Medullary thyroid carcinomas in transgenic mice expressing a Polyoma carboxyl-terminal truncated middle-T and wild type small-T antigens A Felici* ,1,2,3 , M Giorgio 1,8 , N Krauzewicz 4 , C Della Rocca 5 , M Santoro 6 , P Rovere 5 , I Manni 7 , P Amati 2 and L Pozzi 1,2 1 Centro di Ricerca Sperimentale, Istituto Regina Elena, 00158 Rome, Italy; 2 Dip. di Biotecnologie Cellulari ed Ematologia, Universita Á degli Studi di Roma `La Sapienza', 00161 Rome, Italy; 3 Lab. di Patologia Vascolare, Istituto Dermopatico dell'Immacolata, 00167 Rome, Italy; 4 Department of Virology, Royal Postgraduate Medical School, London W12 0NN, UK; 5 Dip. di Medicina Sperimentale, Universita Á degli Studi di Roma `La Sapienza', 00161 Rome, Italy; 6 Centro di Endocrinologia ed Oncologia Sperimentale del CNR, c/o Dip. di Biologia e Patologia Cellulare e Molecolare, Facolta Á di Medicina e Chirurgia, Universita Á degli Studi di Napoli, 80131 Naples, Italy; 7 Lab. di Oncogenesi Molecolare, Istituto Regina Elena, 00158 Rome, Italy Medullary thyroid carcinoma (MTC) is a rare human tumor aecting the calcitonin-secreting c-cells of the thyroid. Here we report that two independent strains of transgenic mice expressing a Polyomavirus (Py) trun- cated middle-T antigen (DMT), consisting of the amino- terminal 304 amino acids, and the full length Py small-T antigen, developed multifocal bilateral MTCs with 100% penetrance. Occasionally one strain also developed mammary and bone tumors. Furthermore, ospring from both transgenic lines displayed pronounced waviness of the whiskers and fur, previously associated with defective epidermal growth factor receptor signaling. Transgene transcription, driven by the homologous early promoter/ enhancer, and the corresponding translation products were detected in tumors and in many other organs which did not develop pathologies. The subcellular distribution of DMT and its interactions with the adapter proteins of the SHC family have also been analysed. Our study describes a novel murine model of MTC and provides evidence that the N-terminal 304 amino acid fragment of Py middle-T antigen, possibly in co-operation with small- T antigen, acts as a potent oncogene in c-cells of the thyroid. Keywords: transgenic; oncogene; middle-T; medullary thyroid carcinoma Introduction The tumorigenic potential of Polyomavirus (Py) is localized to the early region of its genome which encodes three proteins: large-T (LT), middle-T (MT) and small-T (ST) antigen. Py early genes have been used together or separately in a transgenic context to dissect the oncogenic potential of this DNA tumor virus. In contrast to the wide spectrum of tumors obtained after Py infection of newborn or immuno- compromized mice (reviewed in Dawe, 1980), only a limited array of malignant pathologies have been observed in transgenic mice expressing Py T-antigens (reviewed in Kiefer et al., 1994). Several studies on transgenic models led us to unambiguously identify MT as the transforming gene of Py. In particular, these studies have shown that LT can be sucient for the development of a neoplastic phenotype in transgenic mice, although with a long latency (Chalifour et al., 1992; Paquis-Flucklinger et al., 1993; Helseth et al., 1992; Lebel et al., 1995), while ST is not oncogenic in vivo (Bautch, 1989). In contrast, MT alone, driven by its natural promoter, causes endotheliomas in transgenic mice within 4 ± 10 weeks after birth (Bautch et al.,1987). In addition to vascular tumors, bone tumors and lymphangiomas were observed when MT was studied in the context of the entire Py early region (Wang and Bautch, 1991). Moreover, under the control of heterologous tissue speci®c promoters, MT has been shown to induce tumors in a variety of tissues. Under these conditions, MT has been shown to transform mammary gland (Guy et al., 1992), urogenital and prostatic epithelium (Tehranian et al., 1996) and even a wider organ spectrum including salivary, mammary and thyroid gland (Rassoulzadegan et al., 1990). Here we report that transgenic mice expressing the 75% N-terminal fragment of MT antigen and the full length ST antigen consistently develop medullary thyroid carcinomas (MTC), a rare tumor type aecting the calcitonin (CT)-secreting c-cells of the thyroid. Unexpectedly, the MTC was associated with a second dominant phenotype consisting of waved hair and curly whiskers, very similar to that of the spontaneous mouse mutants wa-1 (Crew, 1933) and wa-2 (McLaren and Bowman, 1969). Results The NBL.Py lineages The transgene (NBL.Py) was constructed from 1.2 kbp of the Py virus early region under the transcriptional control of its own early promoter (Figure 1, upper panel). To overcome the in vivo low expression level of wild type Py early regulatory sequences (Krippl et al., 1988), we cloned the transgene under the control of the enhancer derived from a Py mutant virus (NB11/1) which displays a wide host cell range (Maione et al., 1985; De Simone and Amati, 1987). Indeed this 0.4 kb *Correspondence: A Felici 8 Current address: Department of Experimental Oncology, Istituto Europeo di Oncologia, 20141 Milan, Italy Received 12 August 1998; revised 22 October 1998; accepted 12 November 1998 Oncogene (1999) 18, 2387 ± 2395 ã 1999 Stockton Press All rights reserved 0950 ± 9232/99 $12.00 http://www.stockton-press.co.uk/onc