Differential mRNA localization in astroglial cells in culture.

Abstract

Messenger RNA (mRNA) targeting to specific subcellular domains has been studied extensively in many cell types, and there is increasing evidence suggesting that mRNA sorting also occurs in astrocytes. As a step toward developing strategies to evaluate the signals that govern mRNA sorting in astrocytes, the authors studied the subcellular distribution of several representative mRNAs, poly(A) RNA and ribosomal RNA, in process-bearing (type-2) astroglial cells in culture. Nonradioactive in situ hybridization analysis revealed a gradual increase in the expression of glial fibrillary acidic protein (GFAP) mRNA as type-2 astrocytes differentiated in culture. In mature cells, labeling was present in both cell bodies and processes. GFAP mRNA labeling was granular in nature and was particularly concentrated at branch points and at the tips of the processes. Unlike GFAP mRNA, vimentin, beta-tubulin, and beta- and gamma-actin mRNAs were mainly confined to the cell bodies, with only occasional labeling seen in the processes. Nonradioactive and radioactive in situ hybridization analysis of poly(A) and ribosomal RNA, respectively, revealed labeling in cell bodies and processes of immature and differentiated astrocytes. Treatment with nocodazole, a microtubule depolymerizing agent, resulted in a substantial reduction of GFAP mRNA labeling in the processes, whereas treatment with cytochalasin D, a microfilament-disrupting agent, did not alter GFAP mRNA distribution. The results indicate that cultured type-2 astrocytes have the capacity to sort mRNAs to different subcellular domains and that the localization of GFAP mRNA to astrocyte processes requires intact microtubules.