Concurrent overexpression of RET/PTC1 and TTF1 confers tumorigenicity to thyrocytes

Toyoshi Endo,Tetsuro Kobayashi

doi:10.1530/erc-13-0310

Abstract

A variant located on 14q13.3 nearest to thyroid transcription factor-1 (TTF1) predisposes individuals to thyroid cancer, but whether this variant is related to the RET/PTC rearrangement associated with human papillary thyroid carcinomas (PTCs) is unknown. The aims of this study were to investigate the effects of RET/PTC1 on the expression of thyroid-specific genes in thyrocytes and their relationship with malignant transformation of the thyrocytes. In the absence or presence of TSH, an extracellular signal-regulated kinase was phosphorylated in FRTL5 cells that stably expressed RET/PTC1, and these cells grew independently of TSH. FRTL (RET/PTC1) cells produced 566% more thyroglobulin mRNA and 474% more Na+/I− symporter mRNA than did the control FRTL (pcDNA) cells. FRTL (RET/PTC1) cells expressed 468% more Ttf1 mRNA than did FRTL (pcDNA) cells, but these two cell types did not differ significantly with respect to Pax8 or Ttf2 mRNA levels. When FRTL (RET/PTC1) cells and FRTL (pcDNA), cells were injected into each of nine nude mice, each mouse developed a single tumor at the site of FRTL (RET/PTC1) cell injection; in contrast, tumor formation never occurred at sites of FRTL (cDNA) cells injection. Tumors resulting from FRTL (RET/PTC1) cells retained 125I-uptake activity; moreover, the cells invaded into surrounding skeletal muscle. When overexpression of Ttf1 in FRTL (RET/PTC1) cells was silenced, the cells completely lost their tumorigenic potential. Exogenous TTF1 cDNA enhanced the tumorigenicity of BHP18-21v cells, human PTC cells that express RET/PTC1, in nude mice. These results indicated that concurrent overexpression of RET/PTC1 and TTF1 confers tumorigenicity to FRTL5 and BHP18-21v cells in nude mice.

Highlights

Expression of Tg and NaC/IK symporter (Nis) mRNAs in FRTL (RET/PTC1) cells differed significantly from that in FRTL cells; we studied the effect of RET/PTC1 on expression of three genes encoding thyroid-specific transcription factors – Ttf1, Ttf2, and Pax8 – in FRTL (RET/PTC1) and FRTL cells
Expression of RET/PTC1 in FRTL5 cells caused nuclei to become irregularly shaped and cell proliferation to become independent of TSH
Pax8 expression was significantly lower in PC Cl (RET/PTC1) cells than in the parental PC Cl cells, but transcription factor-1 (TTF1) expression was essentially unaltered by RET/PTC1 expression; function of TTF1 might be inactive (De Vita et al 1998)

Summary

Introduction

Papillary thyroid carcinomas (PTCs) are the most frequent cancers of the thyroid gland, and they are usually well differentiated given their ability to i) take up iodine, ii) secrete thyroglobulin (TG), and iii) be responsive to thyroidstimulating hormone (TSH; Nikiforov & Nikiforova 2011). The RET/PTC rearrangement and the BRAFV600E point mutation are the two most common genetic alterations associated with PTCs; the prevalence of RET/PTC varies from 2.5 to 78% (Zou et al 1994, Nikiforov et al 1997), and the prevalence of BRAFV600E varies from 23 to 62% (Xing et al 2005, Fugazzola et al 2006). Constitutive activation of ERK caused by a RET/PTC oncogene may or may not be sufficient to induce all hallmarks of cancer in vivo. Santoro et al (1996) found that some RET/PTC1 transgenic mice developed thyroid tumors, but others developed only thyroid hyperplasia. Knostman et al (2007) reported that doxycycline-induced expression of RET/PTC1 led to ERK phosphorylation in mice that carried a doxycyclineregulated RET/PTC1 transgene; thyroid lesions were not found in any of these mice

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