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J. Biol. Chem., Vol. 275, Issue 49, 38286-38295, December 8, 2000
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From the Biochemistry Department, The University of Queensland,
Queensland 4072, Australia
Heterogeneous nuclear ribonucleoprotein (hnRNP)
A2 binds a 21-nucleotide myelin basic protein mRNA response
element, the A2RE, and A2RE-like sequences in other localized
mRNAs, and is a trans-acting factor in oligodendrocyte
cytoplasmic RNA trafficking. Recombinant human hnRNPs A1 and A2 were
used in a biosensor to explore interactions with A2RE and the cognate
oligodeoxyribonucleotide. Both proteins have a single site that bound
oligonucleotides with markedly different sequences but did not bind in
the presence of heparin. Both also possess a second, specific site that
bound only A2RE and was unaffected by heparin. hnRNP A2 bound A2RE in
the latter site with a Kd near 50 nM,
whereas the Kd for hnRNP A1 was above 10 µM. UV cross-linking assays led to a similar conclusion.
Mutant A2RE sequences, that in earlier qualitative studies appeared not to bind hnRNP A2 or support RNA trafficking in oligodendrocytes, had
dissociation constants above 5 µM for this protein. The
two concatenated RNA recognition motifs (RRMs), but not the individual RRMs, mimicked the binding behavior of hnRNP A2. These data highlight the specificity of the interaction of A2RE with these hnRNPs and suggest that the sequence-specific A2RE-binding site on hnRNP A2 is
formed by both RRMs acting in cis.
Binding of an RNA Trafficking Response Element to Heterogeneous
Nuclear Ribonucleoproteins A1 and A2*
*
This work was supported by grants from the Australian
National Health and Medical Research Council (to R. S.) and from
the Australian Research Council (to R. S. and D. J. W.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
To whom correspondence should be addressed: Biochemistry Dept.,
University of Queensland, Qld 4072, Australia. Tel.: 61-7-3365-4627; Fax: 61-7-3365-4699; E-mail ross@biosci.uq.edu.au.
Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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