Skip to main content
PMC home page
The EMBO Journal logoLink to The EMBO Journal
. 1998 Nov 2;17(21):6276–6288. doi: 10.1093/emboj/17.21.6276

Requirement of the mouse I-mfa gene for placental development and skeletal patterning.

N Kraut 1, L Snider 1, C M Chen 1, S J Tapscott 1, M Groudine 1
PMCID: PMC1170953  PMID: 9799236

Abstract

The bHLH-repressor protein I-mfa binds to MyoD family members, inhibits their activity, and blocks their nuclear import and binding to DNA. In situ hybridization analysis demonstrated that mouse I-mfa was highly expressed in extraembryonic lineages, in the sclerotome, and subsequently within mesenchymal precursors of the axial and appendicular skeleton, before chondrogenesis occurs. Targeted deletion of I-mfa in a C57Bl/6 background resulted in embryonic lethality around E10.5, associated with a placental defect and a markedly reduced number of trophoblast giant cells. Overexpression of I-mfa in rat trophoblast (Rcho-1) stem cells induced differentiation into trophoblast giant cells. I-mfa interacted with the bHLH protein Mash2, a negative regulator of trophoblast giant cell formation, and inhibited its transcriptional activity in cell culture. In contrast, I-mfa did not interfere with the activity of the bHLH protein Hand1, a positive regulator of giant cell differentiation. Interestingly, I-mfa-null embryos on a 129/Sv background had no placental defect, generally survived to adulthood, and exhibited delayed caudal neural tube closure and skeletal patterning defects that included fusions of ribs, vertebral bodies and abnormal formation of spinous processes. Our results indicate that I-mfa plays an important role in trophoblast and chondrogenic differentiation by negatively regulating a subset of lineage-restricted bHLH proteins.

Full Text

The Full Text of this article is available as a PDF (1.1 MB).

Selected References

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

  1. Bain G., Maandag E. C., Izon D. J., Amsen D., Kruisbeek A. M., Weintraub B. C., Krop I., Schlissel M. S., Feeney A. J., van Roon M. E2A proteins are required for proper B cell development and initiation of immunoglobulin gene rearrangements. Cell. 1994 Dec 2;79(5):885–892. doi: 10.1016/0092-8674(94)90077-9. [DOI] [PubMed] [Google Scholar]
  2. Braun T., Rudnicki M. A., Arnold H. H., Jaenisch R. Targeted inactivation of the muscle regulatory gene Myf-5 results in abnormal rib development and perinatal death. Cell. 1992 Oct 30;71(3):369–382. doi: 10.1016/0092-8674(92)90507-9. [DOI] [PubMed] [Google Scholar]
  3. Burgess R., Rawls A., Brown D., Bradley A., Olson E. N. Requirement of the paraxis gene for somite formation and musculoskeletal patterning. Nature. 1996 Dec 12;384(6609):570–573. doi: 10.1038/384570a0. [DOI] [PubMed] [Google Scholar]
  4. Candia A. F., Hu J., Crosby J., Lalley P. A., Noden D., Nadeau J. H., Wright C. V. Mox-1 and Mox-2 define a novel homeobox gene subfamily and are differentially expressed during early mesodermal patterning in mouse embryos. Development. 1992 Dec;116(4):1123–1136. doi: 10.1242/dev.116.4.1123. [DOI] [PubMed] [Google Scholar]
  5. Carney E. W., Prideaux V., Lye S. J., Rossant J. Progressive expression of trophoblast-specific genes during formation of mouse trophoblast giant cells in vitro. Mol Reprod Dev. 1993 Apr;34(4):357–368. doi: 10.1002/mrd.1080340403. [DOI] [PubMed] [Google Scholar]
  6. Chen C. M., Kraut N., Groudine M., Weintraub H. I-mf, a novel myogenic repressor, interacts with members of the MyoD family. Cell. 1996 Sep 6;86(5):731–741. doi: 10.1016/s0092-8674(00)80148-8. [DOI] [PubMed] [Google Scholar]
  7. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  8. Copp A. J. Death before birth: clues from gene knockouts and mutations. Trends Genet. 1995 Mar;11(3):87–93. doi: 10.1016/S0168-9525(00)89008-3. [DOI] [PubMed] [Google Scholar]
  9. Copp A. J. Genetic models of mammalian neural tube defects. Ciba Found Symp. 1994;181:118–143. doi: 10.1002/9780470514559.ch8. [DOI] [PubMed] [Google Scholar]
  10. Cross J. C., Flannery M. L., Blanar M. A., Steingrimsson E., Jenkins N. A., Copeland N. G., Rutter W. J., Werb Z. Hxt encodes a basic helix-loop-helix transcription factor that regulates trophoblast cell development. Development. 1995 Aug;121(8):2513–2523. doi: 10.1242/dev.121.8.2513. [DOI] [PubMed] [Google Scholar]
  11. Cross J. C., Werb Z., Fisher S. J. Implantation and the placenta: key pieces of the development puzzle. Science. 1994 Dec 2;266(5190):1508–1518. doi: 10.1126/science.7985020. [DOI] [PubMed] [Google Scholar]
  12. Cserjesi P., Brown D., Ligon K. L., Lyons G. E., Copeland N. G., Gilbert D. J., Jenkins N. A., Olson E. N. Scleraxis: a basic helix-loop-helix protein that prefigures skeletal formation during mouse embryogenesis. Development. 1995 Apr;121(4):1099–1110. doi: 10.1242/dev.121.4.1099. [DOI] [PubMed] [Google Scholar]
  13. Cserjesi P., Brown D., Lyons G. E., Olson E. N. Expression of the novel basic helix-loop-helix gene eHAND in neural crest derivatives and extraembryonic membranes during mouse development. Dev Biol. 1995 Aug;170(2):664–678. doi: 10.1006/dbio.1995.1245. [DOI] [PubMed] [Google Scholar]
  14. Deutsch U., Dressler G. R., Gruss P. Pax 1, a member of a paired box homologous murine gene family, is expressed in segmented structures during development. Cell. 1988 May 20;53(4):617–625. doi: 10.1016/0092-8674(88)90577-6. [DOI] [PubMed] [Google Scholar]
  15. Ellmeier W., Weith A. Expression of the helix-loop-helix gene Id3 during murine embryonic development. Dev Dyn. 1995 Jun;203(2):163–173. doi: 10.1002/aja.1002030205. [DOI] [PubMed] [Google Scholar]
  16. Faria T. N., Soares M. J. Trophoblast cell differentiation: establishment, characterization, and modulation of a rat trophoblast cell line expressing members of the placental prolactin family. Endocrinology. 1991 Dec;129(6):2895–2906. doi: 10.1210/endo-129-6-2895. [DOI] [PubMed] [Google Scholar]
  17. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  18. Firulli A. B., McFadden D. G., Lin Q., Srivastava D., Olson E. N. Heart and extra-embryonic mesodermal defects in mouse embryos lacking the bHLH transcription factor Hand1. Nat Genet. 1998 Mar;18(3):266–270. doi: 10.1038/ng0398-266. [DOI] [PubMed] [Google Scholar]
  19. Gavrieli Y., Sherman Y., Ben-Sasson S. A. Identification of programmed cell death in situ via specific labeling of nuclear DNA fragmentation. J Cell Biol. 1992 Nov;119(3):493–501. doi: 10.1083/jcb.119.3.493. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Goulding M. D., Chalepakis G., Deutsch U., Erselius J. R., Gruss P. Pax-3, a novel murine DNA binding protein expressed during early neurogenesis. EMBO J. 1991 May;10(5):1135–1147. doi: 10.1002/j.1460-2075.1991.tb08054.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Guillemot F., Nagy A., Auerbach A., Rossant J., Joyner A. L. Essential role of Mash-2 in extraembryonic development. Nature. 1994 Sep 22;371(6495):333–336. doi: 10.1038/371333a0. [DOI] [PubMed] [Google Scholar]
  22. Hollenberg S. M., Sternglanz R., Cheng P. F., Weintraub H. Identification of a new family of tissue-specific basic helix-loop-helix proteins with a two-hybrid system. Mol Cell Biol. 1995 Jul;15(7):3813–3822. doi: 10.1128/mcb.15.7.3813. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hurlin P. J., Quéva C., Koskinen P. J., Steingrímsson E., Ayer D. E., Copeland N. G., Jenkins N. A., Eisenman R. N. Mad3 and Mad4: novel Max-interacting transcriptional repressors that suppress c-myc dependent transformation and are expressed during neural and epidermal differentiation. EMBO J. 1995 Nov 15;14(22):5646–5659. doi: 10.1002/j.1460-2075.1995.tb00252.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Ilgren E. B. Review article: control of trophoblastic growth. Placenta. 1983 Jul-Sep;4(3):307–328. doi: 10.1016/s0143-4004(83)80010-1. [DOI] [PubMed] [Google Scholar]
  25. Jan Y. N., Jan L. Y. HLH proteins, fly neurogenesis, and vertebrate myogenesis. Cell. 1993 Dec 3;75(5):827–830. doi: 10.1016/0092-8674(93)90525-u. [DOI] [PubMed] [Google Scholar]
  26. Jen Y., Weintraub H., Benezra R. Overexpression of Id protein inhibits the muscle differentiation program: in vivo association of Id with E2A proteins. Genes Dev. 1992 Aug;6(8):1466–1479. doi: 10.1101/gad.6.8.1466. [DOI] [PubMed] [Google Scholar]
  27. Kraut N., Frampton J., McNagny K. M., Graf T. A functional Ets DNA-binding domain is required to maintain multipotency of hematopoietic progenitors transformed by Myb-Ets. Genes Dev. 1994 Jan;8(1):33–44. doi: 10.1101/gad.8.1.33. [DOI] [PubMed] [Google Scholar]
  28. Kraut N. The gene encoding I-mf (Mdfi) maps to human chromosome 6p21 and mouse chromosome 17. Mamm Genome. 1997 Aug;8(8):618–619. doi: 10.1007/s003359900517. [DOI] [PubMed] [Google Scholar]
  29. Lee J. E. Basic helix-loop-helix genes in neural development. Curr Opin Neurobiol. 1997 Feb;7(1):13–20. doi: 10.1016/s0959-4388(97)80115-8. [DOI] [PubMed] [Google Scholar]
  30. Lescisin K. R., Varmuza S., Rossant J. Isolation and characterization of a novel trophoblast-specific cDNA in the mouse. Genes Dev. 1988 Dec;2(12A):1639–1646. doi: 10.1101/gad.2.12a.1639. [DOI] [PubMed] [Google Scholar]
  31. Liu Y., Watanabe H., Nifuji A., Yamada Y., Olson E. N., Noda M. Overexpression of a single helix-loop-helix-type transcription factor, scleraxis, enhances aggrecan gene expression in osteoblastic osteosarcoma ROS17/2.8 cells. J Biol Chem. 1997 Nov 21;272(47):29880–29885. doi: 10.1074/jbc.272.47.29880. [DOI] [PubMed] [Google Scholar]
  32. Nagy A., Gócza E., Diaz E. M., Prideaux V. R., Iványi E., Markkula M., Rossant J. Embryonic stem cells alone are able to support fetal development in the mouse. Development. 1990 Nov;110(3):815–821. doi: 10.1242/dev.110.3.815. [DOI] [PubMed] [Google Scholar]
  33. Neubüser A., Koseki H., Balling R. Characterization and developmental expression of Pax9, a paired-box-containing gene related to Pax1. Dev Biol. 1995 Aug;170(2):701–716. doi: 10.1006/dbio.1995.1248. [DOI] [PubMed] [Google Scholar]
  34. Riley P., Anson-Cartwright L., Cross J. C. The Hand1 bHLH transcription factor is essential for placentation and cardiac morphogenesis. Nat Genet. 1998 Mar;18(3):271–275. doi: 10.1038/ng0398-271. [DOI] [PubMed] [Google Scholar]
  35. Rudnicki M. A., Jaenisch R. The MyoD family of transcription factors and skeletal myogenesis. Bioessays. 1995 Mar;17(3):203–209. doi: 10.1002/bies.950170306. [DOI] [PubMed] [Google Scholar]
  36. Rupp R. A., Weintraub H. Ubiquitous MyoD transcription at the midblastula transition precedes induction-dependent MyoD expression in presumptive mesoderm of X. laevis. Cell. 1991 Jun 14;65(6):927–937. doi: 10.1016/0092-8674(91)90545-a. [DOI] [PubMed] [Google Scholar]
  37. Sasai Y., Kageyama R., Tagawa Y., Shigemoto R., Nakanishi S. Two mammalian helix-loop-helix factors structurally related to Drosophila hairy and Enhancer of split. Genes Dev. 1992 Dec;6(12B):2620–2634. doi: 10.1101/gad.6.12b.2620. [DOI] [PubMed] [Google Scholar]
  38. Shida M. M., Ng Y. K., Soares M. J., Linzer D. I. Trophoblast-specific transcription from the mouse placental lactogen-I gene promoter. Mol Endocrinol. 1993 Feb;7(2):181–188. doi: 10.1210/mend.7.2.8469232. [DOI] [PubMed] [Google Scholar]
  39. Shivdasani R. A., Mayer E. L., Orkin S. H. Absence of blood formation in mice lacking the T-cell leukaemia oncoprotein tal-1/SCL. Nature. 1995 Feb 2;373(6513):432–434. doi: 10.1038/373432a0. [DOI] [PubMed] [Google Scholar]
  40. Sibilia M., Wagner E. F. Strain-dependent epithelial defects in mice lacking the EGF receptor. Science. 1995 Jul 14;269(5221):234–238. doi: 10.1126/science.7618085. [DOI] [PubMed] [Google Scholar]
  41. Soriano P., Montgomery C., Geske R., Bradley A. Targeted disruption of the c-src proto-oncogene leads to osteopetrosis in mice. Cell. 1991 Feb 22;64(4):693–702. doi: 10.1016/0092-8674(91)90499-o. [DOI] [PubMed] [Google Scholar]
  42. Spicer D. B., Rhee J., Cheung W. L., Lassar A. B. Inhibition of myogenic bHLH and MEF2 transcription factors by the bHLH protein Twist. Science. 1996 Jun 7;272(5267):1476–1480. doi: 10.1126/science.272.5267.1476. [DOI] [PubMed] [Google Scholar]
  43. Srivastava D., Thomas T., Lin Q., Kirby M. L., Brown D., Olson E. N. Regulation of cardiac mesodermal and neural crest development by the bHLH transcription factor, dHAND. Nat Genet. 1997 Jun;16(2):154–160. doi: 10.1038/ng0697-154. [DOI] [PubMed] [Google Scholar]
  44. Tanaka M., Gertsenstein M., Rossant J., Nagy A. Mash2 acts cell autonomously in mouse spongiotrophoblast development. Dev Biol. 1997 Oct 1;190(1):55–65. doi: 10.1006/dbio.1997.8685. [DOI] [PubMed] [Google Scholar]
  45. Threadgill D. W., Dlugosz A. A., Hansen L. A., Tennenbaum T., Lichti U., Yee D., LaMantia C., Mourton T., Herrup K., Harris R. C. Targeted disruption of mouse EGF receptor: effect of genetic background on mutant phenotype. Science. 1995 Jul 14;269(5221):230–234. doi: 10.1126/science.7618084. [DOI] [PubMed] [Google Scholar]
  46. Turner D. L., Weintraub H. Expression of achaete-scute homolog 3 in Xenopus embryos converts ectodermal cells to a neural fate. Genes Dev. 1994 Jun 15;8(12):1434–1447. doi: 10.1101/gad.8.12.1434. [DOI] [PubMed] [Google Scholar]
  47. Tybulewicz V. L., Crawford C. E., Jackson P. K., Bronson R. T., Mulligan R. C. Neonatal lethality and lymphopenia in mice with a homozygous disruption of the c-abl proto-oncogene. Cell. 1991 Jun 28;65(7):1153–1163. doi: 10.1016/0092-8674(91)90011-m. [DOI] [PubMed] [Google Scholar]
  48. Wallin J., Wilting J., Koseki H., Fritsch R., Christ B., Balling R. The role of Pax-1 in axial skeleton development. Development. 1994 May;120(5):1109–1121. doi: 10.1242/dev.120.5.1109. [DOI] [PubMed] [Google Scholar]
  49. Wang Y., Benezra R., Sassoon D. A. Id expression during mouse development: a role in morphogenesis. Dev Dyn. 1992 Jul;194(3):222–230. doi: 10.1002/aja.1001940307. [DOI] [PubMed] [Google Scholar]
  50. Weintraub H., Davis R., Tapscott S., Thayer M., Krause M., Benezra R., Blackwell T. K., Turner D., Rupp R., Hollenberg S. The myoD gene family: nodal point during specification of the muscle cell lineage. Science. 1991 Feb 15;251(4995):761–766. doi: 10.1126/science.1846704. [DOI] [PubMed] [Google Scholar]
  51. Weintraub H. The MyoD family and myogenesis: redundancy, networks, and thresholds. Cell. 1993 Dec 31;75(7):1241–1244. doi: 10.1016/0092-8674(93)90610-3. [DOI] [PubMed] [Google Scholar]
  52. Wolf C., Thisse C., Stoetzel C., Thisse B., Gerlinger P., Perrin-Schmitt F. The M-twist gene of Mus is expressed in subsets of mesodermal cells and is closely related to the Xenopus X-twi and the Drosophila twist genes. Dev Biol. 1991 Feb;143(2):363–373. doi: 10.1016/0012-1606(91)90086-i. [DOI] [PubMed] [Google Scholar]
  53. Yoon J. K., Olson E. N., Arnold H. H., Wold B. J. Different MRF4 knockout alleles differentially disrupt Myf-5 expression: cis-regulatory interactions at the MRF4/Myf-5 locus. Dev Biol. 1997 Aug 15;188(2):349–362. doi: 10.1006/dbio.1997.8670. [DOI] [PubMed] [Google Scholar]
  54. Zhuang Y., Soriano P., Weintraub H. The helix-loop-helix gene E2A is required for B cell formation. Cell. 1994 Dec 2;79(5):875–884. doi: 10.1016/0092-8674(94)90076-0. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES