LNX functions as a RING type E3 ubiquitin ligase that targets the cell fate determinant Numb for ubiquitin-dependent degradation.
LNX is a RING finger and PDZ domain containing protein that interacts with the cell fate determinant Numb. To investigate the function of LNX, we tested its RING finger domain for ubiquitin ligase activity. The isolated RING finger domain was able to function as an E2-dependent, E3 ubiquitin ligase in vitro and mutation of a conserved cysteine residue within the RING domain abolished its activity, indicating that LNX is the first described PDZ domain-containing member of the E3 ubiquitin ligase family. We have identified Numb as a substrate of LNX E3 activity in vitro and in vivo. In addition to the RING finger, a region of LNX, including the Numb PTB domain-binding site and the first PDZ domain, is required for Numb ubiquitylation. Expression of wild-type but not mutant LNX causes proteasome-dependent degradation of Numb and can enhance Notch signalling. These results suggest that the levels of mammalian Numb protein and therefore, by extension, the processes of asymmetric cell division and cell fate determination may be regulated by ubiquitin-dependent proteolysis.
Nie
Jing
J
The Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children and Department of Medical Biophysics, University of Toronto, 555 University Avenue, Toronto, Ontario M5G 1X8, Canada.
McGill
Melanie A
MA
Dermer
Matt
M
Dho
Sascha E
SE
Wolting
Cheryl D
CD
McGlade
C Jane
CJ
D000818:Animals; D001665:Binding Sites; D016466:CHO Cells; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D006224:Cricetinae; D004285:Dogs; D006801:Humans; D008025:Ligases / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D025801:Ubiquitin / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases
D002352:Carrier Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; C106293:Numb protein, mouse; D011993:Recombinant Fusion Proteins; D025801:Ubiquitin; C484733:LNX1 protein, human; C111865:Lnx1 protein, mouse; D044767:Ubiquitin-Protein Ligases; D008025:Ligases