Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1992 Mar 15;89(6):2091–2095. doi: 10.1073/pnas.89.6.2091

Activation of the cytotactin promoter by the homeobox-containing gene Evx-1.

F S Jones 1, G Chalepakis 1, P Gruss 1, G M Edelman 1
PMCID: PMC48602  PMID: 1372434

Abstract

Cytotactin is a morphoregulatory molecule of the extracellular matrix affecting cell shape, division, and migration that appears in a characteristic and complex site-restricted pattern during embryogenesis. The promoter region of the gene that encodes chicken cytotactin contains a variety of potential regulatory sequences. These include putative binding sites for homeodomain proteins and a phorbol 12-O-tetradecanoate 13-acetate response element (TRE)/AP-1 element, a potential target for transcription factors thought to be involved in growth-factor signal transduction. To determine the effects of homeobox-containing genes on cytotactin promoter activity, we conducted a series of cotransfection experiments on NIH 3T3 cells using cytotactin promoter-chloramphenicol acetyltransferase (CAT) reporter gene constructs and plasmids driving the expression of mouse homeobox genes Evx-1 and Hox-1.3. cotransfection with Evx-1 stimulated cytotactin promoter activity whereas cotransfection in control experiments with Hox-1.3 had no effect. To localize the sequences required for Evx-1 activation, we tested a series of deletions in the cytotactin promoter. An 89-base-pair region containing a consensus TRE/AP-1 element was found to be required for activation. An oligonucleotide segment containing this TRE/AP-1 site was found to confer Evx-1 inducibility on a simian virus 40 minimal promoter; mutation of the TRE/AP-1 site abolished this activity. To explore the potential role of growth factors in cytotactin promoter activation, chicken embryo fibroblasts, which are known to synthesize cytotactin, were first transfected with cytotactin promoter constructs and cultured under minimal conditions in 1% fetal bovine serum. Although the cells exhibited only low levels of CAT activity under these conditions, cells exposed for 12 h to 10% (vol/vol) fetal bovine serum showed a marked increase in CAT activity. Cotransfection with Evx-1 and cytotactin promoter constructs of cells cultured in 1% fetal bovine serum was sufficient, however, to produce high levels of CAT activity. These findings are consistent with the hypothesis that Evx-1, a homeobox-containing gene, may activate the cytotactin promoter by a mechanism involving a growth-factor signal transduction pathway. More generally, the results support the hypothesis that the place-dependent expression of morphoregulatory molecules may depend upon local cues provided by homeobox genes and their encoded proteins.

Full text

PDF
2091

Images in this article

2092Image
on p.2092
2092Image
on p.2092
2093Image
on p.2093
2093Image
on p.2093
2094Image
on p.2094

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Angel P., Allegretto E. A., Okino S. T., Hattori K., Boyle W. J., Hunter T., Karin M. Oncogene jun encodes a sequence-specific trans-activator similar to AP-1. Nature. 1988 Mar 10;332(6160):166–171. doi: 10.1038/332166a0. [DOI] [PubMed] [Google Scholar]
  2. Angel P., Imagawa M., Chiu R., Stein B., Imbra R. J., Rahmsdorf H. J., Jonat C., Herrlich P., Karin M. Phorbol ester-inducible genes contain a common cis element recognized by a TPA-modulated trans-acting factor. Cell. 1987 Jun 19;49(6):729–739. doi: 10.1016/0092-8674(87)90611-8. [DOI] [PubMed] [Google Scholar]
  3. Awgulewitsch A., Bieberich C., Bogarad L., Shashikant C., Ruddle F. H. Structural analysis of the Hox-3.1 transcription unit and the Hox-3.2--Hox-3.1 intergenic region. Proc Natl Acad Sci U S A. 1990 Aug;87(16):6428–6432. doi: 10.1073/pnas.87.16.6428. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bastian H., Gruss P. A murine even-skipped homologue, Evx 1, is expressed during early embryogenesis and neurogenesis in a biphasic manner. EMBO J. 1990 Jun;9(6):1839–1852. doi: 10.1002/j.1460-2075.1990.tb08309.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bohmann D., Keller W., Dale T., Schöler H. R., Tebb G., Mattaj I. W. A transcription factor which binds to the enhancers of SV40, immunoglobulin heavy chain and U2 snRNA genes. Nature. 1987 Jan 15;325(6101):268–272. doi: 10.1038/325268a0. [DOI] [PubMed] [Google Scholar]
  6. Chiquet-Ehrismann R., Mackie E. J., Pearson C. A., Sakakura T. Tenascin: an extracellular matrix protein involved in tissue interactions during fetal development and oncogenesis. Cell. 1986 Oct 10;47(1):131–139. doi: 10.1016/0092-8674(86)90374-0. [DOI] [PubMed] [Google Scholar]
  7. Cho K. W., De Robertis E. M. Differential activation of Xenopus homeo box genes by mesoderm-inducing growth factors and retinoic acid. Genes Dev. 1990 Nov;4(11):1910–1916. doi: 10.1101/gad.4.11.1910. [DOI] [PubMed] [Google Scholar]
  8. Crossin K. L. Cytotactin binding: inhibition of stimulated proliferation and intracellular alkalinization in fibroblasts. Proc Natl Acad Sci U S A. 1991 Dec 15;88(24):11403–11407. doi: 10.1073/pnas.88.24.11403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Crossin K. L., Hoffman S., Grumet M., Thiery J. P., Edelman G. M. Site-restricted expression of cytotactin during development of the chicken embryo. J Cell Biol. 1986 May;102(5):1917–1930. doi: 10.1083/jcb.102.5.1917. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Curran T., Franza B. R., Jr Fos and Jun: the AP-1 connection. Cell. 1988 Nov 4;55(3):395–397. doi: 10.1016/0092-8674(88)90024-4. [DOI] [PubMed] [Google Scholar]
  11. Dollé P., Duboule D. Two gene members of the murine HOX-5 complex show regional and cell-type specific expression in developing limbs and gonads. EMBO J. 1989 May;8(5):1507–1515. doi: 10.1002/j.1460-2075.1989.tb03535.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Faissner A., Kruse J. J1/tenascin is a repulsive substrate for central nervous system neurons. Neuron. 1990 Nov;5(5):627–637. doi: 10.1016/0896-6273(90)90217-4. [DOI] [PubMed] [Google Scholar]
  13. Franza B. R., Jr, Rauscher F. J., 3rd, Josephs S. F., Curran T. The Fos complex and Fos-related antigens recognize sequence elements that contain AP-1 binding sites. Science. 1988 Mar 4;239(4844):1150–1153. doi: 10.1126/science.2964084. [DOI] [PubMed] [Google Scholar]
  14. Friedlander D. R., Hoffman S., Edelman G. M. Functional mapping of cytotactin: proteolytic fragments active in cell-substrate adhesion. J Cell Biol. 1988 Dec;107(6 Pt 1):2329–2340. doi: 10.1083/jcb.107.6.2329. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Graham A., Papalopulu N., Krumlauf R. The murine and Drosophila homeobox gene complexes have common features of organization and expression. Cell. 1989 May 5;57(3):367–378. doi: 10.1016/0092-8674(89)90912-4. [DOI] [PubMed] [Google Scholar]
  16. Graham A., Papalopulu N., Krumlauf R. The murine and Drosophila homeobox gene complexes have common features of organization and expression. Cell. 1989 May 5;57(3):367–378. doi: 10.1016/0092-8674(89)90912-4. [DOI] [PubMed] [Google Scholar]
  17. Grierson J. P., Petroski R. E., Ling D. S., Geller H. M. Astrocyte topography and tenascin cytotactin expression: correlation with the ability to support neuritic outgrowth. Brain Res Dev Brain Res. 1990 Aug 1;55(1):11–19. doi: 10.1016/0165-3806(90)90100-d. [DOI] [PubMed] [Google Scholar]
  18. Gulcher J. R., Nies D. E., Marton L. S., Stefansson K. An alternatively spliced region of the human hexabrachion contains a repeat of potential N-glycosylation sites. Proc Natl Acad Sci U S A. 1989 Mar;86(5):1588–1592. doi: 10.1073/pnas.86.5.1588. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Halfter W., Chiquet-Ehrismann R., Tucker R. P. The effect of tenascin and embryonic basal lamina on the behavior and morphology of neural crest cells in vitro. Dev Biol. 1989 Mar;132(1):14–25. doi: 10.1016/0012-1606(89)90200-5. [DOI] [PubMed] [Google Scholar]
  20. Hoffman S., Crossin K. L., Edelman G. M. Molecular forms, binding functions, and developmental expression patterns of cytotactin and cytotactin-binding proteoglycan, an interactive pair of extracellular matrix molecules. J Cell Biol. 1988 Feb;106(2):519–532. doi: 10.1083/jcb.106.2.519. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Jones F. S., Burgoon M. P., Hoffman S., Crossin K. L., Cunningham B. A., Edelman G. M. A cDNA clone for cytotactin contains sequences similar to epidermal growth factor-like repeats and segments of fibronectin and fibrinogen. Proc Natl Acad Sci U S A. 1988 Apr;85(7):2186–2190. doi: 10.1073/pnas.85.7.2186. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jones F. S., Crossin K. L., Cunningham B. A., Edelman G. M. Identification and characterization of the promoter for the cytotactin gene. Proc Natl Acad Sci U S A. 1990 Sep;87(17):6497–6501. doi: 10.1073/pnas.87.17.6497. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Jones F. S., Hoffman S., Cunningham B. A., Edelman G. M. A detailed structural model of cytotactin: protein homologies, alternative RNA splicing, and binding regions. Proc Natl Acad Sci U S A. 1989 Mar;86(6):1905–1909. doi: 10.1073/pnas.86.6.1905. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Jones F. S., Prediger E. A., Bittner D. A., De Robertis E. M., Edelman G. M. Cell adhesion molecules as targets for Hox genes: neural cell adhesion molecule promoter activity is modulated by cotransfection with Hox-2.5 and -2.4. Proc Natl Acad Sci U S A. 1992 Mar 15;89(6):2086–2090. doi: 10.1073/pnas.89.6.2086. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kessel M., Gruss P. Murine developmental control genes. Science. 1990 Jul 27;249(4967):374–379. doi: 10.1126/science.1974085. [DOI] [PubMed] [Google Scholar]
  26. Kissinger C. R., Liu B. S., Martin-Blanco E., Kornberg T. B., Pabo C. O. Crystal structure of an engrailed homeodomain-DNA complex at 2.8 A resolution: a framework for understanding homeodomain-DNA interactions. Cell. 1990 Nov 2;63(3):579–590. doi: 10.1016/0092-8674(90)90453-l. [DOI] [PubMed] [Google Scholar]
  27. Lee W., Haslinger A., Karin M., Tjian R. Activation of transcription by two factors that bind promoter and enhancer sequences of the human metallothionein gene and SV40. Nature. 1987 Jan 22;325(6102):368–372. doi: 10.1038/325368a0. [DOI] [PubMed] [Google Scholar]
  28. Mackie E. J., Tucker R. P., Halfter W., Chiquet-Ehrismann R., Epperlein H. H. The distribution of tenascin coincides with pathways of neural crest cell migration. Development. 1988 Jan;102(1):237–250. doi: 10.1242/dev.102.1.237. [DOI] [PubMed] [Google Scholar]
  29. Müller M., Affolter M., Leupin W., Otting G., Wüthrich K., Gehring W. J. Isolation and sequence-specific DNA binding of the Antennapedia homeodomain. EMBO J. 1988 Dec 20;7(13):4299–4304. doi: 10.1002/j.1460-2075.1988.tb03328.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Nohno T., Noji S., Koyama E., Ohyama K., Myokai F., Kuroiwa A., Saito T., Taniguchi S. Involvement of the Chox-4 chicken homeobox genes in determination of anteroposterior axial polarity during limb development. Cell. 1991 Mar 22;64(6):1197–1205. doi: 10.1016/0092-8674(91)90274-3. [DOI] [PubMed] [Google Scholar]
  31. Otting G., Qian Y. Q., Billeter M., Müller M., Affolter M., Gehring W. J., Wüthrich K. Protein--DNA contacts in the structure of a homeodomain--DNA complex determined by nuclear magnetic resonance spectroscopy in solution. EMBO J. 1990 Oct;9(10):3085–3092. doi: 10.1002/j.1460-2075.1990.tb07505.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Pearson C. A., Pearson D., Shibahara S., Hofsteenge J., Chiquet-Ehrismann R. Tenascin: cDNA cloning and induction by TGF-beta. EMBO J. 1988 Oct;7(10):2977–2982. doi: 10.1002/j.1460-2075.1988.tb03160.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Prieto A. L., Jones F. S., Cunningham B. A., Crossin K. L., Edelman G. M. Localization during development of alternatively spliced forms of cytotactin mRNA by in situ hybridization. J Cell Biol. 1990 Aug;111(2):685–698. doi: 10.1083/jcb.111.2.685. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Rauscher F. J., 3rd, Sambucetti L. C., Curran T., Distel R. J., Spiegelman B. M. Common DNA binding site for Fos protein complexes and transcription factor AP-1. Cell. 1988 Feb 12;52(3):471–480. doi: 10.1016/s0092-8674(88)80039-4. [DOI] [PubMed] [Google Scholar]
  35. Reuter R., Panganiban G. E., Hoffmann F. M., Scott M. P. Homeotic genes regulate the spatial expression of putative growth factors in the visceral mesoderm of Drosophila embryos. Development. 1990 Dec;110(4):1031–1040. doi: 10.1242/dev.110.4.1031. [DOI] [PubMed] [Google Scholar]
  36. Ruiz i Altaba A., Melton D. A. Interaction between peptide growth factors and homoeobox genes in the establishment of antero-posterior polarity in frog embryos. Nature. 1989 Sep 7;341(6237):33–38. doi: 10.1038/341033a0. [DOI] [PubMed] [Google Scholar]
  37. Ryseck R. P., Macdonald-Bravo H., Mattéi M. G., Ruppert S., Bravo R. Structure, mapping and expression of a growth factor inducible gene encoding a putative nuclear hormonal binding receptor. EMBO J. 1989 Nov;8(11):3327–3335. doi: 10.1002/j.1460-2075.1989.tb08494.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Sassone-Corsi P., Lamph W. W., Kamps M., Verma I. M. fos-associated cellular p39 is related to nuclear transcription factor AP-1. Cell. 1988 Aug 12;54(4):553–560. doi: 10.1016/0092-8674(88)90077-3. [DOI] [PubMed] [Google Scholar]
  39. Spring J., Beck K., Chiquet-Ehrismann R. Two contrary functions of tenascin: dissection of the active sites by recombinant tenascin fragments. Cell. 1989 Oct 20;59(2):325–334. doi: 10.1016/0092-8674(89)90294-8. [DOI] [PubMed] [Google Scholar]
  40. Tan S. S., Crossin K. L., Hoffman S., Edelman G. M. Asymmetric expression in somites of cytotactin and its proteoglycan ligand is correlated with neural crest cell distribution. Proc Natl Acad Sci U S A. 1987 Nov;84(22):7977–7981. doi: 10.1073/pnas.84.22.7977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Tan S. S., Prieto A. L., Newgreen D. F., Crossin K. L., Edelman G. M. Cytotactin expression in somites after dorsal neural tube and neural crest ablation in chicken embryos. Proc Natl Acad Sci U S A. 1991 Aug 1;88(15):6398–6402. doi: 10.1073/pnas.88.15.6398. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Treisman J., Gönczy P., Vashishtha M., Harris E., Desplan C. A single amino acid can determine the DNA binding specificity of homeodomain proteins. Cell. 1989 Nov 3;59(3):553–562. doi: 10.1016/0092-8674(89)90038-x. [DOI] [PubMed] [Google Scholar]
  43. Wehrle B., Chiquet M. Tenascin is accumulated along developing peripheral nerves and allows neurite outgrowth in vitro. Development. 1990 Oct;110(2):401–415. doi: 10.1242/dev.110.2.401. [DOI] [PubMed] [Google Scholar]
  44. Weller A., Beck S., Ekblom P. Amino acid sequence of mouse tenascin and differential expression of two tenascin isoforms during embryogenesis. J Cell Biol. 1991 Jan;112(2):355–362. doi: 10.1083/jcb.112.2.355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Yokouchi Y., Sasaki H., Kuroiwa A. Homeobox gene expression correlated with the bifurcation process of limb cartilage development. Nature. 1991 Oct 3;353(6343):443–445. doi: 10.1038/353443a0. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES