Involvement of a proline-rich motif and RING-H2 finger of Deltex in the regulation of Notch signaling.
The Notch pathway is an evolutionarily conserved signaling mechanism that is essential for cell-cell interactions. The Drosophila deltex gene regulates Notch signaling in a positive manner, and its gene product physically interacts with the intracellular domain of Notch through its N-terminal domain. Deltex has two other domains that are presumably involved in protein-protein interactions: a proline-rich motif that binds to SH3-domains, and a RING-H2 finger motif. Using an overexpression assay, we have analyzed the functional involvement of these Deltex domains in Notch signaling. The N-terminal domain of Deltex that binds to the CDC10/Ankyrin repeats of the Notch intracellular domain was indispensable for the function of Deltex. A mutant form of Deltex that lacked the proline-rich motif behaved as a dominant-negative form. This dominant-negative Deltex inhibited Notch signaling upstream of an activated, nuclear form of Notch and downstream of full-length Notch, suggesting the dominant-negative Deltex might prevent the activation of the Notch receptor. We found that Deltex formed a homo-multimer, and mutations in the RING-H2 finger domain abolished this oligomerization. The same mutations in the RING-H2 finger motif of Deltex disrupted the function of Deltex in vivo. However, when the same mutant was fused to a heterologous dimerization domain (Glutathione-S-Transferase), the chimeric protein had normal Deltex activity. Therefore, oligomerization mediated by the RING-H2 finger motif is an integral step in the signaling function of Deltex.
Matsuno
Kenji
K
Department of Biological Science and Technology, Science University of Tokyo, 2641 Yamazaki, Noda, Chiba 278-8510, Japan. matsuno@rs.noda.sut.ac.jp
Ito
Mikiko
M
Hori
Kazuya
K
Miyashita
Fumiyasu
F
Suzuki
Satoshi
S
Kishi
Noriyuki
N
Artavanis-Tsakonas
Spyros
S
Okano
Hideyuki
H
D020816:Amino Acid Motifs; D000818:Animals; D030801:Animals, Genetically Modified; D004330:Drosophila / Q000235:genetics / Q000254:growth & development; D029721:Drosophila Proteins; D019476:Insect Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D008565:Membrane Proteins / Q000378:metabolism*; D016296:Mutagenesis; D011392:Proline; D051880:Receptors, Notch; D015398:Signal Transduction
C086679:DX protein, Drosophila; D029721:Drosophila Proteins; D019476:Insect Proteins; D008565:Membrane Proteins; C497759:N protein, Drosophila; D051880:Receptors, Notch; D011392:Proline