Expression of the delta 1-crystallin gene: salt-dependent alteration in the cell-free synthesis of delta-crystallin polypeptides.
We have investigated the origins of the two major size classes of delta-crystallin polypeptides (48 kDa and 50 kDa on sodium dodecyl sulfate-urea-polyacrylamide gels) in the chicken lens. Both the 48-kDa and 50-kDa polypeptides were produced by cell-free translation in a salt-dependent (Na+, K+, acetate and Cl-) pattern from mRNA derived from a cloned delta 1-crystallin cDNA. The salt-dependent alteration in the ratio of cell-free synthesis of the 48-kDa to 50-kDa delta-crystallin polypeptides was greatly enhanced by capping of the delta 1 mRNA. Translation of the delta 1 mRNA containing a premature termination codon suggested that the larger delta-crystallin band contains two polypeptides which differ in their N-terminal one-third. In vitro transcription/translation analysis of several mutant delta 1 cDNA clones and immunoblot analysis of authentic delta-crystallin with antisera raised to peptides contained in delta-crystallin showed that neither alternative initiation at two in-phase AUG codons nor alternative termination at sites on the delta 1 mRNA are responsible for generating the two sizes of the delta-crystallin polypeptides. Taken together our data suggest (but do not prove) that delta-crystallin heterogeneity is generated by co-translational modification of the primary polypeptide encoded in the delta 1-crystallin gene.
Laboratory of Molecular and Developmental Biology, National Eye Institute, Bethesda, MD 20892.
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EF
Wawrousek
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J
Piatigorsky
D000085:Acetates / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D002474:Cell-Free System; D002645:Chickens; D002712:Chlorides / Q000494:pharmacology; D003001:Cloning, Molecular; D003459:Crystallins / Q000096:biosynthesis / Q000235:genetics*; D005796:Genes; D007908:Lens, Crystalline / Q000378:metabolism*; D008969:Molecular Sequence Data; D010452:Peptide Biosynthesis; D011188:Potassium / Q000494:pharmacology; D014176:Protein Biosynthesis / Q000187:drug effects*; D011994:Recombinant Proteins; D012964:Sodium / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects*
D000085:Acetates; D002712:Chlorides; D003459:Crystallins; D011994:Recombinant Proteins; D012964:Sodium; D011188:Potassium