The hα-globin transcript contains a C-rich polypyrimidine tract located immediately 3′ to the cognate intron 1 splice donor that is a binding target for PCBP1 and PCBP2). (A) The hα-globin transcript and an expanded view of the intron 1 sequence. The hα-globin gene transcript contains 3 exons; the start codon in exon 1 and the stop codon in exon 3 are indicated. There are 3 defined C-rich tracts within the hα-globin transcript, each indicated by a red box: a C-rich element immediately 3′ to the cognate splice donor of intron 1, a second intronic C-rich element overlying the branch point polypyrimidine tract of the splice acceptor of intron 1, and a third C-rich tract is located within the 3′ UTR. The full intron 1 sequence is shown below the gene diagram, with the 2 C-rich tracts highlighted in red font. The C-rich region at the polypyrimidine tract of intron 1 and that within the 3′ UTR have both been previously linked to functions in transcript processing^10,25,27  and mRNA stability.^7,8,11,12  The function of the C-rich tract located immediately 3′ to the cognate intron 1 splice donor site has not been previously assessed for function. (B) The C-rich tract 3′ to the cognate intron 1 splice donor site of the hα-globin transcript is recognized by PCBP1 and PCBP2. A diagram of the region of the hα-globin transcript extending from exon 1 into exon 2 is shown, and the C-rich segment immediately 3′ to the cognate splice donor is boxed in red. Two Cs within this segment were converted to Gs to test for C-dependent activity of this region. The WT and mutant (MUT) probes were labeled with ³²P and incubated with HeLa cell extracts. After UV crosslinking, the extracts were analyzed on a sodium dodecyl sulfate polyacrylamide gel electrophoresis gel. A prominent complex (PCBPs) formed on the WT but not on the MUT probe; this complex comigrates with the complexes in the crosslinked extract that were immunoprecipitated with affinity-purified isotype-specific antiseras to PCBP1 and PCBP2. IgG, immunoglobulin G.