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The ETS family of transcription factors is involved in several normal remodeling events and pathological processes including tumor progression. ETS transcription factors are divided into subfamilies based on the sequence and location of the ETS domain. ETV5 (Ets variant gene 5; also known as ERM) is a member of the PEA3 subfamily. Our meta-analysis of normal, benign, and malignant thyroid samples demonstrated that ETV5 expression is upregulated in papillary thyroid cancer and was predominantly associated with BRAF V600E or RAS mutations. However, the precise role of ETV5 in these lesions is unknown. In this study, we used the KTC1 cell line as a model for human advanced papillary thyroid cancer (PTC) because the cells harbor the heterozygous BRAF (V600E) mutation together with the C250T TERT promoter mutation. The role of ETV5 in PTC proliferation was tested using RNAi followed by high-throughput screening. Signaling pathways driving ETV5 expression were identified using specific pharmacological inhibitors. To determine if ETV5 influences the expression of epithelial-to-mesenchymal (EMT) markers in these cells, an EMT PCR array was used, and data were confirmed by qPCR and ChIP-qPCR. We found that ETV5 is critical for PTC cell growth, is expressed downstream of the MAPK pathway, and directly upregulates the transcription factor TWIST1, a known marker of intravasation and metastasis. Increased ETV5 expression could therefore be considered as a marker for advanced PTCs and a possible future therapeutic target.

NHERF1/EBP50, an adaptor molecule that interacts with β-catenin, YAP, and PTEN, has been recently implicated in the progression of various human malignancies, including colorectal cancer. We report here that NHERF1 acts as a tumor suppressor in vivo for intestinal adenoma development. NHERF1 is highly expressed at the apical membrane of mucosa intestinal epithelial cells (IECs) and serosa mesothelial cells. NHERF1 -deficient mice show overall longer small intestine and colon that most likely could be attributed to a combination of defects, including altered apical brush border of absorbtive IECs and increased number of secretory IECs. NHERF1 deficiency in Apc Min/+ mice resulted in significantly shorter animal survival due to markedly increased tumor burden. This resulted from a moderate increase of the overall tumor density, more pronounced in females than males, and a massive increase in the number of large adenomas in both genders. The analysis of possible pathways controlling tumor size showed upregulation of Wnt-β-catenin pathway, higher expression of unphosphorylated YAP, and prominent nuclear expression of cyclin D1 in NHERF1-deficient tumors. Similar YAP changes, with relative decrease of phosphorylated YAP and increase of nuclear YAP expression, were observed as early as the adenoma stages in the progression of human colorectal cancer. This study discusses a complex role of NHERF1 for intestinal morphology and presents indisputable evidence for its in vivo tumor suppressor function upstream of Wnt-β-catenin and Hippo-YAP pathways.

Publisher Summary This chapter focuses on the role played by RET Proto-oncogene in Multiple Endocrine Neoplasia Type 2 (MEN2). The RET proto-oncogene is a tyrosine kinase receptor that regulates neuronal differentiation and kidney development and which is expressed in the neural crest during early embryonic life. Migration of RET-positive cells from the neural crest leads to development of the gastrointestinal neuronal system and the dorsal sympathetic system. Two broad categories of mutations cause hereditary MTC or MEN 2. The most common are extracellular mutations of the cysteine-rich dimerization domain and the less common are mutations that change single amino acids in the intracellular tyrosine kinase domain. The extracellular domain mutations associated with MEN 2 or familial MTC affect codons 609, 611, 618, 620, 630, and 634, and change a highly conserved cysteine to another amino acid. Mutations at codon 634 mutations are most common, accounting for approximately 80% of all mutations in MEN 2, where a single change (TGC to CGC; cysteine to arginine) accounts for approximately 50% of all RET mutations found in hereditary MTC. Missense mutations of RET cause transformation by one of two different mechanisms. Mutations of the extracellular domain (609, 611, 618, 620, 630, and 634) cause dimerization and activation of the receptor, presumably in the absence of GDN. Effective therapeutic options for treatment of the neoplastic components of MEN 2 have been limited to surgical removal of neoplastic components.

Publisher Summary Calcitonin is produced by the C (or calcitonin-producing) cells of the thyroid gland. The active principle was subsequently shown to be a 32 amino acid peptide, released in response to an increase in the plasma calcium concentration. The calcitonin gene family includes at least four separate genes. The first (CALC I or CGRP a) encodes calcitonin and a second peptide, calcitonin gene-related peptide (CGRP). Primary RNA transcript of second gene is processed to produce only CGRPb. The third is a pseudogene and is not expressed. The fourth encodes the peptide, amylin. Based on primary structure, the CTs can be divided into three groups: artiodactyl, including porcine, bovine, and ovine, which differ by four amino acids; primate/rodent, including human and rat calcitonins, which differ by two amino acids; and teleost/avian, including salmon, eel, goldfish, and chicken calcitonins, which differ by three amino acids. Calcitonin secretion is regulated by the extracellular calcium concentration. Increasing the extracellular calcium concentration results in increased secretion of CT. This effect is mediated by the interaction of calcium with the calcium-sensing receptor (CaR), a G-protein-coupled receptor expressed in high levels on the thyroidal C cell. In contrast to the parathyroid cell, where increasing calcium concentration activates the CaR and links to pathways that inhibit parathyroid hormone secretion, activation of this same receptor in the C cell causes increased secretion of CT. Calcitonin and its related peptides (CGRPa, CGRPb amylin, and adrenomedullin) are small peptides that exert their biological effect by binding to and activating a family of G-protein-coupled receptors. This receptor is expressed on osteoclasts, renal tubular cells, neurons in the central and peripheral nervous system, and other cell types. Several different isoforms of the CT receptor are produced by alternative RNA processing, which couple differently to downstream transduction pathways.

Publisher Summary This chapter discusses the molecular mechanisms of calcitonin (CT) gene transcription and posttranscriptional RNA processing. The primary site of CT production in mammals is the parafollicular cell or C cell of the thyroid gland. This cell type originates in the neural crest and migrates, during embryonic life, to the fourth pharyngeal pouch. A mass of the C-cell precursors, collectively known as the ultimobranchial body, fuses with the thyroid gland as it migrates caudally during neck morphogenesis. Transcription of protein-encoding genes by eukaryotic RNA polymerase II is regulated by interaction of trans -acting factors with specific DNA sequences, cis elements. The cell-restricted transcription of a gene is mediated by one or a combination of cis elements that function to enhance or repress transcription. The removal of intronic sequence between adjacent exons occurs in two steps. Exon sequence is first defined by recognition processing signals and then splicing proceeds by removal of sequence between defined exons These elements are located in regions flanking the transcription unit or within the transcription unit itself. The key event in the processing of the CT pre-mRNA is the inclusion or exclusion of exon four.