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A novel human insulinoma-associated cDNA, IA-1, encodes a protein with "zinc-finger" DNA-binding motifs
Y Goto1, M G De Silva, A Toscani
1Laboratory of Oral Medicine, National Institute of Dental Research, National Institutes of Health, Bethesda, Maryland 20892.
Abstract:
A subtraction library was constructed from human insulinoma (beta cell tumor) and glucagonoma (alpha cell tumor) cDNA phagemid libraries. Differential screening of 153 clones with end-labeled mRNAs from insulinoma, glucagonoma, and HeLa cells resulted in the isolation of a novel cDNA clone designated IA-1. This cDNA clone has a 2838-base pair sequence consisting of an open reading frame of 1530 nucleotides, which translates into a protein of 510 amino acids with a pI value of 9.1 and a molecular mass of 52,923 daltons. At the 3'-untranslated region there are seven ATTTA sequences between two polyadenylation signals (AATAAA). The IA-1 protein can be divided into two domains based upon the features of its amino acid sequence. The NH2-terminal domain of the deduced protein sequence (amino acids 1-250) has four classical pro-hormone dibasic conversion sites and an amidation signal sequence, Pro-Gly-Lys-Arg. The COOH-terminal domain (amino acids 251-510) contains five putative "zinc-finger" DNA-binding motifs of the form X3-Cys-X2-4-Cys-X12-His-X3-4-His-X4 which has been described as a consensus sequence for members of the Cys2-His2 DNA-binding protein class. Northern blot analysis revealed IA-1 mRNA in five of five human insulinoma and three of three murine insulinoma cell lines. Expression of this gene was undetectable in normal tissues. Additional tissue studies revealed that the message is expressed in several tumor cell lines of neuroendocrine origin including pheochromocytoma, medullary thyroid carcinoma, insulinoma, pituitary tumor, and small cell lung carcinoma. The restricted tissue distribution and unique sequence motifs suggest that this novel cDNA clone may encode a protein associated with the transformation of neuroendocrine cells.
Insights
Researchers identified a novel gene, IA-1, expressed in neuroendocrine tumors but not normal tissues. This gene may play a role in the transformation of neuroendocrine cells, offering potential diagnostic or therapeutic targets.
Area of Science:
- Molecular Biology
- Genetics
- Oncology
Background:
- Human insulinoma and glucagonoma cell lines were used to construct subtraction cDNA libraries.
- Differential screening identified a novel cDNA clone, IA-1, from these libraries.
Purpose of the Study:
- To isolate and characterize a novel gene expressed in human insulinoma and glucagonoma.
- To investigate the potential role of this gene in neuroendocrine tumor development.
Main Methods:
- Subtraction hybridization and differential screening of cDNA libraries.
- DNA sequencing and protein analysis (amino acid composition, pI, molecular mass).
- Northern blot analysis for gene expression in various cell lines and tissues.
Main Results:
- A novel cDNA clone, IA-1, of 2838 base pairs was isolated, encoding a 510-amino acid protein.
- The IA-1 protein contains potential pro-hormone processing sites and five zinc-finger DNA-binding motifs.
- IA-1 mRNA was detected in human and murine insulinomas and various neuroendocrine tumor cell lines (pheochromocytoma, medullary thyroid carcinoma, small cell lung carcinoma), but not in normal tissues.
Conclusions:
- The IA-1 gene product exhibits characteristics suggesting a role in DNA binding and protein processing.
- The restricted expression pattern in neuroendocrine tumors indicates IA-1 may be a marker or contributor to neuroendocrine cell transformation.