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Alternate splicing of human thromboxane synthase mRNA
1Division of Hematology, University of Texas Health Science Center at Houston 77030.
Archives of Biochemistry and Biophysics
|December 1, 1994
Summary
Alternate splicing of thromboxane synthase (TXS) RNA generates two mRNA variants. This process limits the cellular biosynthesis of thromboxane A2, a key inflammatory mediator.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- Two human thromboxane synthase (TXS) cDNA variants, TXS-I and TXS-II, were previously identified.
- TXS-II is characterized by a 163-base pair deletion near the 3'-end of its coding region compared to TXS-I.
- Both TXS mRNA variants are expressed in various cultured blood and lung cells.
Purpose of the Study:
- To investigate the molecular mechanism generating TXS-I and TXS-II mRNA variants.
- To determine the functional consequences of TXS-II formation on thromboxane A2 biosynthesis.
Main Methods:
- Analysis of exon-intron boundaries in TXS genomic DNA.
- Expression of recombinant TXS-I and TXS-II in a baculovirus system.
- Enzymatic assays to measure thromboxane A2 and 12-hydroxyheptadecatrienoic acid (HHT) production.
Main Results:
- The two TXS mRNA species arise from alternate splicing, specifically by skipping a 163-bp exon.
- This skipped exon encodes a heme-binding cysteine residue conserved in P450 enzymes.
- Recombinant TXS-I produced thromboxane A2 and HHT, while recombinant TXS-II lacked this enzymatic activity.
Conclusions:
- Alternate splicing of TXS RNA transcripts is a mechanism for regulating thromboxane A2 production.
- The skipping of a key exon in TXS-II leads to a non-functional enzyme.
- This regulatory pathway may control cellular levels of thromboxane A2.