The autoantigen La/SS-B: analysis of the expression of alternatively spliced La mRNA isoforms

Abstract.

The gene for the nuclear autoantigen La/SS-B encodes two La mRNA isoforms. In order to study the function and expression of both La mRNA forms, an in situ hybridization procedure was developed allowing the selective identification of either exon 1 or exon 1′. For this purpose, digoxigenin-labeled exon-specific sense and anti-sense probes were prepared by in vitro transcription from plasmids that contained the respective exon sequence. Detection of the probes was carried out by using rhodamine-conjugated anti-digoxigenin antibody and confocal laser scanning microscopy. Both La mRNAs were found in the cytoplasm of endothelial cells but not in smooth muscle cells. In addition to the in situ technique, an assay system was established allowing the expression ratio of the two mRNA forms to be determined. The estimation was based on the amplification of exon 1 and 1′ La cDNAs in parallel by using a three primer polymerase chain reaction. The ratio of the exon 1 to exon 1′ La mRNA forms was determined to be about 5:1 in liver tissue and endothelial cells. The data support the conclusion that both La mRNA forms represent finally processed cytoplasmic mRNAs that are up- or downregulated in parallel.