Regulation of rate of cartilage differentiation by Indian Hedgehog and PTH-related protein

Science

Volumn 273, Issue 5275, 1996, Pages 613-622

  Vortkamp, Andrea ^a   Lee, Kaechoong ^b   Lanske, Beate ^b   Segre, Gino V ^b   Kronenberg, Henry M ^b   Tabin, Clifford J ^a  

^a HARVARD MEDICAL SCHOOL   (United States)

^b MASSACHUSETTS GENERAL HOSPITAL   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

PARATHYROID HORMONE RELATED PROTEIN;

ANIMAL CELL; BONE DEVELOPMENT; CARTILAGE CELL; CELL DIFFERENTIATION; CHICKEN; CONTROLLED STUDY; NEGATIVE FEEDBACK; NONHUMAN; PRIORITY JOURNAL; PROTEIN EXPRESSION; PROTEIN ISOLATION; REVIEW; SEQUENCE HOMOLOGY;

ANIMALIA; ERINACEIDAE; GALLUS GALLUS;

EID: 0029750190     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.273.5275.613     Document Type: Review

References (74)

1. J. R. Hinchliffe and D. R. Johnson, in The Development of the Vertebrate Limb (Oxford Univ. Press, New York, 1980), pp. 76-83.
2. A. Erlebacher, E. H. Filvaroff, S. E. Gitelman, R. Derynck, Cell 80, 371 (1995).
3. F. Rousseau et al., Nature 371, 252 (1994).
4. R. Shiang et al., Cell 78, 335 (1994).
5. C. Deng, A. Wynshaw-Boris, F. Zhou, A. Kuo, P. Leder, ibid. 84, 911 (1996).
6. J. Baker, J. P. Liu, E. J. Robertson, A. Efstratiadis, ibid. 75, 73 (1993).
7. A. C. Karaplis et al., Genes Dev, 8, 277 (1994).
8. N. Amizuka, H. Warshawsky, J. E. Henderson, D. Goltzman, A. C. Karaplis, J. Cell Biol. 126, 1611 (1994).
9. M. J. Bitgood and A. P. McMahon, Dev. Biol. 172, 126 (1995).
10. C. Nüsslein-Volhard and E. Wieschaus, Nature 287, 795 (1980).
11. J. J. Lee, K. D. Von, S. Parks, P. A. Beachy, Cell 71, 33 (1992).
12. J. Mohler and K. Vani, Development 115, 957 (1992).
13. R. D. Riddle, R. L. Johnson, E. Laufer, C. Tabin, Cell 75, 1401 (1993).
14. Y. Echelard et al., ibid., p. 1417.
15. S. Krauss, J.-P. Concordet, P. W. Ingham, ibid., p. 1431.
16. M. Levin, R. L. Johnson, C. D. Stern, M. Kuehn, C. Tabin, ibid. 82, 803 (1995).
17. H. Roelink et al., ibid. 76, 761 (1994).
18. R. L. Johnson, E. Laufer, R. D. Riddle, C. Tabin, ibid. 79, 1165 (1994).
19. C. M. Fan and M. Tessier-Lavigne, ibid., p. 1175.
20. D. J. Roberts et al., Development 121, 3163 (1995).
21. M. Bitgood, L. Shen, A. McMahon, Curr. Biol. 6, 298 (1996).
22. J. E. Hooper and M. P. Scott, Cell 59, 751 (1989).
23. Y. Nakano et al., Nature 341, 508 (1989).
24. T. Orenic, D. C. Slusarski, K. L. Kroll, R. A. Holmgren, Genes Dev. 124, 50 (1990).
25. P. W. Ingham, A. M. Taylor, Y. Nakano, Nature 353, 184 (1991).
26. J. Capdevila, M. P. Estrada, E. Sánchez-Herrero, I. Guerrero, EMBO J. 13, 71 (1994).
27. A. J. Forbes, Y. Nakano, A. M. Taylor, P. W. Ingham, Dev. Suppl. 115 (1993).
28. V. Marigo, M. P. Scott, R. L. Johnson, L. V. Goodrich, C. J. Tabin, Development 122, 1225 (1996).
29. L. V. Goodrich, R. L. Johnson, L. Milenkovic, J. A. McMahon, M. P. Scott, Genes Dev. 10, 301 (1996).
30. J. M. Ruppert et al., Mol. Cell. Biol. 8, 3104 (1988).
31. V. Marigo and C. J. Tabin, unpublished results.
32. _, Proc. Natl. Acad. Sci. U.S.A., in press.
33. K. Basler and G. Struhl, Nature 368, 208 (1994).
34. J. Capdevila and I. Guerrero, EMBO J. 13, 4459 (1994).
35. R. Tabata and T. Kornberg, Cell 76, 89 (1994).
36. E. Laufer, C. E. Nelson, R. L. Johnson, B. A. Morgan, C. J. Tabin, ibid. 79, 993 (1994).
37. 6) of the unamplified library were plated and transferred to duplicate sets of nylon filters (Colony Plaque Screen, New England Nuclear). A genomic lhh DNA probe, plhhex3, homologous to base pairs (bp) 908 to 1180 of the Shh cDNA and containing 400 bp of 3′ untranslated sequence, and the Shh cDNA probe pHH2 (13), were mixed and used as a hybridization probe. Hybridization was carried out in 0.5 M phosphate buffer, 7% SDS, 1 mM EDTA, and herring sperm DNA (100 μg/ml) at 65°C overnight. Filters were washed in 50 mM phosphate buffer, with 1% SDS at 65°C as the final stringency (62), and exposed to Kodak XAR-5 films for 16 to 72 hours. After two rounds of subcloning, three positive clones were identified that hybridized strongly to plhhex3 and less strongly to pHH2. Nucleotide sequences of the three clones were determined with Sequenase 2.0 (US Biochemicals) and sequence-specific oligonucleotides as sequencing primers. DNA and amino acid sequences were analyzed with Genetics computer Group (Devreux) and DNA star software. A cDNA contig was assembled, consisting of 1958 bp with an open reading frame encoding a protein of 408 amino acids, flanked by 0.3 kb of untranslated cDNA on the 5′ side and 0.4 kb on the 3′ side. Comparison of the predicted amino acid sequence with the previously cloned vertebrate Hedgehog proteins showed highest conservation with the predicted mouse (73%) and human (75%) lhh proteins and less but substantial conservation with the other Hedgehog proteins (55 to 61%), indicating that the isolated cDNA represents the chicken lhh gene (GenBank accession number U58511).
38. J. J. Lee et al., Science 266, 1528 (1994).
39. J. A. Porter et al., Nature 374, 363 (1995).
40. D. A. Bumcrot, R. Takada, A. P. McMahon, Mol. Cell. Biol, 15, 2294 (1995).
41. C. M. Fan et al., Cell 81, 457 (1995).
42. E. Marti, D. A. Bumcrot, R. Takada, A. P. McMahon, Nature 375, 322 (1995).
43. H. Roelink et al., Cell 81, 445 (1995).
44. P. Currie and P. Ingham, Nature, In press.
45. V. Hamburger and H. L. Hamilton, J. Exp. Morphol. 88, 49 (1951).
46. Misexpression of lhh: The complete lhh coding region and part of the 3′ untranslated cDNA [nucleotides (nt) 328 to 1691] were cloned into replication-competent retroviral vectors (RCAS) with various host ranges with the Slacks13 shuttle vector (13, 63-65). lhh-expressing RCAS(E)-type virus, which has a limited host range, was used to infect susceptible chicken primary fibroblast, prepared from chick line 15b. The lhh-expressing cells were pelleted and transplanted into the anterior limb mesenchyme of a virally resistant outbred host strain, which resulted in a localized expression of lhh and prevented the spread of the virus into the surrounding tissue. All other misexpression experiments were carried out with lhh-expressing RCAS(A)-type viruses, which directly infected the outbred chicken host strain and spread throughout the surrounding tissue after infection (13), resulting in high lhh expression in the infected limb tissue. Outbred specific pathogen-free eggs were obtained from SPAFAS (Norwich, CT).
47. A. Vortkamp and C. J. Tabin, unpublished results.
48. 33P-UTP-labeled probes on 6-μm sections as described (66). Hybridization was done at 52° to 70°C in 50% formamid, and posthybridization washes were carried out at a final stringency of 55°C in 50% formamid and 2X SSC (sodium chloride/sodium citrate). For serial sections, three continuous sections were successively collected on six alternating slides. Probes used for hybridizations were as follows: lhh: plhh-R1-4sac contains nt 1 to 557 of the cloned lhh sequence and corresponds to the 5′ untranslated region and to exon 1. The probe detects the endogenous and the viral lhh transcript. Endogenous lhh: plhh-R1-4AE contains nt 1 to 276 of the lhh cDNA, which correspond to the 5′ untranslated lhh message and are not included in the viral construct. This probe detects only the endogenous lhh transcript. Human lhh (67), Gli (31), Shh (13), Hoxd-11, Hoxd-13, and Bmp2 (36), Ptc (28), Col-X (68), Col-IX (69), Bmp6 (B. Houston), PTHrP (70), PTH/PTHrP receptor (59), and rat PTH/PTHrP receptor (71).
49. T. F. Linsenmayer et al., Development 111, 191 (1991).
50. D. M. Kingsley, Trends Genet. 10, 16 (1994).
51. BMP expression: Because members of the Bmps/ dpp family are often induced by Hedgehog proteins, we also looked at the expression of Bmp-2, Bmp-4, Bmp-5, and Bmp-7. All were expressed in the perichondrium as described (50); however, none of their expression domains correlated with the domain of lhh expression.
52. Weigert-Safranin staining: 16-μm paraffin sections of chicken wings were fixed in Bouins fixative and stained with a series of Weigert hematoxylin, fast green, and safranin O.
53. L. J. Suva et al., Science 237, 893 (1987).
54. A. Broadus and A. Stewart, in The Parathyroids, J. Bilezikiaan, M. Levine, R. Marcus, Eds. (Raven, New York, 1994), pp. 259-294.
55. K. Lee, J. D. Deeds, G. V. Segre, Endocrinology 136, 453 (1995).
56. K. Lee et al., in preparation.
57. H. Juppner et al., Science 254, 1024 (1991).
58. B. Lanske et al., ibid. 273, 663 (1996).
59. PTH/PTHrP receptor probe: pRP-R1/4, corresponding to amino acids 219 to 454 of PTH/PTHrP receptor, was amplified from reverse-transcribed RNA of day 8 chicken limbs and subcloned into pBluescript SK-. Degenerate primers [GACGGATCC(CA)GIAA (CT)TA(CT)AT(CTA)CA(CT)ATGCA and GACGAATTC(TC)TC(AG)TA(AG)TGCAT(TC)TGIAC(TC)TGCCA] were based on the human, mouse, rat, pig, opossum, and Xenopus cDNA sequences. Polymerase chain reaction (PCR) conditions: 5 min at 94°C, 30 cycles of 30 s at 94°C, 30 s at 55°C, and 30 s at 72°C, fallowed by a final extension of 5 min at 72°C. Sequence analysis revealed that the resulting probe pRP-R1/4 shows highest homology to the opossum PTH/PTHrP receptor cDNA.
60. b medium containing 0.1% bovine serum albumin) alone from day 2 for 4 days. The samples were never exposed to serum. At the termination of culture, the hind limbs were fixed by 10% formalin in phosphate-buffered saline.
61. F. M. Ausubel et al., Current Protocols in Molecular Biology (Greene and Wiley Interscience, New York, 1989).
62. G. M. Church and W. Gilbert, Proc. Natl. Acad. Sci. U.S.A. 81, 1991 (1984).
63. D. M. Fekete and C. L. Cepko, Mol. Cell. Biol. 13, 2604 (1993).
64. _, Proc. Natl. Acad. Sci. U.S.A. 90, 2350 (1993).
65. S. H. Hughes, J. J. Greenhouse, C. J. Petropoulos, P. Sutrave, J. Virol. 61, 3004 (1987).
66. L. Tessarollo, L. Nagarajan, L. F. Parada, Development 115, 11 (1992).
67. V. Marigo et al., Genomics 28, 44 (1995).
68. P. LuValle, Y. Ninomiya, N. D. Rosenblum, B. R. Olsen, J. Biol. Chem. 263, 18378 (1988).
69. I. Nishimura, Y. Muragaki, B. R. Olsen, ibid. 264, 20033 (1989).
70. M. A. Thiede and S. J. Rutledge, Nucleic Acids Res. 18, 3062 (1990).
71. A. B. Abou-Samra et al., Adv. Nephrol. Necker Hosp. 23, 247 (1994).
72. A. Vortkampe et al., data not shown.
73. Our model suggests that PTHrP represses chondrocyte differentiation, acting on prehypertrophic cells just before the onset of lhh expression. However, the inhibitory PTHrP signal may also act, to some extent, on the proliferating chondrocytes before the prehypertrophic stage. In the mouse, both antibodies to the receptor and PTH-binding studies suggest that a very low level of PTH/PTHrP receptor may be present on the surface of proliferating cartilage cells even though the levels of PTH/PTHrP receptor mRNA are too low to be detected (72).
74. We thank R. L. Johnson for the genomic lhh probes and sequence information; V. Marigo for the probes for Gli and Ptc; B. R. Olsen for the Col-IX and Col-X probes; B. Houston for the Bmp-6 probes; and T. Woolf, L. Wang, and Ontogeny for providing Sonic hedgehog protein, prepared from an unpublished clone provided by H. Roelink. We are grateful to V. Marigo, J. Capdevilla, E. Laufer, and M. Belliveau for critical discussion of the manuscript and to the members of the Tabin lab for technical advice. A.V. was supported by a fellowship of the Human Frontiers Science Program (HFSP) (LT246/94). Supported by a grant from the HFSP (to C.J.T.) and by National Institutes of Health grants DK47038 (to H.M.K.) and DK4723 to (G.V.S.).