Human COL2Al-directed SV40 T Antigen Expression in Transgenic and Chimeric Mice Results in Abnormal Skeletal Development

The ability of SV40 T antigen to cause abnormalities in cartilage development in transgenic mice and chimeras has been tested. The cis-regulatory elements of the COL2A1 gene were used to target expression of SV40 T antigen to differentiating chondrocytes in transgenic mice and chimeras derived from embryonal stem (ES) cells bearing the same transgene. The major phenotypic consequences of transgenic (pAL21) expression are malformed skeleton, disproportionate dwarfism, and perinatal/neonatal death. Expression of T antigen was tissue specific and in the main characteristic of the mouse od(ID collagen gene. Chondrocyte densities and levels of ot I(ID collagen mRNAs were reduced in the transgenic mice. Islands of cells which express cartilage characteristic genes such as type fib procollagen, long form or(IX) collagen, o~2(XI) collagen, and aggrecan were found in the articular and growth cartilages of pAL21 chimeric fetuses and neonates. But these cells, which were expressing T antigen, were not properly organized into columns of proliferating chondrocytes. Levels of od0l) collagen mRNA were reduced in these chondroeytes. In addition, these cells did not express type X collagen, a marker for hypertrophic chondroeytes. The skeletal abnormality in pAL21 mice may therefore be due to a retardation of chondrocyte maturation or an impaired ability of chondroeytes to complete terminal differentiation and an associated paucity of some cartilage matrix components. URING vertebrate development, the establishment of a cartilage template involves the orderly differentiation of chondrocytes. This is important as the template forms a temporary framework for the formation of endochondral bone (Horton, 1993). Cartilage also is important because it has to withstand high cyclical loads and absorb shocks. This is achieved by the synthesis of a complex extracellular matrix, consisting of hyaluronate-proteoglycan complexes (Rosenberg and Buckwalter, 1986) and at least five different collagens, type II, VI, IX, X, and XI (Mayne and Irwin, 1986), one of the major components of which is type II collagen. There is growing evidence that mutations in the human oil(l/) collagen gene (COL2A1) ~ can cause some inherited chondrodysplasias, disorders which result in skeletal abnormalities characterized by disproportionate The current address of Ms. Alison Levy is Medical Research Council Collaborative Centre, Mill Hill, London NW7 lAD, United Kingdom. Address all correspondence to Dr. Kathryn S. E. Cheah, Department of Biochemistry, Hoag Kong University, Sassoon Rd., Hong Kong. Tel.: (852) 819 9170. Fax: (852) 855 1254. 1. Abbreviatious used in this paper: COL2A1, oil(H) collagen gene; d.p.c., days post coitum; ES, embryonal stem cells; Tag, SV40 T antigen. dwarfism (Spranger and Maroteaux, 1990; Horton and Hecht, 1993). Mutations in COL2A1 have been implicated in Stickler syndrome (Francomano et al, 1987; Ahmad et al., 1991, 1993; Brown et al., 1992;; W'mterpacht et al., 1993), Kniest dysplasia (Winterpacht et al., 1993; Poole et al., 1988), lethal (Langer-Saldino) aehondrogenesis (Vissing et al., 1989), spondyloepiphyseal dysplasia (Lee et al., 1989; Tiller et al., 1990; Cole et al., 1993; Vikkttla et al., 1993a), Goldblatt syndrome (Bonaventure et al., 1992), hypochondrogenesis (Horton et al., 1992), and also in primary generalized osteoarthfitis (Palotie et al., 1989; Vikkula et al., 1993b; Knowlton et al., 1990; Ala-Kokko et al., 1990). Dominant negative mutations in the ~ 1(I1) collagen gene expressed in transgenic mice result in severe chondrodysplasia (Mets~ranta et al., 1992; Garofalo et al., 1991; Vandenberg et al., 1991). These studies confirm the association of mutations in COL2A1 with some forms of human ehondrodysplasia. However, over 150 different ehondrodysplastic conditions have been described in humans varying from being perinatal lethal, to deforming during life, to being so mild that they are difficult to detect (Spranger and Maroteaux, 1990; Horton and Hecht, 1993). The molecular bases underlying these disorders are largely unknown. SV40 T antigen (Tag) is the major multifuncfional regula© The Rockefeller University Press, 0021-9525/95/01/223/15 $2.00 The Journal of Cell Biology, Volume 128, Numbers 1 &2, January 1995 223-237 223 on A ril 3, 2017 D ow nladed fom Published January 1, 1995