Neuralized-like 1 (Neurl1) Targeted to the Plasma Membrane by N-Myristoylation Regulates the Notch Ligand Jagged1

Abstract

Notch signaling constitutes an evolutionarily conserved mechanism that mediates cell-cell interactions in various developmental processes. Numerous regulatory proteins interact with the Notch receptor and its ligands and control signaling at multiple levels. Ubiquitination and endocytosis followed by endosomal sorting of both the receptor and its ligands is essential for Notch-mediated signaling. The E3 ubiquitin ligases, Neuralized (Neur) and Mind Bomb (Mib1), are crucial for regulating the activity and stability of Notch ligands in Drosophila; however, biochemical evidence that the Notch ligands are directly targeted for ubiquitination by Neur and/or Mib1 has been lacking. In this report, we explore the function of Neurl1, a mouse ortholog of Drosophila Neur. We show that Neurl1 can function as an E3 ubiquitin ligase to activate monoubiquitination in vitro of Jagged1, but not other mammalian Notch ligands. Neurl1 expression decreases Jagged1 levels in cells and blocks signaling from Jagged1-expressing cells to neighboring Notch-expressing cells. We demonstrate that Neurl1 is myristoylated at its N terminus, and that myristoylation of Neurl1 targets it to the plasma membrane. Point mutations abolishing either Neurl1 myristoylation and plasma membrane localization or Neurl1 ubiquitin ligase activity impair its ability to down-regulate Jagged1 expression and to block signaling. Taken together, our results argue that Neurl1 at the plasma membrane can affect the signaling activity of Jagged1 by directly enhancing its ubiquitination and subsequent turnover. Notch signaling constitutes an evolutionarily conserved mechanism that mediates cell-cell interactions in various developmental processes. Numerous regulatory proteins interact with the Notch receptor and its ligands and control signaling at multiple levels. Ubiquitination and endocytosis followed by endosomal sorting of both the receptor and its ligands is essential for Notch-mediated signaling. The E3 ubiquitin ligases, Neuralized (Neur) and Mind Bomb (Mib1), are crucial for regulating the activity and stability of Notch ligands in Drosophila; however, biochemical evidence that the Notch ligands are directly targeted for ubiquitination by Neur and/or Mib1 has been lacking. In this report, we explore the function of Neurl1, a mouse ortholog of Drosophila Neur. We show that Neurl1 can function as an E3 ubiquitin ligase to activate monoubiquitination in vitro of Jagged1, but not other mammalian Notch ligands. Neurl1 expression decreases Jagged1 levels in cells and blocks signaling from Jagged1-expressing cells to neighboring Notch-expressing cells. We demonstrate that Neurl1 is myristoylated at its N terminus, and that myristoylation of Neurl1 targets it to the plasma membrane. Point mutations abolishing either Neurl1 myristoylation and plasma membrane localization or Neurl1 ubiquitin ligase activity impair its ability to down-regulate Jagged1 expression and to block signaling. Taken together, our results argue that Neurl1 at the plasma membrane can affect the signaling activity of Jagged1 by directly enhancing its ubiquitination and subsequent turnover. Cells have evolved a complex machinery of signaling pathways to achieve coordination and fine programming of events that specify different cell types. Notch signaling plays an important role in cellular differentiation, proliferation, and apoptotic events at all stages of development (1Bray S.J. Nat. Rev. Mol. Cell Biol. 2006; 7: 678-689Crossref PubMed Scopus (1984) Google Scholar). Notch proteins are members of a family of evolutionarily conserved transmembrane receptors that bind ligands of the Delta/Serrate/Lag1 (DSL) 5The abbreviations used are: DSLDelta/Serrate/Lag1NHRneuralized homology repeatNeurl1neuralized-like 1HAhemagglutininGFPgreen fluorescent proteinGSTglutathione S-transferase. type presented on juxtaposed cells and specify cell fate in a wide variety of tissues through local cell-cell interactions (2Artavanis-Tsakonas S. Rand M.D. Lake R.J. Science. 1999; 284: 770-776Crossref PubMed Scopus (4918) Google Scholar). Genetic studies in Drosophila and Caenorhabditis elegans indicate that small differences in Notch receptor or ligand expression in adjacent cells are sufficient to alter Notch signaling, resulting in changes in cell fate decisions (3Greenwald I. Genes Dev. 1998; 12: 1751-1762Crossref PubMed Scopus (460) Google Scholar, 4Heitzler P. Simpson P. Cell. 1991; 64: 1083-1092Abstract Full Text PDF PubMed Scopus (586) Google Scholar). Nevertheless, activation of the Notch pathway can also be fine-tuned by additional mechanisms, including post-translational modifications and control of intracellular localization, because dramatic differences in signaling and signal reception between cells do not always correlate with obvious differences in the expression levels of ligands or receptors (5Schweisguth F. Curr. Biol. 2004; 14: R129-R138Abstract Full Text Full Text PDF PubMed Google Scholar). Delta/Serrate/Lag1 neuralized homology repeat neuralized-like 1 hemagglutinin green fluorescent protein glutathione S-transferase. Recent advances have demonstrated essential roles for ubiquitination and endocytosis of receptors and ligands in Notch-mediated signaling (6Le Borgne R. Bardin A. Schweisguth F. Development. 2005; 132: 1751-1762Crossref PubMed Scopus (277) Google Scholar). Key components for the endocytosis of Notch ligands have been identified in Drosophila, zebrafish, and mice. Among these are proteins with general roles in the endocytic pathway, including liquid facets/epsin and auxilin, as well as RING finger ubiquitin ligases Mind bomb (Mib)-1, Mib-2, and Neuralized (Neur) (7Chen W. Corliss D.C. Dev. Biol. 2004; 267: 361-373Crossref PubMed Scopus (55) Google Scholar, 8Hagedorn E.J. Bayraktar J.L. Kandachar V.R. Bai T. Englert D.M. Chang H.C. J. Cell Biol. 2006; 173: 443-452Crossref PubMed Scopus (47) Google Scholar, 9Itoh M. Kim C.-H. Palardy G. Oda T. Jiang Y.-J. Maust D. Yeo S.-Y. Lorick K. Wright G.J. Ariza-McNaughton L. Weissman A.M. Lewis J. Chandrasekharappa S.C. Chitnis A.B. Dev. Cell. 2003; 4: 67-82Abstract Full Text Full Text PDF PubMed Scopus (660) Google Scholar, 10Koo B.-H. Lim H.-S. Song R. Yoon M.-J. Yoon K.-J. Moon J.-S. Kim Y.-W. Kwon M.-C. Yoo K.-W. Kong M.-P. Lee J. Chitnis A.B. Kim C.-H. Kong Y.-Y. Development. 2005; 132: 3459-3470Crossref PubMed Scopus (211) Google Scholar, 11Koo B.-H. Yoon K.-J. Yoo K.-W. Lim H.-S. Song R. Yoon M.-J. Moon J.-S. Kim Y.-W. So J.-H. Kim C.-H. Kong Y.-Y. J. Biol. Chem. 2005; 280: 22335-22342Abstract Full Text Full Text PDF PubMed Scopus (81) Google Scholar, 12Lai E.C. Roegiers F. Qin X. Jan Y.N. Rubin G. Development. 2005; 132: 2319-2332Crossref PubMed Scopus (131) Google Scholar, 13Le Borgne R. Remaud S. Hamel S. Schweisguth F. PLoS. 2005; 3: e96Crossref Scopus (140) Google Scholar, 14Overstreet E. Fitch E. Fischer J.A. Development. 2004; 131: 5355-5366Crossref PubMed Scopus (153) Google Scholar, 15Pitsouli C. Delidakis C. Development. 2005; 132: 4041-4050Crossref PubMed Scopus (95) Google Scholar, 16Wang W. Struhl G. Development. 2004; 131: 5367-5380Crossref PubMed Scopus (205) Google Scholar, 17Wang W. Struhl G. Development. 2005; 132: 2883-2894Crossref PubMed Scopus (148) Google Scholar). In Drosophila, Neur and Mib regulate the Notch ligands Delta and Serrate by inducing their internalization and eventual degradation (9Itoh M. Kim C.-H. Palardy G. Oda T. Jiang Y.-J. Maust D. Yeo S.-Y. Lorick K. Wright G.J. Ariza-McNaughton L. Weissman A.M. Lewis J. Chandrasekharappa S.C. Chitnis A.B. Dev. Cell. 2003; 4: 67-82Abstract Full Text Full Text PDF PubMed Scopus (660) Google Scholar, 10Koo B.-H. Lim H.-S. Song R. Yoon M.-J. Yoon K.-J. Moon J.-S. Kim Y.-W. Kwon M.-C. Yoo K.-W. Kong M.-P. Lee J. Chitnis A.B. Kim C.-H. Kong Y.-Y. Development. 2005; 132: 3459-3470Crossref PubMed Scopus (211) Google Scholar, 12Lai E.C. Roegiers F. Qin X. Jan Y.N. Rubin G. Development. 2005; 132: 2319-2332Crossref PubMed Scopus (131) Google Scholar, 13Le Borgne R. Remaud S. Hamel S. Schweisguth F. PLoS. 2005; 3: e96Crossref Scopus (140) Google Scholar, 15Pitsouli C. Delidakis C. Development. 2005; 132: 4041-4050Crossref PubMed Scopus (95) Google Scholar, 17Wang W. Struhl G. Development. 2005; 132: 2883-2894Crossref PubMed Scopus (148) Google Scholar, 18Lai E.C. Deblandre G.A. Kintner C. Rubin G.M. Dev. Cell. 2001; 1: 783-794Abstract Full Text Full Text PDF PubMed Scopus (288) Google Scholar, 19Pavlopoulos E. Pitsouli C. Klueg K.M. Muskavitch M.A.T. Moschonas N.K. Delidakis C. Dev. Cell. 2001; 1: 1-10Abstract Full Text Full Text PDF PubMed Scopus (226) Google Scholar). Ubiquitination of the ligands seems to be necessary not only for their turnover, but also for downstream trafficking events that render the ligands active for signaling; however, the nature of these downstream events remains unclear (6Le Borgne R. Bardin A. Schweisguth F. Development. 2005; 132: 1751-1762Crossref PubMed Scopus (277) Google Scholar). In mice, there are two Neuralized-like genes, Neurl1 and Neurl2 (20Pavlopoulos E. Kokkinaki M. Koutelou E. Mitsiadis T.A. Prinos P. Delidakis C. Kilpatrick M.W. Tsipouras P. Moschonas N.K. Biochim. Biophys. Acta. 2002; 1574: 375-382Crossref PubMed Scopus (13) Google Scholar, 21Ruan Y. Tecott L. Jiang M.-M. Jan L.Y. Jan Y.N. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 9907-9912Crossref PubMed Scopus (51) Google Scholar, 22Song R. Koo B.-K. Yoon K.-J. Yoon M.-J. Yoo K.-W. Kim H.-T. Oh H.-J. Kim Y.-Y. Han J.-K. Kim C.-H. Kong Y.-Y. J. Biol. Chem. 2006; 281: 36391-36400Abstract Full Text Full Text PDF PubMed Scopus (47) Google Scholar, 23Vollrath B. Pudney J. Asa S. Leder P. Fitzgerald K. Mol. Cell Biol. 2001; 21: 7481-7494Crossref PubMed Scopus (47) Google Scholar). Like Drosophila Neur, they encode proteins that consist of a C-terminal RING finger domain and two neuralized homology repeat (NHR) domains, NHR1 and NHR2. Recently, Neurl2 has been shown to exhibit a nonspecific ubiquitin ligase activity in vitro. Coexpression of Neurl2 with mammalian Delta-like 1 (Dll1) was shown to promote increased Dll1 ubiquitination in cells. In addition, Neurl2 functioned cooperatively with Mib1 to promote endocytosis of XDelta to Hrs (hepatocyte growth factor-regulated tyrosine kinase substrate)-positive early endosomes (22Song R. Koo B.-K. Yoon K.-J. Yoon M.-J. Yoo K.-W. Kim H.-T. Oh H.-J. Kim Y.-Y. Han J.-K. Kim C.-H. Kong Y.-Y. J. Biol. Chem. 2006; 281: 36391-36400Abstract Full Text Full Text PDF PubMed Scopus (47) Google Scholar); however, unequivocal evidence that Dll1 is a direct target of Neurl2 ubiquitin ligase activity is lacking. Although Neurl1 has also been shown to exhibit a nonspecific ubiquitin ligase activity in vitro (22Song R. Koo B.-K. Yoon K.-J. Yoon M.-J. Yoo K.-W. Kim H.-T. Oh H.-J. Kim Y.-Y. Han J.-K. Kim C.-H. Kong Y.-Y. J. Biol. Chem. 2006; 281: 36391-36400Abstract Full Text Full Text PDF PubMed Scopus (47) Google Scholar), putative Neurl1 substrates have not been identified. In addition, whether Neurl1 plays a role in Notch signaling is because exhibit cell fate and two in Notch signaling has been shown to be Y. Tecott L. Jiang M.-M. Jan L.Y. Jan Y.N. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 9907-9912Crossref PubMed Scopus (51) Google Scholar, 23Vollrath B. Pudney J. Asa S. Leder P. Fitzgerald K. Mol. Cell Biol. 2001; 21: 7481-7494Crossref PubMed Scopus (47) Google Scholar). In this report, we demonstrate that Neurl1 is a E3 ubiquitin ligase that can regulate Notch signaling in Neurl1 targets the intracellular domain of Jagged1, but not of other Notch for monoubiquitination in vitro. we show that Neurl1 expression degradation of Jagged1 in cells and blocks signaling from Jagged1-expressing cells to neighboring Notch-expressing cells. we evidence that of Jagged1 and of Notch signaling myristoylation and plasma membrane localization of Neurl1 was from the The Neurl1 was by and or of Neurl1 from Neurl1 and with an or C-terminal or for and from as PubMed Scopus Google Scholar). a mammalian expression Jagged1 ligand J. G. Cell. Full Text PDF Scopus Google was by G. of and Y. S. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google was by P. J. of and was by in of and was by in and of Cell cell from the and as cell Jagged1 or Neurl1 and Jagged1 or cells by and or expression was by with to or followed by and cells in and Cells with of cells in of and by and to and by with either or cells with the and in and for with of in the The with and and proteins with and cells on with in with in and and mouse and a and of in with the expression cells at in with and with cells in and at for 1 at the resulting was with of or for The with and and proteins from the with and or In Ubiquitination type or Neurl1, in and from was for 1 at with or and with or of in and mouse was in cells and by as K. K. T. A. Proc. Natl. Acad. Sci. U. S. A. 1999; PubMed Scopus Google Scholar). with an and a C-terminal in E. and by K. K. T. A. Proc. Natl. Acad. Sci. U. S. A. 1999; PubMed Scopus Google Scholar). and from and and was from In Ubiquitination cells in with and either or Cells in of and by for 1 at proteins by proteins by cells at The cells or cells either or and at a to the of cells. was or of cells used in the of they with the and on was of with the to the the was and the to the in in at activation of is as a of to and in with to and in cells. of Neurl1 by Neurl1 was from cells and by D. Nat. 2001; PubMed Scopus Google Scholar, Chem. 2001; PubMed Scopus Google as in to myristoylated Neurl1 Jagged1 for Ubiquitination in we that Neurl1, in and from cells as a protein of with the and the and whether the activity of Neurl1 is we a of Neurl1 the RING domain and a Neurl1 in two conserved RING domain and have been to NHR1 of in vitro and In a Neurl1 has an RING domain but was active with that an RING domain is sufficient for nonspecific ubiquitin ligase activity S. A.M. S. Weissman A.M. Proc. Natl. Acad. Sci. U. S. A. 1999; PubMed Scopus Google Scholar). In addition, we a in the of at and that Neurl1 can be the role of Neurl1 in ubiquitination of Notch we used a biochemical to whether of the ligands can be directly targeted by mammalian of Jagged1 intracellular domain in and from cells and but not the of or with and to the of a of the of Jagged1 that this was we that it we ubiquitin with has a ubiquitin monoubiquitination of Jagged1 on the of the Neurl1 RING because to ubiquitination Although it from our biochemical that Neurl1 ubiquitin ligase activity is for Jagged1, it is important to that by ubiquitin ligases post-translational of the R. Rev. Cell Dev. Biol. 1999; PubMed Scopus Google Scholar). we the that Neurl1 to ubiquitination of other ligands in our because they are not in cells. The Neurl1 with and Neuralized have been to interact with Delta through the Neur NHR1 domain C. Mol. Biol. Cell. PubMed Scopus Google Scholar). the of Neurl1 for with Jagged1, we Neurl1 or Neurl1 with Jagged1 in cells and and with Jagged1 in cells we to of and or Jagged1 in from or cell The RING domain was for these In of the NHR1 domain with the Jagged1 the was was a small of Jagged1 with NHR1 from a was both or was and Taken together, these results that is for the of Neurl1 with the Jagged1 Neurl1 and to the of the mouse Neurl1 that Neurl1 be myristoylated at its that this is myristoylated in Neurl1 in cells that all or Neurl1 is other Neuralized proteins are myristoylated remains to be however, in with that of the two of Drosophila but not mouse has a myristoylation has a myristoylation Although Drosophila Neur has been to the plasma Neurl1 has been to have a or localization on of Neurl1 at its N to or other (20Pavlopoulos E. Kokkinaki M. Koutelou E. Mitsiadis T.A. Prinos P. Delidakis C. Kilpatrick M.W. Tsipouras P. Moschonas N.K. Biochim. Biophys. Acta. 2002; 1574: 375-382Crossref PubMed Scopus (13) Google Scholar, 21Ruan Y. Tecott L. Jiang M.-M. Jan L.Y. Jan Y.N. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 9907-9912Crossref PubMed Scopus (51) Google Scholar, 23Vollrath B. Pudney J. Asa S. Leder P. Fitzgerald K. Mol. Cell Biol. 2001; 21: 7481-7494Crossref PubMed Scopus (47) Google Scholar), be to with to explore the of Neurl1 we the localization of Neurl1 and a Neurl1 in the was to of we that a of Neurl1 was to the plasma membrane in of cells. In was to the plasma membrane in only of cells and was in in the and of the myristoylation not only Neurl1 localization, but also levels of Neurl1 protein in cells. Drosophila and Neuralized are at levels to degradation by the E.C. Deblandre G.A. Kintner C. Rubin G.M. Dev. Cell. 2001; 1: 783-794Abstract Full Text Full Text PDF PubMed Scopus (288) Google Scholar, C. P. P. Genes Dev. 2002; PubMed Scopus Google Scholar). we that Neurl1 is and at levels by but that it is by of cells with the or of the RING domain a on Neurl1 expression and as well as Neurl1 the NHR1 to levels and that Neurl1 to the plasma membrane by myristoylation is Neurl1 Jagged1 that Neurl1 with Jagged1 and monoubiquitination of the Jagged1 in vitro that Neurl1 Jagged1 and regulate its stability in cells. with that we that expression of Jagged1 with Neurl1, but not Neurl1 with a RING to of Jagged1 protein in cells. In the of and cells with a Jagged1, with or Neurl1 or Neurl1 and the of Jagged1 expression was by with at various Jagged1 was whether or not Neurl1 was In the of Neurl1, Jagged1 levels the of the in cells Neurl1, Jagged1 was by Jagged1 levels in the of that of Jagged1 on Neurl1 ubiquitin ligase activity In addition, can interact with Jagged1 Jagged1 levels in the of of Jagged1 on Neurl1 myristoylation and/or plasma membrane whether Neurl1 expression the ubiquitination of Jagged1 in Jagged1 was with or ubiquitin and Neurl1 or the Neurl1 RING Cell to to proteins that to shown in the of Jagged1 from was increased by of Neurl1, but not the Neurl1 RING The of Jagged1 from cells ubiquitin and ubiquitin has and was that Neurl1 monoubiquitination of Jagged1 in cells. Ubiquitination can as a signal proteins for degradation by the in the of transmembrane for internalization and degradation by the L. Nat. Rev. Mol. Cell Biol. 2001; PubMed Scopus Google Scholar). We the on Jagged1 expression in cells of a and a that blocks to a Cell Biol. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar), and an of function or on Jagged1 to a In Jagged1 expression the of the in the of that of Jagged1 by Neurl1 ubiquitin ligase is on Neurl1 Jagged1 in Notch signaling in cells to the activation of target mammalian of a protein that blocks cellular T. L. Y. J. Cell 2003; PubMed Scopus Google Scholar). directly and the of mouse Neurl1 on Notch signaling, cells with Jagged1 or Jagged1 and cells with and C. P. P. Genes Dev. 2002; PubMed Scopus Google Scholar). of the Notch signaling pathway results in of and intracellular domain the cell and activation of the was was of cells with cells Jagged1 Notch signaling was by expression of Neurl1, because was not of cells with cells with both Jagged1 and with our that Jagged1 was at levels in the of the Neurl1 RING domain or Notch signaling was in cells either of these Neurl1 the results in we Jagged1 expression and activation of the in cells Jagged1 or both Jagged1 and in Neurl1 expression to a dramatic of Jagged1 expression in cells this was to a in Jagged1 because Jagged1 expression be by of cells with the with the results in signaling was in cells Neuralized signaling was by of cells with Although mouse Neurl1 was Y. Tecott L. Jiang M.-M. Jan L.Y. Jan Y.N. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 9907-9912Crossref PubMed Scopus (51) Google Scholar, 23Vollrath B. Pudney J. Asa S. Leder P. Fitzgerald K. Mol. Cell Biol. 2001; 21: 7481-7494Crossref PubMed Scopus (47) Google Scholar), its role in ubiquitination and of Notch pathway components has not been In Drosophila, it is well that the ubiquitin ligase of Neuralized and/or are for and degradation of Notch all on ubiquitination of the ligand intracellular E.C. Roegiers F. Qin X. Jan Y.N. Rubin G. Development. 2005; 132: 2319-2332Crossref PubMed Scopus (131) Google Scholar, 13Le Borgne R. Remaud S. Hamel S. Schweisguth F. PLoS. 2005; 3: e96Crossref Scopus (140) Google Scholar, 15Pitsouli C. Delidakis C. Development. 2005; 132: 4041-4050Crossref PubMed Scopus (95) Google Scholar, 17Wang W. Struhl G. Development. 2005; 132: 2883-2894Crossref PubMed Scopus (148) Google Scholar, M. Pitsouli C. C. Delidakis C. S. J. 2006; Scholar). expression of a Neur was shown to ubiquitination of Dll1 in cells (22Song R. Koo B.-K. Yoon K.-J. Yoon M.-J. Yoo K.-W. Kim H.-T. Oh H.-J. Kim Y.-Y. Han J.-K. Kim C.-H. Kong Y.-Y. J. Biol. Chem. 2006; 281: 36391-36400Abstract Full Text Full Text PDF PubMed Scopus (47) Google Scholar), and a ortholog was shown to promote ubiquitination of in vitro G.A. E.C. Kintner C. Dev. Cell. 2001; 1: Full Text Full Text PDF PubMed Scopus Google Scholar). Nevertheless, the it not been a that a Neuralized protein directly ubiquitination of a we an in vitro ubiquitination with to direct biochemical evidence that Neurl1 as a ubiquitin ligase that monoubiquitination of In addition, we show that of Neurl1 to of Jagged1 in with the that the ubiquitin ligase activity in vitro to Neurl1 activity in cells. Drosophila and mammalian Neur and Mib have been shown to interact with various Notch ligands B.-H. Lim H.-S. Song R. Yoon M.-J. Yoon K.-J. Moon J.-S. Kim Y.-W. Kwon M.-C. Yoo K.-W. Kong M.-P. Lee J. Chitnis A.B. Kim C.-H. Kong Y.-Y. Development. 2005; 132: 3459-3470Crossref PubMed Scopus (211) Google Scholar, 12Lai E.C. Roegiers F. Qin X. Jan Y.N. Rubin G. 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Although we also interactions of Neurl1 with and in mammalian only Jagged1 was in our in vitro and only Jagged1 to Neurl1 the proteins in cells not We do not for this however, it is that the of Neurl1 with and additional proteins and/or on post-translational modifications that are only in mammalian cells. The localization of Neurl1 has with its role as a of Notch ligand ubiquitination and Drosophila neuralized has been shown to be a plasma protein E. Pitsouli C. Klueg K.M. Muskavitch M.A.T. Moschonas N.K. Delidakis C. Dev. Cell. 2001; 1: 1-10Abstract Full Text Full Text PDF PubMed Scopus (226) Google Scholar). In Neurl1 has been to exhibit a and (20Pavlopoulos E. Kokkinaki M. Koutelou E. Mitsiadis T.A. Prinos P. Delidakis C. Kilpatrick M.W. Tsipouras P. Moschonas N.K. Biochim. Biophys. Acta. 2002; 1574: 375-382Crossref PubMed Scopus (13) Google Scholar, 21Ruan Y. Tecott L. Jiang M.-M. Jan L.Y. Jan Y.N. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 9907-9912Crossref PubMed Scopus (51) Google or a T. K. G. T. Mol. Cell 2002; PubMed Scopus Google localization on of evidence that localization of Neurl1 to the plasma membrane its by myristoylation an for this The of and other to the N of proteins protein to the plasma membrane W. S. M.D. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). or post-translational of proteins on an is by T.A. G. J. Biol. Chem. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar, Rev. PubMed Google and only subsequent to of the by an of Neurl1 to an or block with the that the in studies to localization of Neurl1 to the plasma membrane was to the of the we that Neurl1 to the plasma Neurl1, the by not membrane localization of Neurl1 important for Neurl1 ability to interact with the can be from cell with Jagged1, myristoylation is not essential for of Neurl1 with Nevertheless, down-regulate Jagged1 expression in can it block its signaling activity in an that that Jagged1 and be targeted to the plasma membrane they can interact with in cells. of Jagged1 expression and activity that Neurl1 is targeted to plasma membrane by In this it is that are family and the can be targeted to by A. L. J.A. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google Scholar, M.D. Nat. Chem. Biol. 2006; PubMed Scopus Google Scholar, I. S. M. 2006; PubMed Scopus Google Scholar). that Neurl1 expression Jagged1 activity is in of evidence from Drosophila that as a of Notch signaling. The of signaling activity of Jagged1 degradation of cells with that the of ability to signal to Notch is to ligand In other ligand degradation has been shown to with signaling activation Borgne R. Remaud S. Hamel S. Schweisguth F. PLoS. 2005; 3: e96Crossref Scopus (140) Google Scholar, 16Wang W. Struhl G. Development. 2004; 131: 5367-5380Crossref PubMed Scopus (205) Google Scholar, 19Pavlopoulos E. Pitsouli C. Klueg K.M. Muskavitch M.A.T. Moschonas N.K. Delidakis C. Dev. Cell. 2001; 1: 1-10Abstract Full Text Full Text PDF PubMed Scopus (226) Google Scholar). is not whether the ubiquitination that are for ligand activation also to a degradation is that there are two of ubiquitin ligases for of Neur the of Drosophila activation E.C. Deblandre G.A. Kintner C. Rubin G.M. Dev. Cell. 2001; 1: 783-794Abstract Full Text Full Text PDF PubMed Scopus (288) Google Scholar, 19Pavlopoulos E. Pitsouli C. Klueg K.M. Muskavitch M.A.T. Moschonas N.K. Delidakis C. Dev. Cell. 2001; 1: 1-10Abstract Full Text Full Text PDF PubMed Scopus (226) Google Scholar, R. Rev. Cell Dev. Biol. 1999; PubMed Scopus Google Scholar), as mouse Neurl1 down-regulate the to is that the Neuralized ortholog has also been to Notch signaling by levels of G.A. E.C. Kintner C. Dev. Cell. 2001; 1: Full Text Full Text PDF PubMed Scopus Google Scholar). it is that all Neuralized proteins can either activate or on with this in Drosophila expression neuralized Notch Notch activation E.C. Rubin G.M. Dev. Biol. 2001; PubMed Scopus Google Scholar). the of Jagged1 by Neurl1 its function or whether it is also of Notch signaling at levels or in different cellular remains to be We and for and and for Jagged1 and expression We are to Delidakis for and of the with