A novel 3.6-kb cDNA, IA-2, with a 2,937-bp open reading frame was isolated from a human insulinoma subtraction library (ISL-153). The predicted amino acid sequence and in vitro-translated product of IA-2 cDNA revealed a 979-amino-acid protein with a pI value of 7.09 and a molecular mass of 105,847 daltons. The protein sequence is consistent with a signal peptide, an extracellular domain, a transmembrane region, and an intracellular domain. The extracellular domain contains an unusual cysteine-rich region following the signal peptide. The intracellular cytoplasmic domain of IA-2 possesses highly conserved regions similar to the catalytic domains found in members of the protein tyrosine phosphatase (PTP) family. Northern blot analysis showed that IA-2 mRNA was expressed in five of five freshly isolated human insulinomas, rat and mouse insulinoma cell lines, and enriched normal mouse islets. It also was found in normal human brain, pituitary, pancreas, and brain tumor cell lines, but not in a variety of other normal or tumor tissues. Based on the sequence and expression data, it appears that IA-2 is a new member of the receptor-type PTP family that is expressed in islet and brain tissues.

We have previously reported the establishment of cytotoxic T-cell lines, from pancreatic cancer patients, by continuously stimulating tumor-draining lymph node cells with allogeneic pancreatic tumor cell lines. After the preliminary characterization of their phenotype and tumor specificity, detailed studies performed with one of the cell lines, W.D., show that it recognizes a specific antigen, a large and heavily glycosylated mucin molecule, expressed on pancreatic and breast tumors and tumor cell lines. Although this recognition appears major histocompatibility complex (MHC)-unrestricted, the antigen receptor used by the cytotoxic T cell is the a/fl heterodimer, typically found on MHC-restricted T cells.The target antigen is atypical, however, in its ability to directly bind and activate the T cells in the absence of self MHC, presumably by abundant and regularly repeated antigenic epitopes. These rmfings are important because they demonstrate a specific T-cell response against a human tumorassociated antigen. In addition to pancreatic and breast tumors, various mucin molecules are known to be produced by other tumors of epithelial cell origin and could be expected to stimulate similar T-cell-mediated immune responses.

Neurogenin 3 (Ngn3) is key for endocrine cell specification in the embryonic pancreas and induction of a neuroendocrine cell differentiation program by misexpression in adult pancreatic duct cells. We identify the gene encoding IA1, a zinc-finger transcription factor, as a direct target of Ngn3 and show that it forms a novel branch in the Ngn3-dependent endocrinogenic transcription factor network. During embryonic development of the pancreas, IA1 and Ngn3 exhibit nearly identical spatio-temporal expression patterns. However, embryos lacking Ngn3 fail to express IA1 in the pancreas. Upon ectopic expression in adult pancreatic duct cells Ngn3 binds to chromatin in the IA1 promoter region and activates transcription. Consistent with this direct effect, IA1 expression is normal in embryos mutant for NeuroD1, Arx, Pax4 and Pax6, regulators operating downstream of Ngn3. IA1 is an effector of Ngn3 function as inhibition of IA1 expression in embryonic pancreas decreases the formation of insulin-and glucagon-positive cells by 40%, while its ectopic expression amplifies neuroendocrine cell differentiation by Ngn3 in adult duct cells. IA1 is therefore a novel Ngn3-regulated factor required for normal differentiation of pancreatic endocrine cells.