Liver-specific RNA processing of the ubiquitously transcribed rat fibrinogen γ-chain gene

Abstract

Fibrinogen γ-chains differ in amino acid sequence at the carboxyterminus due to alternative 3′ RNA processing. Previous studies reported differences between humans and rats in the mechanism of γ-chain RNA processing and that it was a nonregulated event. To test the hypothesis that rat γ-chain RNA processing involves both alternative splicing and poly(A) site selection and that it is regulated in a tissue-specific manner, we determined the tissue distribution of γ-chain mRNA expression and the pattern of γ-chain pre-mRNA processing. The results of in situ hybridization demonstrated that γA and γB transcripts were localized to and codistributed in liver hepatocytes, indicating that no subset of cells process γB mRNA. The ubiquitous expression of the fibrinogen γ-chain promoter was demonstrated in marrow, lung, brain, and liver by RNase protection using a 5′-specific γ-chain probe. RNase protection studies to map 3′ RNA processing sites suggested that, in addition to the distal poly(A) signal previously identified, two alternative poly(A) signals within the last intron (ATTAAA and AATAAA) were used only in liver to produce γB transcripts. Approximately equal usage of the three poly(A) signals (27%, 37%, and 36%, respectively) to form the 3′ end of mature γB transcripts suggested that poly(A) site selection is random. These results indicate that splicing of the last intron to produce γA mRNA is the ubiquitous and constitutive pattern of γ-chain RNA processing, while retention of the last intron to produce γB mRNAs is the tissue-specific and regulated pattern of γ-chain RNA processing. The pattern of rat γ-chain RNA processing is similar to human, implying that the mechanism is conserved. These data support a mechanism of tissue-specific splice site selection predominating over poly(A) site selection in γ-chain pre-mRNA processing. The expression of both fibrinogen γ-chain transcripts in liver, rather than mutually exclusive expression in liver and other tissues, provides a new model for studying tissue-specific alternative 3′ end formation regulatory mechanisms. © 1992 by The American Society of Hematology.