Three N-terminal Variants of the AE2 Cl/HCO Exchanger Are Encoded by mRNAs Transcribed from Alternative Promoters

doi:10.1074/jbc.271.13.7835

Three N-terminal Variants of the AE2 Cl/HCO Exchanger Are Encoded by mRNAs Transcribed from Alternative Promoters (*)

From the Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati College of Medicine, Cincinnati, Ohio 45267-0524

^¶To whom correspondence should be addressed:
Dept. of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati College of Medicine, Bethesda Ave., ML 524, Cincinnati, OH 45267-0524.
Tel.: 513-558-0056; Fax: 513-558-8474.

↵^§ Present address: Dept. of Cardiothoracic Surgery, Stanford University, Stanford, CA 94305.

Abstract

Multiple AE2 Cl/HCO exchanger mRNAs have been identified in rat. To determine the genetic basis for these mRNAs and whether they encode different variants of the exchanger, we used both rapid amplification of cDNA ends and S1 nuclease protection protocols and examined the organization of the gene. mRNAs encoding three N-terminal variants of AE2 (AE2a, AE2b, and AE2c) were identified and shown to be transcribed from alternative promoters. The AE2a transcription unit consists of 23 exons, with exons 1 and 2 containing 5′-untranslated sequence and the first 17 codons. The first exon of AE2b is located in intron 2; it contains 5′-untranslated sequence and an alternative 3-amino acid N-terminal coding sequence and is spliced to exon 3. The first exon of AE2c is located in intron 5; it consists of 5′-untranslated sequence and is spliced to exon 6, which contains the translation initiation codon corresponding to Met-200 of AE2a. Northern analysis shows that AE2a is expressed in all tissues, AE2b exhibits a more restricted distribution with highest levels in stomach, and AE2c is expressed only in stomach. Thus, the use of alternative promoters leads to the production of three N-terminal variants of AE2 that exhibit tissue-specific patterns of expression.

Footnotes

↵* This work was supported by National Institutes of Health Grants DK39626 and HL41496. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The nucleotide sequence(s) reported in this paper has been submitted to the GenBank(™)/EMBL Data Bank with accession number(s) U45885-U45887.
↵¹ The abbreviations used are:

AE

anion exchanger (the specific gene is designated by a number and variants produced by the use of alternative promoters are designated by a lowercase letter)

kb

kilobase(s)

nt

nucleotide(s)

bp

base pair(s)

RACE

rapid amplification of cDNA ends

PCR

polymerase chain reaction.
↵²These PCR artifacts consisted of several products with sequences derived entirely from the anchor oligonucleotide and the P2 primer. In some of these products, the P2 primer was partially duplicated.
↵³The transcription initiation site for the long form of AE3 has not yet been identified. Analysis of sequences extending several hundred bp upstream of the known 5′-most exon of AE3 do not reveal consensus acceptor splice sites (S. Linn and G. E. Shull, unpublished data), and the relative sizes of the brain and cardiac AE3 mRNAs suggest that there is little, if any, additional sequence in the larger mRNA besides that present in the cDNA. Therefore, in the discussion and Fig. 9 we have assumed that the 5′-most exon that was identified in the AE3 gene (11) is exon 1. However, the reader should be aware that the possibility of additional upstream exons has not been rigorously excluded.
- Received October 30, 1995.