EXPRESSION OF THE PACAP GENE IN A HUMAN NEUROBLASTOMA CELL-LINE - CDNA CLONING AND ANALYSES OF THE UPSTREAM REGULATORY REGION

作者：OHKUBO S; OGI K; KIMURA C; OKAZAKI K; ONDA H; FUJINO M

来源：Endocrine, 1994, 2(2): 135-145.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a novel peptide that occurs in both the brain and testis. A human neuroblastoma cell line, IMR-32, was shown by Northern blot analysis and enzyme immunoassay to express the PACAP gene and secrete immunoreactive PACAP, respectively. The size of PACAP mRNA was heterogeneous in these cells. PACAP cDNAs were cloned from IMR-32 cells. Comparison of the nucleotide sequences of the PACAP cDNAs and the gene confirmed the presence of intron-1 in the 5'-noncoding region. A cDNA lacking exon-4 was found and the polymerase chain reaction showed that a mRNA without exon-4 was routinely synthesized in these cells.
The transcriptional activities of the upstream regions of the PACAP gene were measured in IMR-32 cells with chloramphenicol acetyltransferase and luciferase as reporter genes. The assays showed that the PACAP promoter region was limited to about 400 bp. At least two promoter regions, designated P1 and P2, occurred in this sequence; neither region had TATA- or CCAAT-boxes. The downstream promoter, P1, had an Initiator (Inr)-like sequence, while the P2 promoter region had CC-boxes and a CT-rich sequence. Primer extension analysis specified a transcription initiation site for the P1 promoter in the Inr-like sequence, supporting the hypothesis that tissue-specific expression of the PACAP Inr-like element instead of TATA-boxes.

• 出版日期1994-2