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. 1997 Oct;17(10):5688–5698. doi: 10.1128/mcb.17.10.5688

Functional analysis of the nuclear LIM domain interactor NLI.

L W Jurata 1, G N Gill 1
PMCID: PMC232417  PMID: 9315627

Abstract

LIM homeodomain and LIM-only (LMO) transcription factors contain two tandemly arranged Zn2+-binding LIM domains capable of mediating protein-protein interactions. These factors have restricted patterns of expression, are found in invertebrates as well as vertebrates, and are required for cell type specification in a variety of developing tissues. A recently identified, widely expressed protein, NLI, binds with high affinity to the LIM domains of LIM homeodomain and LMO proteins in vitro and in vivo. In this study, a 38-amino-acid fragment of NLI was found to be sufficient for the association of NLI with nuclear LIM domains. In addition, NLI was shown to form high affinity homodimers through the amino-terminal 200 amino acids, but dimerization of NLI was not required for association with the LIM homeodomain protein Lmxl. Chemical cross-linking analysis revealed higher-order complexes containing multiple NLI molecules bound to Lmx1, indicating that dimerization of NLI does not interfere with LIM domain interactions. Additionally, NLI formed complexes with Lmx1 on the rat insulin I promoter and inhibited the LIM domain-dependent synergistic transcriptional activation by Lmx1 and the basic helix-loop-helix protein E47 from the rat insulin I minienhancer. These studies indicate that NLI contains at least two functionally independent domains and may serve as a negative regulator of synergistic transcriptional responses which require direct interaction via LIM domains. Thus, NLI may regulate the transcriptional activity of LIM homeodomain proteins by determining specific partner interactions.

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Selected References

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  1. Agulnick A. D., Taira M., Breen J. J., Tanaka T., Dawid I. B., Westphal H. Interactions of the LIM-domain-binding factor Ldb1 with LIM homeodomain proteins. Nature. 1996 Nov 21;384(6606):270–272. doi: 10.1038/384270a0. [DOI] [PubMed] [Google Scholar]
  2. Ahlgren U., Pfaff S. L., Jessell T. M., Edlund T., Edlund H. Independent requirement for ISL1 in formation of pancreatic mesenchyme and islet cells. Nature. 1997 Jan 16;385(6613):257–260. doi: 10.1038/385257a0. [DOI] [PubMed] [Google Scholar]
  3. Arber S., Caroni P. Specificity of single LIM motifs in targeting and LIM/LIM interactions in situ. Genes Dev. 1996 Feb 1;10(3):289–300. doi: 10.1101/gad.10.3.289. [DOI] [PubMed] [Google Scholar]
  4. Bach I., Carrière C., Ostendorff H. P., Andersen B., Rosenfeld M. G. A family of LIM domain-associated cofactors confer transcriptional synergism between LIM and Otx homeodomain proteins. Genes Dev. 1997 Jun 1;11(11):1370–1380. doi: 10.1101/gad.11.11.1370. [DOI] [PubMed] [Google Scholar]
  5. Bach I., Rhodes S. J., Pearse R. V., 2nd, Heinzel T., Gloss B., Scully K. M., Sawchenko P. E., Rosenfeld M. G. P-Lim, a LIM homeodomain factor, is expressed during pituitary organ and cell commitment and synergizes with Pit-1. Proc Natl Acad Sci U S A. 1995 Mar 28;92(7):2720–2724. doi: 10.1073/pnas.92.7.2720. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. 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]
  7. Begley C. G., Lipkowitz S., Göbel V., Mahon K. A., Bertness V., Green A. R., Gough N. M., Kirsch I. R. Molecular characterization of NSCL, a gene encoding a helix-loop-helix protein expressed in the developing nervous system. Proc Natl Acad Sci U S A. 1992 Jan 1;89(1):38–42. doi: 10.1073/pnas.89.1.38. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Blackwell T. K., Weintraub H. Differences and similarities in DNA-binding preferences of MyoD and E2A protein complexes revealed by binding site selection. Science. 1990 Nov 23;250(4984):1104–1110. doi: 10.1126/science.2174572. [DOI] [PubMed] [Google Scholar]
  9. Bourgouin C., Lundgren S. E., Thomas J. B. Apterous is a Drosophila LIM domain gene required for the development of a subset of embryonic muscles. Neuron. 1992 Sep;9(3):549–561. doi: 10.1016/0896-6273(92)90192-g. [DOI] [PubMed] [Google Scholar]
  10. Campos-Ortega J. A. Mechanisms of early neurogenesis in Drosophila melanogaster. J Neurobiol. 1993 Oct;24(10):1305–1327. doi: 10.1002/neu.480241005. [DOI] [PubMed] [Google Scholar]
  11. Cohen B., McGuffin M. E., Pfeifle C., Segal D., Cohen S. M. apterous, a gene required for imaginal disc development in Drosophila encodes a member of the LIM family of developmental regulatory proteins. Genes Dev. 1992 May;6(5):715–729. doi: 10.1101/gad.6.5.715. [DOI] [PubMed] [Google Scholar]
  12. Dawid I. B., Toyama R., Taira M. LIM domain proteins. C R Acad Sci III. 1995 Mar;318(3):295–306. [PubMed] [Google Scholar]
  13. Diaz-Benjumea F. J., Cohen S. M. Interaction between dorsal and ventral cells in the imaginal disc directs wing development in Drosophila. Cell. 1993 Nov 19;75(4):741–752. doi: 10.1016/0092-8674(93)90494-b. [DOI] [PubMed] [Google Scholar]
  14. Freyd G., Kim S. K., Horvitz H. R. Novel cysteine-rich motif and homeodomain in the product of the Caenorhabditis elegans cell lineage gene lin-11. Nature. 1990 Apr 26;344(6269):876–879. doi: 10.1038/344876a0. [DOI] [PubMed] [Google Scholar]
  15. German M. S., Blanar M. A., Nelson C., Moss L. G., Rutter W. J. Two related helix-loop-helix proteins participate in separate cell-specific complexes that bind the insulin enhancer. Mol Endocrinol. 1991 Feb;5(2):292–299. doi: 10.1210/mend-5-2-292. [DOI] [PubMed] [Google Scholar]
  16. German M. S., Moss L. G., Wang J., Rutter W. J. The insulin and islet amyloid polypeptide genes contain similar cell-specific promoter elements that bind identical beta-cell nuclear complexes. Mol Cell Biol. 1992 Apr;12(4):1777–1788. doi: 10.1128/mcb.12.4.1777. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. German M. S., Wang J., Chadwick R. B., Rutter W. J. Synergistic activation of the insulin gene by a LIM-homeo domain protein and a basic helix-loop-helix protein: building a functional insulin minienhancer complex. Genes Dev. 1992 Nov;6(11):2165–2176. doi: 10.1101/gad.6.11.2165. [DOI] [PubMed] [Google Scholar]
  18. German M. S., Wang J. The insulin gene contains multiple transcriptional elements that respond to glucose. Mol Cell Biol. 1994 Jun;14(6):4067–4075. doi: 10.1128/mcb.14.6.4067. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Gill G. N. The enigma of LIM domains. Structure. 1995 Dec 15;3(12):1285–1289. doi: 10.1016/s0969-2126(01)00265-9. [DOI] [PubMed] [Google Scholar]
  20. Guan K. L., Dixon J. E. Eukaryotic proteins expressed in Escherichia coli: an improved thrombin cleavage and purification procedure of fusion proteins with glutathione S-transferase. Anal Biochem. 1991 Feb 1;192(2):262–267. doi: 10.1016/0003-2697(91)90534-z. [DOI] [PubMed] [Google Scholar]
  21. Guillemot F., Lo L. C., Johnson J. E., Auerbach A., Anderson D. J., Joyner A. L. Mammalian achaete-scute homolog 1 is required for the early development of olfactory and autonomic neurons. Cell. 1993 Nov 5;75(3):463–476. doi: 10.1016/0092-8674(93)90381-y. [DOI] [PubMed] [Google Scholar]
  22. Henthorn P., Kiledjian M., Kadesch T. Two distinct transcription factors that bind the immunoglobulin enhancer microE5/kappa 2 motif. Science. 1990 Jan 26;247(4941):467–470. doi: 10.1126/science.2105528. [DOI] [PubMed] [Google Scholar]
  23. Hu J. S., Olson E. N., Kingston R. E. HEB, a helix-loop-helix protein related to E2A and ITF2 that can modulate the DNA-binding ability of myogenic regulatory factors. Mol Cell Biol. 1992 Mar;12(3):1031–1042. doi: 10.1128/mcb.12.3.1031. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Johnson J. D., Zhang W., Rudnick A., Rutter W. J., German M. S. Transcriptional synergy between LIM-homeodomain proteins and basic helix-loop-helix proteins: the LIM2 domain determines specificity. Mol Cell Biol. 1997 Jul;17(7):3488–3496. doi: 10.1128/mcb.17.7.3488. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Jurata L. W., Kenny D. A., Gill G. N. Nuclear LIM interactor, a rhombotin and LIM homeodomain interacting protein, is expressed early in neuronal development. Proc Natl Acad Sci U S A. 1996 Oct 15;93(21):11693–11698. doi: 10.1073/pnas.93.21.11693. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Karlsson O., Edlund T., Moss J. B., Rutter W. J., Walker M. D. A mutational analysis of the insulin gene transcription control region: expression in beta cells is dependent on two related sequences within the enhancer. Proc Natl Acad Sci U S A. 1987 Dec;84(24):8819–8823. doi: 10.1073/pnas.84.24.8819. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Kreider B. L., Benezra R., Rovera G., Kadesch T. Inhibition of myeloid differentiation by the helix-loop-helix protein Id. Science. 1992 Mar 27;255(5052):1700–1702. doi: 10.1126/science.1372755. [DOI] [PubMed] [Google Scholar]
  28. Lai J. S., Herr W. Ethidium bromide provides a simple tool for identifying genuine DNA-independent protein associations. Proc Natl Acad Sci U S A. 1992 Aug 1;89(15):6958–6962. doi: 10.1073/pnas.89.15.6958. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Larson R. C., Lavenir I., Larson T. A., Baer R., Warren A. J., Wadman I., Nottage K., Rabbitts T. H. Protein dimerization between Lmo2 (Rbtn2) and Tal1 alters thymocyte development and potentiates T cell tumorigenesis in transgenic mice. EMBO J. 1996 Mar 1;15(5):1021–1027. [PMC free article] [PubMed] [Google Scholar]
  30. Lassar A. B., Davis R. L., Wright W. E., Kadesch T., Murre C., Voronova A., Baltimore D., Weintraub H. Functional activity of myogenic HLH proteins requires hetero-oligomerization with E12/E47-like proteins in vivo. Cell. 1991 Jul 26;66(2):305–315. doi: 10.1016/0092-8674(91)90620-e. [DOI] [PubMed] [Google Scholar]
  31. Lee J. E., Hollenberg S. M., Snider L., Turner D. L., Lipnick N., Weintraub H. Conversion of Xenopus ectoderm into neurons by NeuroD, a basic helix-loop-helix protein. Science. 1995 May 12;268(5212):836–844. doi: 10.1126/science.7754368. [DOI] [PubMed] [Google Scholar]
  32. Leonard J., Serup P., Gonzalez G., Edlund T., Montminy M. The LIM family transcription factor Isl-1 requires cAMP response element binding protein to promote somatostatin expression in pancreatic islet cells. Proc Natl Acad Sci U S A. 1992 Jul 15;89(14):6247–6251. doi: 10.1073/pnas.89.14.6247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Murre C., McCaw P. S., Baltimore D. A new DNA binding and dimerization motif in immunoglobulin enhancer binding, daughterless, MyoD, and myc proteins. Cell. 1989 Mar 10;56(5):777–783. doi: 10.1016/0092-8674(89)90682-x. [DOI] [PubMed] [Google Scholar]
  34. Murre C., McCaw P. S., Vaessin H., Caudy M., Jan L. Y., Jan Y. N., Cabrera C. V., Buskin J. N., Hauschka S. D., Lassar A. B. Interactions between heterologous helix-loop-helix proteins generate complexes that bind specifically to a common DNA sequence. Cell. 1989 Aug 11;58(3):537–544. doi: 10.1016/0092-8674(89)90434-0. [DOI] [PubMed] [Google Scholar]
  35. Nelson C., Shen L. P., Meister A., Fodor E., Rutter W. J. Pan: a transcriptional regulator that binds chymotrypsin, insulin, and AP-4 enhancer motifs. Genes Dev. 1990 Jun;4(6):1035–1043. doi: 10.1101/gad.4.6.1035. [DOI] [PubMed] [Google Scholar]
  36. Osada H., Grutz G., Axelson H., Forster A., Rabbitts T. H. Association of erythroid transcription factors: complexes involving the LIM protein RBTN2 and the zinc-finger protein GATA1. Proc Natl Acad Sci U S A. 1995 Oct 10;92(21):9585–9589. doi: 10.1073/pnas.92.21.9585. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Peverali F. A., Ramqvist T., Saffrich R., Pepperkok R., Barone M. V., Philipson L. Regulation of G1 progression by E2A and Id helix-loop-helix proteins. EMBO J. 1994 Sep 15;13(18):4291–4301. doi: 10.1002/j.1460-2075.1994.tb06749.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Pfaff S. L., Mendelsohn M., Stewart C. L., Edlund T., Jessell T. M. Requirement for LIM homeobox gene Isl1 in motor neuron generation reveals a motor neuron-dependent step in interneuron differentiation. Cell. 1996 Jan 26;84(2):309–320. doi: 10.1016/s0092-8674(00)80985-x. [DOI] [PubMed] [Google Scholar]
  39. Porcher C., Swat W., Rockwell K., Fujiwara Y., Alt F. W., Orkin S. H. The T cell leukemia oncoprotein SCL/tal-1 is essential for development of all hematopoietic lineages. Cell. 1996 Jul 12;86(1):47–57. doi: 10.1016/s0092-8674(00)80076-8. [DOI] [PubMed] [Google Scholar]
  40. Riddle R. D., Ensini M., Nelson C., Tsuchida T., Jessell T. M., Tabin C. Induction of the LIM homeobox gene Lmx1 by WNT7a establishes dorsoventral pattern in the vertebrate limb. Cell. 1995 Nov 17;83(4):631–640. doi: 10.1016/0092-8674(95)90103-5. [DOI] [PubMed] [Google Scholar]
  41. Robb L., Lyons I., Li R., Hartley L., Köntgen F., Harvey R. P., Metcalf D., Begley C. G. Absence of yolk sac hematopoiesis from mice with a targeted disruption of the scl gene. Proc Natl Acad Sci U S A. 1995 Jul 18;92(15):7075–7079. doi: 10.1073/pnas.92.15.7075. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Shawlot W., Behringer R. R. Requirement for Lim1 in head-organizer function. Nature. 1995 Mar 30;374(6521):425–430. doi: 10.1038/374425a0. [DOI] [PubMed] [Google Scholar]
  43. Shen C. P., Kadesch T. B-cell-specific DNA binding by an E47 homodimer. Mol Cell Biol. 1995 Aug;15(8):4518–4524. doi: 10.1128/mcb.15.8.4518. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Sheng H. Z., Zhadanov A. B., Mosinger B., Jr, Fujii T., Bertuzzi S., Grinberg A., Lee E. J., Huang S. P., Mahon K. A., Westphal H. Specification of pituitary cell lineages by the LIM homeobox gene Lhx3. Science. 1996 May 17;272(5264):1004–1007. doi: 10.1126/science.272.5264.1004. [DOI] [PubMed] [Google Scholar]
  45. 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]
  46. Sánchez-García I., Osada H., Forster A., Rabbitts T. H. The cysteine-rich LIM domains inhibit DNA binding by the associated homeodomain in Isl-1. EMBO J. 1993 Nov;12(11):4243–4250. doi: 10.1002/j.1460-2075.1993.tb06108.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Taira M., Otani H., Saint-Jeannet J. P., Dawid I. B. Role of the LIM class homeodomain protein Xlim-1 in neural and muscle induction by the Spemann organizer in Xenopus. Nature. 1994 Dec 15;372(6507):677–679. doi: 10.1038/372677a0. [DOI] [PubMed] [Google Scholar]
  48. Tsuchida T., Ensini M., Morton S. B., Baldassare M., Edlund T., Jessell T. M., Pfaff S. L. Topographic organization of embryonic motor neurons defined by expression of LIM homeobox genes. Cell. 1994 Dec 16;79(6):957–970. doi: 10.1016/0092-8674(94)90027-2. [DOI] [PubMed] [Google Scholar]
  49. Valge-Archer V. E., Osada H., Warren A. J., Forster A., Li J., Baer R., Rabbitts T. H. The LIM protein RBTN2 and the basic helix-loop-helix protein TAL1 are present in a complex in erythroid cells. Proc Natl Acad Sci U S A. 1994 Aug 30;91(18):8617–8621. doi: 10.1073/pnas.91.18.8617. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Wadman I., Li J., Bash R. O., Forster A., Osada H., Rabbitts T. H., Baer R. Specific in vivo association between the bHLH and LIM proteins implicated in human T cell leukemia. EMBO J. 1994 Oct 17;13(20):4831–4839. doi: 10.1002/j.1460-2075.1994.tb06809.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Walker M. D., Park C. W., Rosen A., Aronheim A. A cDNA from a mouse pancreatic beta cell encoding a putative transcription factor of the insulin gene. Nucleic Acids Res. 1990 Mar 11;18(5):1159–1166. doi: 10.1093/nar/18.5.1159. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Warren A. J., Colledge W. H., Carlton M. B., Evans M. J., Smith A. J., Rabbitts T. H. The oncogenic cysteine-rich LIM domain protein rbtn2 is essential for erythroid development. Cell. 1994 Jul 15;78(1):45–57. doi: 10.1016/0092-8674(94)90571-1. [DOI] [PubMed] [Google Scholar]
  53. Way J. C., Chalfie M. mec-3, a homeobox-containing gene that specifies differentiation of the touch receptor neurons in C. elegans. Cell. 1988 Jul 1;54(1):5–16. doi: 10.1016/0092-8674(88)90174-2. [DOI] [PubMed] [Google Scholar]
  54. Wu R. Y., Gill G. N. LIM domain recognition of a tyrosine-containing tight turn. J Biol Chem. 1994 Oct 7;269(40):25085–25090. [PubMed] [Google Scholar]
  55. Wu R., Durick K., Songyang Z., Cantley L. C., Taylor S. S., Gill G. N. Specificity of LIM domain interactions with receptor tyrosine kinases. J Biol Chem. 1996 Jul 5;271(27):15934–15941. doi: 10.1074/jbc.271.27.15934. [DOI] [PubMed] [Google Scholar]
  56. Xue D., Tu Y., Chalfie M. Cooperative interactions between the Caenorhabditis elegans homeoproteins UNC-86 and MEC-3. Science. 1993 Sep 3;261(5126):1324–1328. doi: 10.1126/science.8103239. [DOI] [PubMed] [Google Scholar]
  57. 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]

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