Glial fibrillary acidic protein filaments can tolerate the incorporation of assembly-compromised GFAP-delta, but with consequences for filament organization and alphaB-crystallin association.
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 (approximately 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 (approximately 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 alphaB-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.
Perng
Ming-Der
MD
School of Biological and Biomedical Sciences, The University of Durham, Durham DH1 3LE, United Kingdom.
Wen
Shu-Fang
SF
Gibbon
Terry
T
Middeldorp
Jinte
J
Sluijs
Jacqueline
J
Hol
Elly M
EM
Quinlan
Roy A
RA
D038261:Alexander Disease / Q000235:genetics* / Q000378:metabolism; D001253:Astrocytes / Q000378:metabolism; D002460:Cell Line; D045744:Cell Line, Tumor; D005904:Glial Fibrillary Acidic Protein / Q000737:chemistry*; D006801:Humans; D048670:MAP Kinase Kinase 4 / Q000378:metabolism; D008954:Models, Biological; D009154:Mutation; D010766:Phosphorylation; D011485:Protein Binding; D020033:Protein Isoforms; D013116:Spinal Cord / Q000378:metabolism; D014162:Transfection; D038203:alpha-Crystallin B Chain / Q000737:chemistry*
D005904:Glial Fibrillary Acidic Protein; D020033:Protein Isoforms; D038203:alpha-Crystallin B Chain; D048670:MAP Kinase Kinase 4