Previous studies suggest that high-level stability of a subset of mammalian mRNAs is linked to a C-rich motif in the 3' untranslated region (3'UTR). High-level expression of human alpha-globin mRNA (h alpha-globin mRNA) in erythroid cells has been specifically attributed to formation of an RNA-protein complex comprised of a 3'UTR C-rich motif and an associated 39-kDa poly(C) binding protein, alpha CP. Documentation of this RNA-protein alpha-complex has been limited to in vitro binding studies, and its impact has been monitored by alterations in steady-state mRNA. Here we demonstrate that alpha CP is stably bound to h alpha-globin mRNA in vivo, that alpha-complex assembly on the h alpha-globin mRNA is restricted to the 3'UTR C-rich motif, and that alpha-complex assembly extends the physical half-life of h alpha-globin mRNA selectively in erythroid cells. Significantly, these studies also reveal that an artificially tethered alpha CP has the same mRNA-stabilizing activity as the native alpha-complex. These data demonstrate a unique contribution of the alpha-complex to h alpha-globin mRNA stability and support a model in which the sole function of the C-rich motif is to selectively tether alpha CP to a subset of mRNAs. Once bound, alpha CP appears to be fully sufficient to trigger downstream events in the stabilization pathway.