UNUSUAL SPLICE SITES IN THE E1A-E1B COTRANSCRIPTS SYNTHESIZED IN ADENOVIRUS TYPE 40-INFECTED A549 CELLS

The adenovirus El DNA region consists of two transcription units, E1A and E1B. In this paper we report that the E1A-E1B cotranscripts contai ning sequences of both the E1A and E1B regions are synthesized during adenovirus type 40 (Ad40) infection of A549 cells. Cytoplasmic RNA was isolated from Ad40-infected A549 cells at 24, 72, and 100h post infec tion (p.i.). The complementary (c) DNA was synthesized by reverse tran scription using an oligo-dT primer and then amplified by the polymeras e chain reaction (PCR) using primers derived from the E1A and E1B regi ons. The cDNAs thus amplified were sequenced either directly or after cloning into bacteriophage M13 vectors. Analysis of cDNA indicated tha t the E1A-E1B cotranscripts are synthesized at 72h p.i., but not at 24 or 100h p.i. Nucleotide sequences of three cDNAs of the E1A-E1B cotra nscripts indicated that the cotranscripts originate from the E1A promo ter and lack sequences for both the E1A poly(A) site and E1B cap site. The splices create open reading frames for E1A-E1B fused polypeptides around the E1A-E1B junctions in these mRNAs. Most interestingly, the sequence analysis showed that the 5' and 3' splice junctions in the tw o E1A-E1B cotranscripts do not conform to the splice consensus GT-AG r ule. Our results thus suggest that factor(s) which lead to unsual spli cing in the E1 mRNAs are present in Ad40-infected A549 cells.