Notch subunit heterodimerization and prevention of ligand-independent proteolytic activation depend, respectively, on a novel domain and the LNR repeats.
Notch proteins are transmembrane receptors that participate in a highly conserved signaling pathway that regulates morphogenesis in metazoans. Newly synthesized Notch receptors are proteolytically cleaved during transit to the cell surface, creating heterodimeric mature receptors comprising noncovalently associated extracellular (N(EC)) and transmembrane (N) subunits. Ligand binding activates Notch by inducing two successive proteolytic cleavages, catalyzed by metalloproteases and gamma-secretase, respectively, that permit the intracellular portion of N to translocate to the nucleus and activate transcription of target genes. Prior work has shown that the presence of N(EC) prevents ligand-independent activation of N, but the mechanisms involved are poorly understood. Here, we define the roles of two regions at the C-terminal end of N(EC) that participate in maintaining the integrity of resting Notch receptors through distinct mechanisms. The first region, a hydrophobic, previously uncharacterized portion of N(EC), is sufficient to form stable complexes with the extracellular portion of N. The second region, consisting of the three Lin12/Notch repeats, is not needed for heterodimerization but acts to protect N from ligand-independent cleavage by metalloproteases. Together, these two contiguous regions of N(EC) impose crucial restraints that prevent premature Notch receptor activation.
Sanchez-Irizarry
Cheryll
C
Department of Pathology, Brigham and Women's Hospital and Harvard Medical School, 77 Ave. Louis Pasteur, Boston, MA 02115, USA.
Carpenter
Andrea C
AC
Weng
Andrew P
AP
Pear
Warren S
WS
Aster
Jon C
JC
Blacklow
Stephen C
SC
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D019281:Dimerization; D005786:Gene Expression Regulation; D006801:Humans; D008024:Ligands; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D051880:Receptors, Notch; D020449:Repetitive Sequences, Amino Acid; D016415:Sequence Alignment; D015398:Signal Transduction
D008024:Ligands; D008565:Membrane Proteins; D021122:Protein Subunits; D051880:Receptors, Notch