HUMAN ADENOVIRUS ENCODES 2 PROTEINS WHICH HAVE OPPOSITE EFFECTS ON ACCUMULATION OF ALTERNATIVELY SPLICED MESSENGER-RNAS

All mRNAs expressed from the adenovirus major late transcription unit have a common, 201-nucleotide-long 5' leader sequence, which consists of three short exons (the tripartite leader). This leader has two vari ants, either with or without the i-leader exon, which, when present, i s spliced between the second and the third exons of the tripartite lea der. Previous studies have shown that adenovirus early region 4 (E4) e ncodes two proteins, E4 open reading frame 3 (E4-ORF3) and E4-ORF6, wh ich are required for efficient expression of mRNAs from the major late transcription unit. These two E4 proteins appear to have redundant ac tivities, and expression of one has been shown to be sufficient for ef ficient major late mRNA accumulation during a lytic virus infection. I n this report, we provide evidence that E4-ORF3 and E4-ORF6 both regul ate major late mRNA accumulation by stimulating constitutive splicing. Moreover, we show that the two proteins have different effects on acc umulation of alternatively spliced tripartite leader exons. In a DNA t ransfection assay, E4-ORF3 was shown to facilitate i-leader exon inclu sion, while E4-ORF6 preferentially favored i-leader exon skipping. In addition, E4-ORF3 and E4-ORF6 had the same effects on accumulation of alternatively spliced chimeric 13-globin transcripts. This finding sug gests that the activities of the two proteins may be of more general r elevance and not restricted to splicing of major late tripartite leade r-containing pre-mRNAs. Interestingly, E4-ORF6 expression was also sho wn to stimulate i-leader exon skipping during a lytic virus infection.