HNRNP-L BINDS A CIS-ACTING RNA SEQUENCE ELEMENT THAT ENABLES INTRON-INDEPENDENT GENE-EXPRESSION

Most pre-mRNAs require an intron for efficient processing in higher eu karyotes. To test the hypothesis that intron-independent gene expressi on involves positive, cis-acting RNA sequence elements, we constructed chimeric genes in which various regions of the naturally intronless H SV-TK gene were inserted into an intronless variant of the highly intr on-dependent human P-globin gene. Using a transient transfection assay , we identified a 119-nucleotide sequence element contained within the transcribed region of the HSV-TK gene that enables efficient cytoplas mic accumulation of globin RNA in the absence of splicing. RNA UV-cros s-linking assays indicated that a 68-kD protein present in nuclear ext racts of HeLa and COS cells specifically binds to this HSV-TK sequence element. This 68-kD protein was found to cross-react with an antiseru m specific to hnRNP L. Recombinant hnRNP L was shown to bind with high sequence specificity to this RNA sequence element. Analysis of substi tution mutants in this element indicated that binding of hnRNP L corre lates with accumulation of the RNA in the cytoplasm. Thus, we conclude that (1) hnRNP L binds in a sequence-specific manner to this RNA sequ ence element that enables intron-independent gene expression, and (2) intron-independent pre-mRNA processing and transport involves sequence -specific RNA-protein interactions between cis-acting RNA sequence ele ments and proteins such as hnRNP L. This sequence element may be of ge neral use for the efficient expression of cDNA versions of intron-depe ndent genes.