Alternative usage of 5' exons in the chicken nerve growth factor gene: refined characterization of a weakly expressed gene.
Nerve growth factor (NGF) is the prototype member of the neurotrophin family. Identification of transcript structures and promoter regions is described here in view of clarifying the molecular basis of chicken NGF gene regulation. Chicken NGF complementary DNA (cDNA) was amplified from heart and brain mRNA using the single-strand ligation to cDNA (SLIC) procedure. Several cloning and sequencing rounds were necessary to elucidate the diversity of NGF transcripts. The chicken NGF gene was shown to possess, in addition to its unique 3' coding exon, five 5' exons grouped into two clusters that have been entirely sequenced. The first cluster encompasses three leader exons (1a, 1b and 1c) and is separated from the second cluster by a approximately 15 kilobases (kb) intronic sequence. "Exon walking" based on reverse transcription-polymerase chain reaction (RT-PCR) allowed to ascertain the length of the three leader exons. The second cluster contains exons 2 and 3, separated from each other by a approximately 2.4 kb intron, and lies approximately 0.5 kb upstream from coding exon 4. Combination of several mechanisms, such as differential usage of leader and internal exons, alternative transcription start inside exon 1b, second donor and acceptor sites in exon 1c and 4, respectively, leads to the production of at least 21 different transcripts. This remarkable diversity may represent a common feature largely underestimated for other weakly expressed genes. Preliminary RT-PCR expression study in a panel of chicken tissues shows that transcripts containing exon 1b are present in most tissues tested. Transcripts containing exon 1a are represented mainly in heart and reproductive organs, whereas transcripts containing exon 1c are mostly represented in peripheral organs other than heart. Complementary data are published as a Web supplement available at.
Bertaux
Odile
O
Génomique Fonctionnelle et Biologie Systémique en Santé-CNRS FRE 2571, 19 rue Guy Môquet, BP 8, Villejuif cedex F-94 801, France. odile.bertaux@wanadoo.fr
Toselli-Mollereau
Esther
E
Auffray
Charles
C
Devignes
Marie-Dominique
MD
D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D002645:Chickens / Q000235:genetics*; D003001:Cloning, Molecular / Q000379:methods; D004247:DNA / Q000737:chemistry / Q000235:genetics; D005091:Exons / Q000235:genetics*; D005260:Female; D020869:Gene Expression Profiling; D008297:Male; D008969:Molecular Sequence Data; D020932:Nerve Growth Factor / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D014158:Transcription, Genetic / Q000235:genetics
D004247:DNA; D020932:Nerve Growth Factor