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. 1997 Apr 15;94(8):3994-9.
doi: 10.1073/pnas.94.8.3994.

Chondrocytes as a specific target of ectopic Fos expression in early development

H Watanabe  1 , K Saitoh, T Kameda, M Murakami, Y Niikura, S Okazaki, Y Morishita, S Mori, Y Yokouchi, A Kuroiwa, H Iba
Affiliations

Chondrocytes as a specific target of ectopic Fos expression in early development

H Watanabe et al. Proc Natl Acad Sci U S A. .

Abstract

The Finkel-Biskis-Jinkins murine sarcoma virus, which carries v-fos, induces osteosarcomas, whereas high-level expression of exogenous c-fos in transgenic and chimeric mice leads to postnatal development of osteogenic and chondrogenic tumors, respectively. To test whether such target cell specificity of an oncogene can be detected even in early development, we induced ectopic expression of fos in chicken limb buds by microinjecting replication-competent retrovirus into the presumptive leg field of stage 10 embryos. This caused cartilage truncation of all the long bones of the injected leg, which was mainly attributable to chondrodysplasia due to severe retardation of differentiation of the proliferating chondrocytes into mature or hypertrophic chondrocytes, as well as a slight delay in precartilagenous condensation. Expression of genes for all the other known members of chicken AP-1, which include such transforming genes as c-jun and fra-2, however, caused no macroscopic abnormalities in limb formation, indicating a specific function of Fos proteins in embryonic endochondral bone differentiation. The extent of truncation was stronger with v-Fos than with c-Fos, and comparative analysis of these proteins, as well as v-Fos mutants, revealed that strong transforming activity of Fos protein is necessary to cause dysplasia, suggesting that common molecular mechanisms are involved in both embryonic chondrodysplasia and bone tumor formation in postnatal mice.

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Figures

Figure 1
Figure 1
Protein structures of Fos proteins used in this study. ∗, A single amino acid exchange from c-Fos; ▤, the leucine zipper motif; ▨, a frame-shift mutation generated by a deletion present in v-Fos of FBJ-MSV. The mutant of v-Fos (FBJ), CD3, terminates at amino acid residue 202, whereas the wild type is composed of 381 amino acid residues.
Figure 2
Figure 2
Effect of v-fos virus expression in limb buds. A concentrated virus stock of recombinant replication-competent retrovirus (subgroup A), FJ-2, which encodes v-fos (originated from FBJ-MSV), was microinjected into the presumptive right leg or wing field at stage 10. Pairs of uninfected left limb (A–F Left) and infected right limb (A–F Right) from the same embryos are shown. The distribution of exogenous v-fos RNA (A) and proteoglycan H mRNA (B) is shown in the embryos at day 4.5, as detected by whole-mount in situ hybridization. (C–F) Dorsal view of wing cartilage (D Upper) or leg cartilage (C and D Lower, and E and F) in limbs at day 6 (C), day 8.5 (D), day 9.5 (E), and day 11.5 (F) after double staining with Alcian blue and Alizarin red. [Bars = 1 mm (A, for A–C) and 3 mm (F, for D–F).]
Figure 2
Figure 2
Effect of v-fos virus expression in limb buds. A concentrated virus stock of recombinant replication-competent retrovirus (subgroup A), FJ-2, which encodes v-fos (originated from FBJ-MSV), was microinjected into the presumptive right leg or wing field at stage 10. Pairs of uninfected left limb (A–F Left) and infected right limb (A–F Right) from the same embryos are shown. The distribution of exogenous v-fos RNA (A) and proteoglycan H mRNA (B) is shown in the embryos at day 4.5, as detected by whole-mount in situ hybridization. (C–F) Dorsal view of wing cartilage (D Upper) or leg cartilage (C and D Lower, and E and F) in limbs at day 6 (C), day 8.5 (D), day 9.5 (E), and day 11.5 (F) after double staining with Alcian blue and Alizarin red. [Bars = 1 mm (A, for A–C) and 3 mm (F, for D–F).]
Figure 3
Figure 3
(A) Suppression of calcification of several long bones is seen in the v-fos virus (FJ-2)-infected limb (right) as compared with the uninfected limb (left) of the same 8.5-day embryo. Cartilage was stained with Alcian blue (blue) and mineralized cartilage with Alizarin red (red). (B) Immunohistochemical analysis of type X collagen in sections near the central part of the tibia in the v-fos virus (FJ-2)-infected (Right) and uninfected (Left) limbs of the same 8.5-day embryo. Sections were counterstained with hematoxylin. The lower part is closer to the center. (Bar = 100 μm.)
Figure 3
Figure 3
(A) Suppression of calcification of several long bones is seen in the v-fos virus (FJ-2)-infected limb (right) as compared with the uninfected limb (left) of the same 8.5-day embryo. Cartilage was stained with Alcian blue (blue) and mineralized cartilage with Alizarin red (red). (B) Immunohistochemical analysis of type X collagen in sections near the central part of the tibia in the v-fos virus (FJ-2)-infected (Right) and uninfected (Left) limbs of the same 8.5-day embryo. Sections were counterstained with hematoxylin. The lower part is closer to the center. (Bar = 100 μm.)
Figure 4
Figure 4
Length ratio of each long bone in the infected right limb to that in the uninfected left limb of the same embryo (day 8.5–9.5). (A) Limbs infected with virus carrying c-fos (mouse), FM4 (•), or virus carrying v-fos derived from FBJ-MSV, FJ-2 (○), as well as mock-infected limbs (□). (B) Limbs infected with virus carrying c-fos (chicken), FC-1 (•), or NK24 (RAV-1), which carries v-fos (○).
Figure 5
Figure 5
(A) Suppression of the type X collagen gene expression in primary chondrocytes by the expression of exogenous fos genes. Chicken chondrocytes from the upper sternum (lanes 1–4) as well as CEF (lane 5) were infected just after preparation with a subgroup B virus carrying no exogenous gene [DS3(B), lane 1], or carrying chicken c-fos [FC1(B), lane 2], mouse c-fos [FM4(B), lane 3], or v-fos from FBJ-MSV [FJ-2(B), lane 4]. Total RNAs (8 μg) were prepared, separated on agarose gels, and hybridized with DNA probes encoding chicken type collagen (II) and type X collagen (X). (B) Western blot analysis of Fos proteins in CEF infected with subgroup A viruses such as DS3 (empty vector, lane 1), FM4 (c-fos virus, lane 2), FJ2 (v-fos virus, lane 4), CD3 (lane 5), and IR2-CD3 (lane 6), or a subgroup B virus, FM4(B) (c-fos virus, lane 3). Five days after infection, cells were disrupted under denaturing conditions, and each protein sample (20 μg) was separated in 10% polyacrylamide gel and analyzed. Molecular masses of c-Fos, v-Fos, and CD3 were 60, 55, and 35 kDa, respectively.
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References

    1. Curran T, Miller A D, Zokas L, Verma I M. Cell. 1984;36:259–268. - PubMed
    1. Curran T, Peters G, Van Beveren C, Teich N M, Verma I M. J Virol. 1982;44:674–682. - PMC - PubMed
    1. Finkel M P, Biskis B O, Jinkins P B. Science. 1966;151:698–701. - PubMed
    1. Cohen D R, Curran T. Mol Cell Biol. 1988;8:2063–2069. - PMC - PubMed
    1. Nishina H, Sato H, Suzuki T, Sato M, Iba H. Proc Natl Acad Sci USA. 1990;87:3619–3623. - PMC - PubMed

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