuries and neurodegeneration. Emerging evidence also suggests that, following traumatic brain and spinal cord injuries and stroke, GFAP and its breakdown products are rapidly released into biofluids, making them strong candidate biomarkers for such neurological disorders.

Figure 1. GFAP tissue specificity

(A) GFAP mRNA expression in human tissue and cells based on BioGPS database [4] (B) GFAP protein expression in various human and rat organs, tissues Human GFAP was extensively truncated to 48-38 kDa bands, likely due to postmortem proteolysis, while rat GFAP mainly exists in brain as 50-48 kDa form [5].

Figure 2. Glial Fibrillary Acidic Protein (GFAP) structure and assembly

(A) linear structure, functional domains and key modifications. (B) 3D-GFAP protein structure based on Ref. [13]; (C) Proposed GFAP dimer and tetramer assembly and oligomerization model, modified after Ref. [14].

Figure 3. Known GFAP isoforms and features

(Left) Description of key features of GFAP isoforms. (Right) Schematic comparison of key GFAP isoforms in linear models.

Figure 4. GFAP patterns in injured and activated glia cells in vitro and in vivo

(A) Rat GFAP staining of resting primary glia cells or Staurosporine (0.5 μM, 5 h)-injured glia or CNTF (200 nM for 24 h)- activated glial cells. (B) Naïve (control) rat cortex or 24 h after experimental TBI showing injured glia cells or 3 day after TBI showing activated glia cells. GFAP was stained red, while counterstain was DAPI for nuclear DNA (blue). (C) Anti-GFAP-immunblotting of control glia cell lysate shows intact GFAP of 50 kDa with two anti-GFAP antibodies. Upon induction of necrosis (induced by 10 μM A23187) or caspase-dominant apoptosis (induced by 5 mM EDTA) [51,112], core-directed antibody shows calpain-mediated 38 kDa limit GFAP-BDP after A23187 treatment, while C-terminal antibody detects two district caspase-mediated GFAP-BDPs of 44K and 20K after EDTA treatment (Results shown in both (A) and (B) are from a published study by us [5], but were not shown as representative examples in a primary publication).

Figure 5. Release of GFAP and GFAP-BDP into biofluid as acute CNS injury biomarker

(Left) List of neuro-diseases and disorders in which GFAP and/or GFAP-BDP is released into biofluid. (Right) Schematic showing how GFAP BDP is generated after TBI and how both GFAP-BDP and to a lesser extent intact GFAP are released into interstitial fluid (ISF)/extracellular fluid. From there, these proteins diffused into the subarachnoid CSF [113]. GFAP-BDP/GFAP then either drain directly into the veins (glymphatic pathway, dashed arrows) or continue to follow the CSF flow through the ventricles and then enter the circulation by diffusing through the BBB. Blood-based GFAP and GFAP-BDP can be detected as a biomarker, but they also can serve as autoantigen, triggering autoantibody response in a subset of patients.