Drosophila neuralized is a ubiquitin ligase that promotes the internalization and degradation of delta.

The Drosophila gene neuralized (neur) has long been recognized to be essential for the proper execution of a wide variety of processes mediated by the Notch (N) pathway, but its role in the pathway has been elusive. In this report, we present genetic and biochemical evidence that Neur is a RING-type, E3 ubiquitin ligase. Next, we show that neur is required for proper internalization of Dl in the developing eye. Finally, we demonstrate that ectopic Neur targets Dl for internalization and degradation in a RING finger-dependent manner, and that the two exist in a physical complex. Collectively, our data indicate that Neur is a ubiquitin ligase that positively regulates the N pathway by promoting the endocytosis and degradation of Dl.

[1]  S. Blair,et al.  The function and regulation of cut expression on the wing margin of Drosophila: Notch, Wingless and a dominant negative role for Delta and Serrate. , 1997, Development.

[2]  P. Simpson,et al.  Altered epidermal growth factor-like sequences provide evidence for a role of Notch as a receptor in cell fate decisions. , 1993, Development.

[3]  Matthew Freeman,et al.  Sprouty, an Intracellular Inhibitor of Ras Signaling , 1999, Cell.

[4]  J. Campos-Ortega,et al.  The Enhancer of split complex and adjacent genes in the 96F region of Drosophila melanogaster are required for segregation of neural and epidermal progenitor cells. , 1992, Genetics.

[5]  T. P. Neufeld,et al.  A genetic screen to identify components of the sina signaling pathway in Drosophila eye development. , 1998, Genetics.

[6]  T. L. Jacobsen,et al.  Feedback regulation is central to Delta-Notch signalling required for Drosophila wing vein morphogenesis. , 1997, Development.

[7]  R. Fehon,et al.  Implications of dynamic patterns of Delta and Notch expression for cellular interactions during Drosophila development. , 1993, Development.

[8]  F. Schweisguth,et al.  Dominant-negative mutation in the beta2 and beta6 proteasome subunit genes affect alternative cell fate decisions in the Drosophila sense organ lineage. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[9]  S. Artavanis-Tsakonas,et al.  The intracellular deletions of Delta and Serrate define dominant negative forms of the Drosophila Notch ligands. , 1996, Development.

[10]  A. Weissman,et al.  RING Finger Proteins Mediators of Ubiquitin Ligase Activity , 2000, Cell.

[11]  J. Campos-Ortega,et al.  The basic-helix-loop-helix domain of Drosophila lethal of scute protein is sufficient for proneural function and activates neurogenic genes , 1994, Cell.

[12]  V. Hartenstein,et al.  The function of the neurogenic genes during epithelial development in the Drosophila embryo. , 1992, Development.

[13]  Y. Li,et al.  Photoreceptor Cell Differentiation Requires Regulated Proteolysis of the Transcriptional Repressor Tramtrack , 1997, Cell.

[14]  L. Hicke.  A New Ticket for Entry into Budding Vesicles—Ubiquitin , 2001, Cell.

[15]  S. Ōmura,et al.  Degradation Process of Ligand-stimulated Platelet-derived Growth Factor β -Receptor Involves Ubiquitin-Proteasome Proteolytic Pathway * , 1995, The Journal of Biological Chemistry.

[16]  T. P. Neufeld,et al.  PHYL Acts to Down-Regulate TTK88, a Transcriptional Repressor of Neuronal Cell Fates, by a SINA-Dependent Mechanism , 1997, Cell.

[17]  A. Ciechanover,et al.  The ubiquitin conjugation system is required for ligand‐induced endocytosis and degradation of the growth hormone receptor. , 1996, The EMBO journal.

[18]  S. Schmid,et al.  Clathrin-independent pinocytosis is induced in cells overexpressing a temperature-sensitive mutant of dynamin , 1995, The Journal of cell biology.

[19]  J. Belote,et al.  Identification of an essential gene, l(3)73Ai, with a dominant temperature-sensitive lethal allele, encoding a Drosophila proteasome subunit. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[20]  L. Neckers,et al.  Polyubiquitination and Proteasomal Degradation of the p185c-erbB-2 Receptor Protein-tyrosine Kinase Induced by Geldanamycin* , 1996, The Journal of Biological Chemistry.

[21]  A. Michelson,et al.  A role for the Drosophila neurogenic genes in mesoderm differentiation , 1991, Cell.

[22]  P. Simpson,et al.  Requirement for dynamin during Notch signaling in Drosophila neurogenesis. , 1997, Developmental biology.

[23]  S. Ōmura,et al.  Degradation of the Met tyrosine kinase receptor by the ubiquitin-proteasome pathway , 1997, Molecular and cellular biology.

[24]  D A Lauffenburger,et al.  The role of tyrosine kinase activity in endocytosis, compartmentation, and down-regulation of the epidermal growth factor receptor. , 1991, The Journal of biological chemistry.

[25]  D. Rotin,et al.  Ubiquitination and Endocytosis of Plasma Membrane Proteins: Role of Nedd4/Rsp5p Family of Ubiquitin-Protein Ligases , 2000, The Journal of Membrane Biology.

[26]  A. Parks,et al.  Ligand endocytosis drives receptor dissociation and activation in the Notch pathway. , 2000, Development.

[27]  Peter Jackson,et al.  The lore of the RINGs: substrate recognition and catalysis by ubiquitin ligases. , 2000, Trends in cell biology.

[28]  E. Lai,et al.  Xenopus neuralized is a ubiquitin ligase that interacts with XDelta1 and regulates Notch signaling. , 2001, Developmental cell.

[29]  Y. Jan,et al.  The Drosophila neurogenic gene neuralized encodes a novel protein and is expressed in precursors of larval and adult neurons. , 1991, The EMBO journal.

[30]  T. Hunter,et al.  The tyrosine kinase negative regulator c-Cbl as a RING-type, E2-dependent ubiquitin-protein ligase. , 1999, Science.

[31]  M. Muskavitch,et al.  Complex proteolytic processing acts on Delta, a transmembrane ligand for Notch, during Drosophila development. , 1998, Molecular biology of the cell.

[32]  I. Guerrero,et al.  Targeted expression of the signaling molecule decapentaplegic induces pattern duplications and growth alterations in Drosophila wings. , 1994, The EMBO journal.

[33]  Howard Riezman,et al.  Ubiquitination of a Yeast Plasma Membrane Receptor Signals Its Ligand-Stimulated Endocytosis , 1996, Cell.

[34]  P. Simpson,et al.  The choice of cell fate in the epidermis of Drosophila , 1991, Cell.

[35]  O. Staub,et al.  Regulation of stability and function of the epithelial Na+ channel (ENaC) by ubiquitination , 1997, The EMBO journal.

[36]  N. Baker,et al.  The R8-photoreceptor equivalence group in Drosophila: fate choice precedes regulated Delta transcription and is independent of Notch gene dose , 1998, Mechanisms of Development.

[37]  A. Laughon,et al.  The Drosophila neuralized gene encodes a C3HC4 zinc finger. , 1993, The EMBO journal.

[38]  T. L. Jacobsen,et al.  Cis-interactions between Delta and Notch modulate neurogenic signalling in Drosophila. , 1998, Development.

[39]  F. Turner,et al.  Relationships between complex Delta expression and the specification of retinal cell fates during Drosophila eye development , 1995, Mechanisms of Development.

[40]  P. Hieter,et al.  The APC11 RING-H2 finger mediates E2-dependent ubiquitination. , 2000, Molecular biology of the cell.

[41]  S. Artavanis-Tsakonas,et al.  Notch signaling: cell fate control and signal integration in development. , 1999, Science.

[42]  G. Rubin,et al.  neuralized is essential for a subset of Notch pathway-dependent cell fate decisions during Drosophila eye development , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[43]  C. Poodry.  shibire, a neurogenic mutant of Drosophila. , 1990, Developmental biology.

[44]  S. Barolo,et al.  GFP and beta-galactosidase transformation vectors for promoter/enhancer analysis in Drosophila. , 2000, BioTechniques.

[45]  R. J. Fleming,et al.  Specific EGF repeats of Notch mediate interactions with Delta and serrate: Implications for notch as a multifunctional receptor , 1991, Cell.

[46]  A. Carmena,et al.  Neurogenic genes control gene expression at the transcriptional level in early neurogenesis and in mesectoderm specification. , 1995, Development.

[47]  Y. Jan,et al.  Ethanol hypersensitivity and olfactory discrimination defect in mice lacking a homolog of Drosophila neuralized , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[48]  G. Boulianne,et al.  Neuralized functions cell autonomously to regulate Drosophila sense organ development , 2000, The EMBO journal.

[49]  N. Perrimon,et al.  Targeted gene expression as a means of altering cell fates and generating dominant phenotypes. , 1993, Development.

[50]  P. Simpson,et al.  Genes of the Enhancer of split and achaete-scute complexes are required for a regulatory loop between Notch and Delta during lateral signalling in Drosophila. , 1996, Development.

[51]  M. Vidal,et al.  Mammalian homologs of seven in absentia regulate DCC via the ubiquitin-proteasome pathway. , 1997, Genes & development.

[52]  G. Rubin,et al.  neuralized functions cell-autonomously to regulate a subset of notch-dependent processes during adult Drosophila development. , 2001, Developmental biology.

[53]  C. Wilson,et al.  P-element-mediated enhancer detection: a versatile method to study development in Drosophila. , 1989, Genes & development.

[54]  A. Brand,et al.  Specificity of bone morphogenetic protein-related factors: cell fate and gene expression changes in Drosophila embryos induced by decapentaplegic but not 60A. , 1994, Cell growth & differentiation : the molecular biology journal of the American Association for Cancer Research.

[55]  Mitsuyoshi Nakao,et al.  Identification of a human homolog of the Drosophila neuralized gene within the 10q25.1 malignant astrocytoma deletion region , 1998, Oncogene.

[56]  M. W. Young,et al.  Antineurogenic phenotypes induced by truncated Notch proteins indicate a role in signal transduction and may point to a novel function for Notch in nuclei. , 1993, Genes & development.

[57]  J. Bonifacino,et al.  Ubiquitin and the control of protein fate in the secretory and endocytic pathways. , 1998, Annual review of cell and developmental biology.

[58]  G. Struhl,et al.  Nuclear Access and Action of Notch In Vivo , 1998, Cell.

[59]  S. Fang,et al.  RING fingers mediate ubiquitin-conjugating enzyme (E2)-dependent ubiquitination. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[60]  M. Freeman,et al.  Reiterative Use of the EGF Receptor Triggers Differentiation of All Cell Types in the Drosophila Eye , 1996, Cell.