Glial Fibrillary Acidic Protein Filaments Can Tolerate the Incorporation of Assembly-compromised GFAP-delta, but with Consequences for Filament Organization and alpha B-Crystallin Association

[[abstract]]The glial fibrillary acidic protein (GFAP) gene is alternatively spliced to give GFAP-alpha, the most abundant isoform, and seven other differentially expressed transcripts including GFAP-delta. GFAP-delta has an altered C-terminal domain that renders it incapable of self-assembly in vitro. When titrated with GFAP-alpha, assembly was restored providing GFAP-delta levels were kept low (similar to 10%). In a range of immortalized and transformed astrocyte derived cell lines and human spinal cord, we show that GFAP-delta is naturally part of the endogenous intermediate filaments, although levels were low (similar to 10%). This suggests that GFAP filaments can naturally accommodate a small proportion of assembly-compromised partners. Indeed, two other assembly-compromised GFAP constructs, namely enhanced green fluorescent protein (eGFP)-tagged GFAP and the Alexander disease-causing GFAP mutant, R416W GFAP both showed similar in vitro assembly characteristics to GFAP-delta and could also be incorporated into endogenous filament networks in transfected cells, providing expression levels were kept low. Another common feature was the increased association of alpha B-crystallin with the intermediate filament fraction of transfected cells. These studies suggest that the major physiological role of the assembly-compromised GFAP-delta splice variant is as a modulator of the GFAP filament surface, effecting changes in both protein - and filament-filament associations as well as Jnk phosphorylation.[[fileno]]2050150010001[[department]]生科