Tissue‐specific heterogeneity of the 3′‐untranslated region of L‐type pyruvate kinase mRNAs

Abstract

A single L-type pyruvate kinase (PK) gene seems to exist per haploid genome. It is expressed in the liver, kidney and small intestine in the form of three mRNA species of 2, 2.2 and 3.2 x l o 3 bases (kb). All three species are polyadenylated and translatable into the same L-type subunit. Primer extension experiments demonstrate that all three PK mRNAs have the same 5' ends. Nuclease S1 protection experiments with various cDNA and 3' genomic probes indicate that the different mRNA species only differ by the length of their 3' noncoding region. The mechanism responsible for the production of the three transcripts seems to be the use of alternative unusual polyadenylation sites. Run-on assays with specific probes recognizing only the 3.2-kb or all three mRNA species show that the transcription proceeds across the gene with similar rate. This means that the process involved in generation of the three transcripts is a posttranscriptional event, probably due to different sites of endonucleolytic cleavage of primary transcripts extending 3' from the gene region encoding the mature mRNAs. The ratio between the different PK mRNA species is, to a certain extent, tissue-specific and changes with development. The role of an 'identifier sequence' located in the 3' noncoding sequence of the 3.2-kb species in such a tissue-specific use of alternative polyadenylation sites is discussed.Many strategies are used by eucaryotic genes to modulate their expression. Accumulating evidence suggests that RNA processing is one of the biochemical events frequently involved in regulation of gene expression. In fact, several processes can be responsible for the production of different messenger RNAs from the same gene. Multiple RNAs could be generated from primary transcripts whose synthesis is controlled by alternative promotors, this phenomenon being then associated with differential splicing events [l -51.These RNAs could be derived from the same primary transcript through differential splicing events or selective choice of several polyadenylation sites eventually coupled with alternative splicing phenomena [6 -231. These alternative RNA processing events sometimes exhibit tissue and developmental specificity [5, 8, 11, 15, 18, 191. We have recently isolated liver L-type pyruvate kinase (PK) cDNA clones and used them to analyze RNAs from different tissues. Three mRNA species of 3.2, 2.2 and 2 kb were found in liver and in tissues which express L-type PK [24,25]. Genetic arguments, Southern blot analysis and detection of a single type of PK genomic clone in a rat DNA library demonstrate that they are encoded by the same gene.We show here that these different messenger RNAs share identical noncoding 5' and coding regions and differ only by the length of their 3' untranslated extensions, probably through the use of alternative polyadenylation sites. Moreover Abbreviations. kb, lo3 bases; SDS, sodium dodecyl sulphate; PK, pyruvate kinase.Enzyme. Pyruvate kinase (EC 2.7.1.40).the three different RNAs are expressed in a relatively tissuespec...

In vivo regulation of glycolytic and gluconeogenic enzyme gene expression in newborn rat liver.

“…Fig. 1 shows that three pyruvate kinase mRNA species of 3.2, 2.2, and 2.0 kb, respectively, hybridize to the probe, as reported by Marie et al (25). A modification ofthe 3.2/2.2 + 2 kb mRNA ratio occurred during development, but the three RNA species underwent largely parallel changes during the entire period studied.…”

“…This corrects for the size of cDNA probe relative to gene length and for the efficiency of hybridization, which averaged 40%. The L-type pyruvate kinase gene is 10 kb long, and we have used two overlapping cDNA clones covering 2.1 kb of the mRNA sequence (25). The PEP carboxykinase and albumin genes are 6 and 15 kb long, respectively.…”

Regulated expression of nuclear protein(s) in myogenic cells that binds to a conserved 3' untranslated region in pro alpha 1 (I) collagen cDNA.

“…The physiological importance of this decision may be reflected by the fact that the differential use of the collagen polyadenylation sites is maintained during evolution. Furthermore, the differential use of polyadenylation sites has been also reported for other gene products, such as the L-PK transcripts (20). In this case, it seems that the ratio of the different L-PK mRNAs which are specified by the length of the 3' UTR is even regulated in a developmentally and tissue-specific manner (20).…”

“…Furthermore, the differential use of polyadenylation sites has been also reported for other gene products, such as the L-PK transcripts (20). In this case, it seems that the ratio of the different L-PK mRNAs which are specified by the length of the 3' UTR is even regulated in a developmentally and tissue-specific manner (20). Nevertheless, the functional significance of these multiple L-PK transcripts remains unclear, since they have similar half-lives and appear to be translatable with comparable efficiencies (20).…”

“…Transcription can also be regulated via sequences in the 3' UTR, as has been shown for the ,-actin gene (8). Furthermore, multiple mRNAs with identical protein-coding regions but varying 3' UTRs are, for example, transcribed from the L-type pyruvate kinase (L-PK) gene (20). The different L-PK transcripts arise by use of different polyadenylation sites.…”