Redistribution of GFAP and αB-crystallin after thermal stress in C6 glioma cell line

Abstract

SummarySome intermediate filament (IF) proteins expressed in the development of glia include nestin, vimentin, and glial fibrillary acidic protein (GFAP). However, GFAP is the major intermediate filament protein of mature astrocytes. To determine the organization of GFAP in glial cells, rat GFAP cDNA tagged with enhanced green fluorescent protein (EGFP) was transfected into the rat C6 glioma cell line. After selection, two stable C6-EGFP-GFAP cell lines were established. Stable C6-EGFP-GFAP cell lines with or without heat shock treatment were analyzed by immunocytochemistry, electron microscopy, and Western blot analysis. In the transient transfection study, EGFP-GFAP transiently expressed in C6 cells formed punctate aggregations in the cytoplasm right after transfection, but gradually a filamentous structure of EGFP-GFAP was observed. The protein level of nestin in the C6-EGFP-GFAP stable clone was similar to that in the pEGFP-C1 transfected C6 stable clones and non-transfected C6 cells, whereas the level of vimentin was reduced in Western blotting. Interestingly, the expression level of small heat shock protein aB-crystallin in C6-EGFP-GFAP cells was also enhanced after transfection. Immunostaining patterns of C6-EGFP-GFAP cells showed that GFAP was dispersed as a fine filamentous structure. However, after heat shock treatment, GFAP formed IF bundles in C6-EGFP-GFAP cells. In the meantime, aB-crystallin also colocalized with IF bundles of GFAP in C6-EGFP-GFAP cells. The heat-induced GFAP reorganization we found suggested that small heat shock protein aB-crystallin may play a functional role regulating the cytoarchitecture of GFAP.

Properties of astrocytes cultured from GFAP over-expressing and GFAP mutant mice

“…αB-crystallin is well known as a binding partner for type III intermediate filaments, including vimentin [37], desmin [38], and GFAP [39]. Studies in both C6 cells and in primary astrocytes demonstrate that αB-crystallin can influence GFAP integration into the cytoskeletal network, both by regulating normal assembly and by assisting in recovery from stress [37,40]. …”

Phosphorylation of Ser45 and Ser59 of αB‐crystallin and p38/extracellular regulated kinase activity determine αB‐crystallin‐mediated protection of rat brain astrocytes from C2‐ceramide‐ and staurosporine‐induced cell death

“…Several reports indicated that a-crystallin may interact with different cellular components, like cytoplasmic membrane, cytoskeletal elements, and mediators of cell death (Cobb and Petrash 2000;Kamradt et al 2001;Liu et al 2004;Mao et al 2004;Li et al 2005). As a result of these interactions, a-crystallin maintains the stability of the cellular structures and inhibits the activity of caspases to reduce the stress-induced damage (Nicholl and Quinlan 1994;Wang and Spector 1996;Djabali et al 1997;Tseng et al 2006;Xi et al 2006).…”

“…As a result of these interactions, α‐crystallin maintains the stability of the cellular structures and inhibits the activity of caspases to reduce the stress‐induced damage (Nicholl and Quinlan 1994; Wang and Spector 1996; Djabali et al. 1997; Tseng et al. 2006; Xi et al.…”

Cortical dysplasia with prominent Rosenthal fiber formation in a case of intractable pediatric epilepsy

“…The GFAP gene is first expressed as astrocytes mature and in the adult is strongly up-regulated in response to central nervous system damage. It has been suggested that RFs may represent a metabolite of upregulation of GFAP in response to stress [13,18,23]. The abundant RFs seen in association with our case of CD may represent a dysregulation of α-B-crystallin aggregating in association with GFAP or be secondary to the severity of the CD that produces a dys/up-regulation of GFAP to levels sufficient to induce RFs and localized leukodystrophy, in some cases, due to astrocytic dysfunction [18].…”