HNRNP A1 BINDS PROMISCUOUSLY TO OLIGORIBONUCLEOTIDES - UTILIZATION OFRANDOM AND HOMO-OLIGONUCLEOTIDES TO DISCRIMINATE SEQUENCE FROM BASE-SPECIFIC BINDING

To understand the range of possible and probable Al functions in pre-m RNA biogenesis, it is important that we quantify the relative ability (or inability) of Al to bind high affinity RNA target sequences and/or structures, Using a fluorescence competition assay we have determined apparent binding affinities for a wide range of 20mer oligos containi ng putative and possible Al targets including the high affinity 'winne r' sequence identified by selection/amplification [Burd,C,G and Dreyfu ss,G. (1994) EMBO J. 13, 1197-1204], AUUUA sequences found in 3'-UTRs of labile mRNAs, 5'- and 3'-splice sites and telomeric sequences, With the exception of a 20mer 'winner' sequence, all other 20mers examined bind Al with a narrow, -10-fold range of affinities extending from 3. 2 x 10(6) to 4.2 x 10(7) M(-1). Studies with homo-oligomers suggest th is range reflects nucleotide base rather than sequence specificity and hence, it was possible to predict reasonably accurate affinities for all other 20mers examined except for the 'winner', whose unusually hig h affinity of 4.0 x 10(8) M(-1) results from a unique higher order str ucture and sequence, Since there is no known physiological role for th e 'winner' 20mer sequence, these data suggest Al generally binds indis criminately to all available pre-mRNA sequences, Both the large abunda nce of Al in vivo and its binding properties are thus consistent with it playing a structural role in pre-mRNA biogenesis.