Identification of ubiquitination sites on the X-linked inhibitor of apoptosis protein.
The execution phase of apoptosis is under the control of members of the inhibitor of apoptosis (IAP) family of zinc finger proteins. Several of these proteins contain a C-terminal RING (really interesting new gene) domain that has been postulated to regulate ubiquitination of themselves or their target proteins, thereby modulating thresholds for apoptosis. We demonstrate that the auto-ubiquitination sites of the X-linked IAP (XIAP) are Lys(322) and Lys(328), located in the third baculovirus IAP repeat domain of the protein. Modification of these sites to arginine dramatically reduces ubiquitination of XIAP, but has no measurable effect on the ability of ectopically expressed IAP to rescue cells from two independent apoptotic inducers. Our data firmly locate the auto-ubiquitination sites, and raise doubts regarding the importance of this event as a mechanism for regulating the levels of XIAP.
The Program in Apoptosis and Cell Death Research, Burnham Institute, 10901 North Torrey Pines Road, La Jolla, CA 92037, USA.
Hwain
H
Shin
Kazuya
K
Okada
John C
JC
Wilkinson
Katherine M
KM
Solomon
Colin S
CS
Duckett
John C
JC
Reed
Guy S
GS
Salvesen
D000595:Amino Acid Sequence; D017209:Apoptosis; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D006801:Humans; D008239:Lysine / Q000378:metabolism; D013058:Mass Spectrometry / Q000379:methods; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D025801:Ubiquitin / Q000378:metabolism*; D051636:X-Linked Inhibitor of Apoptosis Protein
D017931:DNA Primers; D011506:Proteins; D011994:Recombinant Proteins; D025801:Ubiquitin; D051636:X-Linked Inhibitor of Apoptosis Protein; C496866:XIAP protein, human; D008239:Lysine