Aprimary limitation in the development and use of screens to identify factors that regulate mammalian pre-mRNA splicing has been the development of sensitive reporter assays. Alternative splicing typically involves relatively small ( < 10-fold) changes in isoform ratios. Therefore, reporter constructs designed to allow direct analysis of isoform expression historically have at most a10-fold window of discrimination between apositive signal and background. Here we describe the design and application of areporter cell line that makes use of the phenomenon of transcriptional synergy to amplify the detection of changes in splicing, such that athree-to five-fold change in splicing pattern is observed as a30-to 50-fold change in GFP expression. Using this cell line we have identified two small molecules, from alibrary of approximately 300 synthetic compounds, that can induce partial repression of av ariable exon from the CD45 gene. We propose that the concept of transcription-based amplification of signal will allow the development of true high-throughput screening approaches to identify effectors of mammalian alternative splicing.

The present work was planned to study the effect of temperature on the vascular responses in the isolated perfused ear of the rabbit. To this end the rate of inflow was recorded and the net outward filtration of fluid into the interstitial spaces of the ear was measured. The work was partly prompted by the observation that, although this preparation has been used for over forty years

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