Astrocytes play important roles in the development and maintenance of the central nervous system (CNS). To understand and manipulate astrocyte function, this project addresses transcriptional control of the astrocyte~specific gfa gene that encodes the intermediate filament protein, GFAP, or glial fibrillary acidic protein. Using transfection of astrocytoma cells with reporter gene constructs, multiple segments have been identified within the gfa promoter and upstream regions; the segments interact to control its expression. Site~directed mutagenesis is being used to pinpoint the critical specific sequences within these segments. This will be followed by isolation and study of the regulatory proteins acting at these sites. The activity of reporter constructs is also being studied in transgenic mice. A 2,000 base pair 5'~flanking fragment of the gfa gene was sufficient to drive expression of a beta~galactosidase reporter gene in astrocytes throughout the CNS. Deleting from this construct a gfa segment found unimportant for expression in cultured cells also produces expression exclusively in astrocytes, but activity is largely restricted to the cortex. These results indicate that astrocytes are heterogeneous in gene expression and that different regulatory regions of the gfa gene are utilized by different types of astrocytes. Projects are also underway to use the gfa regulatory sequence to express other genes of interest in astrocytes to study brain development and function and to produce models for human diseases. Genes currently being expressed include those encoding the amyloid precursor protein associated with Alzheimer~s disease, somatostatin,, TGF~beta1, and a putative dominant negative GFAP.
