Erythroid cell-specific determinants of alpha-globin mRNA stability.
Although globin mRNAs are considered prototypes of highly stable messages, the mechanisms responsible for their longevity remain largely undefined. As an initial step in identifying potential cis-acting elements or structures which contribute to their stability, we analyzed the defect in expression of a naturally occurring alpha 2-globin mutant, alpha Constant Spring (CS). The CS mutation is a single-base change in the translation termination codon (UAA-->CAA) that allows the ribosome to read through into the 3' nontranslated region (NTR). The presence of CS mRNA in transcriptionally active erythroid precursors and its absence (relative to normal alpha-globin mRNA) in the more differentiated transcriptionally silent erythrocytes suggest that this mutation disrupts some feature of the alpha-globin mRNA required for its stability. Using a transient transfection system, we demonstrate that in murine erythroleukemia cells the CS mRNA is unstable compared with the normal alpha 2-globin mRNA. The analyses of several other naturally occurring and site-directed mutant alpha-globin genes in murine erythroleukemia cells indicate that entry of a translating ribosome into the 3' NTR targets the message for accelerated degradation in erythroid cells. In contrast, both the CS and alpha 2-globin mRNAs are stable in several nonerythroid cell lines. These results suggest that translational readthrough disrupts a determinant associated with the alpha 2-globin 3' NTR which is required for mRNA stability in erythroid cells.
Howard Hughes Medical Institute, University of Pennsylvania, Philadelphia 19104.
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Weiss
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Liebhaber
D000818:Animals; D001483:Base Sequence; D017931:DNA Primers / Q000737:chemistry; D015672:Erythroid Precursor Cells / Q000378:metabolism*; D005914:Globins / Q000235:genetics*; D006455:Hemoglobins, Abnormal / Q000235:genetics*; D066298:In Vitro Techniques; D051379:Mice; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D006455:Hemoglobins, Abnormal; D012333:RNA, Messenger; D005914:Globins; C035541:Hemoglobin Constant Spring