Characterization of Gene Expression Induced by RET with MEN2A or MEN2B Mutation

Abstract

Germ-line point mutations of the RET gene are responsible for multiple endocrine neoplasia (MEN) type 2A and 2B that develop medullary thyroid carcinoma and pheochromocytoma. We performed a differential display analysis of gene expression using NIH 3T3 cells expressing the RET-MEN2A or RET-MEN2B mutant proteins. As a consequence, we identified 10 genes induced by both mutant proteins and eight genes repressed by them. The inducible genes include cyclin D1, cathepsins B and L, and cofilin genes that are known to be involved in cell growth, tumor progression, and invasion. In contrast, the repressed genes include type I collagen, lysyl oxidase, annexin I, and tissue inhibitor of matrix metalloproteinase 3 (TIMP3) genes that have been implicated in tumor suppression. In addition, six RET-MEN2A-and five RET-MEN2B-inducible genes were identified. Among 21 genes induced by RET-MEN2A and/or RET-MEN2B, six genes including cyclin D1, cathepsin B, cofilin, ring finger protein 11 (RNF11), integrin-␣6, and stanniocalcin 1 (STC1) genes were also induced in TGW human neuroblastoma cells in response to glial cell line-derived neurotrophic factor stimulation. Because the STC1 gene was found to be highly induced by both RET-MEN2B and glial cell line-derived neurotrophic factor stimulation, and the expression of its product was detected in medullary thyroid carcinoma with the MEN2B mutation by immunohistochemistry, this may suggest a possible role for STC1 in the development of MEN 2B phenotype. The RET proto-oncogene encodes a receptor tyrosine kinase with a cadherin-related motif and a cysteine-rich domain in the extracellular domain and is located on chromosome 10q11.2.1,2 It has been demonstrated that RET is a functional receptor for four related neurotrophic factors including glial cell line-derived neurotrophic factor (GDNF), neurturin, artemin, and persephin. These factors are known to require glycosylphosphatidylinositol-anchored co-receptors, GFR-␣s, as ligand-binding components and to promote the survival of various central and peripheral neurons in culture.1,2 In addition, gene knockout studies revealed that the GDNF/RET signaling plays a crucial role in the development of the enteric nervous system and the kidney. [3][4][5][6] Germline mutations of the RET gene cause dominant inherited cancer syndromes; multiple endocrine neoplasia (MEN) type 2A and 2B.7-10 MEN 2A is characterized by the development of medullary thyroid carcinoma (MTC), pheochromocytoma, and parathyroid hyperplasia. MEN 2B shows a more complex phenotype with association of MTC, pheochromocytoma, and developmental abnormalities such as mucosal neuroma, hyperganglionosis of the intestinal tract, and marfanoid skeletal changes. The MEN2A mutations were identified in cysteine residues of the RET extracellular domain, leading to ligand-independent RET dimerization. 11,12 The MEN2B mutations were detected in methionine at codon 918 or in alanine at codon 883 in the tyrosine kinase domain and appear to activate RET without dimerization. 12,13 A variety of si...

Regulation of microRNA expression by HMGA1 proteins

“…The modulation of its expression has been revealed in numerous developmental, physiological and pathological processes. Recently, STC1 has also been reported to be induced in a variety of tumors including the breast (McCudden et al, 2004), colon (Gerritsen et al, 2002), ovary (Ismail et al, 2000), lung cancer (Garber et al, 2001), thyroid medullary carcinoma (Watanabe et al, 2002), hepatocarcinoma (Okabe et al, 2001), pheochromocytoma (Eisenhofer et al, 2004), neuroblastoma (Wong et al, 2002), osteosarcoma and fibrosarcoma (Jellinek et al, 2000). The possible use of STC1 expression levels for the diagnosis of human breast, hepatocellular and colorectal cancer has also been proposed (Fujiwara et al, 2000;Wascher et al, 2003).…”

“…The possible use of STC1 expression levels for the diagnosis of human breast, hepatocellular and colorectal cancer has also been proposed (Fujiwara et al, 2000;Wascher et al, 2003). Moreover, it has been shown that the inherited mutations of the RET gene, responsible for the MEN2A and MEN2B, are able to induce STC1 expression (Watanabe et al, 2002). Recently, a critical role of STC1 expression in the formation of tumor vasculature has been proposed.…”

“…The STC1 gene is expressed in a wide variety of tissues, including the kidney, prostate, thyroid, bone and ovary (Chang et al, 1995;Olsen et al, 1996;Varghese et al, 1998). Recently, STC1 overexpression has been found in hepatocellular, colorectal, breast and medullary thyroid carcinomas (Fujiwara et al, 2000;Okabe et al, 2001;Watanabe et al, 2002;McCudden et al, 2004). Three sites in the 3 0 -UTR of ATXN1 gene and three in the 3 0 -UTR of STC1 gene, that match the miR-101b 'seed sequence', were predicted ( Figure 4a).…”

Roles of induced expression of MAPK phosphatase-2 in tumor development in RET-MEN2A transgenic mice

“…To investigate whether MKP-2 expression is induced by the activation of endogenous RET by GDNF, TGW neuroblastoma cells, which have been shown to express both endogenous RET and GFRa1 (Watanabe et al, 2002;Uchida et al, 2006), were treated with GDNF. MKP-2 was weakly expressed in TGW cells in their basal state, and biphasic induction of MKP-2 was observed 15 min and 6 h after GDNF stimulation Figure S1).…”

Evaluation of potential mechanisms underlying genotype–phenotype correlations in multiple endocrine neoplasia type 2

“…Interestingly, it was shown that JNK mediates survival signals after cell transformation by the oncogenic tyrosine kinase BCR-ABL (Hess et al, 2002). In addition, Watanabe et al, (2002) established a number of genes differentially expressed by MEN 2A-RET or MEN 2B-RET. Again, these results imply striking differences in pathways activated by MEN 2B-RET.…”