We have investigated the RNA binding specificity of Hel-Nl, a human neuron-specific RNA-binding protein, which contains three RNA recognition motifs. Hel-Nl is a human homolog of Drosophila melanogaster elav, which plays a vital role in the development of neurons. A random RNA selection procedure revealed that Hel-Nl prefers to bind RNAs containing short stretches of uridylates similar to those found in the 3' untranslated regions (3' UTRs) of oncoprotein and cytokine mRNAs such as c-myc, c-fos, and granulocyte macrophage colony-stimulating factor. Direct binding studies demonstrated that Hel-Nl bound and formed multimers with c-myc 3' UTR mRNA and required, as a minimum, a specific 29-nucleotide stretch containing AUUUG, AUUUA, and GUUUUU. Deletion analysis demonstrated that a fragment of Hel-Nl containing 87 amino acids, encompassing the third RNA recognition motif, forms an RNA binding domain for the c-myc 3' UTR. In addition, Hel-Ni was shown to be reactive with autoantibodies from patients with paraneoplastic encephalomyelitis both before and after binding to c-myc mRNA.RNA-binding proteins are involved in a variety of regulatory and developmental processes such as RNA processing and compartmentalization, mRNA translation, and viral gene expression. One family of RNA-binding proteins is defined by the presence of an 80-amino-acid RNA recognition motif (RRM) which was shown to constitute the core structure of the RNA binding domain of the UlsnRNP-70K protein (39). The motif characteristic of this family of proteins is evident from the conservation of several amino acid residues, most notably RNP1 and RNP2 consensus sequences (1,4,26,27). On the basis of crystallographic and nuclear magnetic resonance spectroscopic studies of the Ul RNA binding domain of the UlsnRNP-A protein, a model of the RRM tertiary structure has been derived (21, 36). The structural model of the domain together with biochemical studies has led to the suggestion that the RNA binding surface resides within a monomeric unit domain containing four antiparallel strands with solvent-exposed aromatic and basic residues (27).More than 100 members of the RRM superfamily have been reported, the majority of which are expressed in all tissues and are conserved throughout phylogeny (27). Tissue-specific members of the RRM superfamily are less common but include X16, which is expressed in pre-B cells (3); Bj6, which is a puff-specific Drosophila melanogaster protein (62); and elav (embryonic lethal abnormal vision), which is a neuron-specific protein in D. melanogaster (44 (43,44). However, to date no known RNA ligands for elav have been elucidated. Selection of RNAs from degenerate pools of in vitro transcripts demonstrated that Hel-Nl preferred to bind RNAs which resemble sequences in the 3' untranslated regions (UTRs) of certain mRNAs. Direct binding studies revealed that Hel-Nl can bind specific portions of the 3' UTR of rapidly degraded mRNAs, such as c-myc, c-fos, and granulocyte macrophage colony-stimulating factor (GM-CSF). Interestingly, des...

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