Distribution of snRNPs, splicing factor SC-35 and actin in interphase nuclei: immunocytochemical evidence for differential distribution during changes in functional states

Abstract

Small nuclear ribonucleoproteins (snRNPs) play an integral role in the processing of pre-mRNA in eukaryotic nuclei. snRNPs often occur in a speckled intranuclear distribution, together with the non-snRNP splicing factor SC-35. snRNPs have also been shown to be associated with actin in the nuclear matrix, suggesting that both actin and snRNPs may be involved in the processing and transport of transcripts. The work reported here was undertaken to compare the spatial relationship of snRNPs, SC-35, and intranuclear actin in neuronal and non-neuronal cell types. In undifferentiated PC12 cells and in non-neuronal cells growing in association with dorsal root ganglion neurons, confocal immunocytochemistry revealed a typical, speckled distribution of snRNP aggregates, which colocalized with the SC-35 splicing factor. In contrast, a unique snRNP distribution was observed in dorsal root ganglion neurons in vitro and in PC12 cells differentiated by nerve growth factor. In nuclei of these cells, snRNPs were predominantly located at the periphery where they formed a spherical shell apposed to the nuclear envelope. Ultrastructural immunogold labelling of snRNPs in dorsal root ganglion neurons in vitro confirmed this distribution. In contrast, SC-35 remained distributed in a speckled pattern throughout nuclei of dorsal root ganglion neurons and PC12 cells, even in cases where snRNPs were almost exclusively positioned at the nuclear periphery. In non-neuronal cells in dorsal root ganglion cultures and in undifferentiated PC12 cells, snRNP aggregates were frequently associated with actin aggregates, as determined by Nearest Neighbor Analyses. In PC12 cells, this spatial relationship was altered during nerve growth factor-induced differentiation, prior to the time at which these cells showed morphological evidence of differentiation. Specifically, Nearest Neighbor Analyses between snRNP and actin aggregates in PC12 cells exposed to nerve growth factor for 4 hours revealed that snRNP and actin aggregates exhibited a closer association than in undifferentiated cells. These results suggest that sites of pre-mRNA processing and transcription may differ between cell types, and that the functions of snRNPs and actin within interphase nuclei may be related. The results also indicate that the distribution of snRNPs is dynamic and that it may depend upon the functional state of the cell as well as upon its state of differentiation.