Proteasomal degradation of ubiquitinated Insig proteins is determined by serine residues flanking ubiquitinated lysines.
Insig-1 and Insig-2 are closely related proteins of the endoplasmic reticulum that play crucial roles in cholesterol homeostasis by inhibiting excessive cholesterol synthesis and uptake. In sterol-depleted cells Insig-1 is degraded at least 15 times more rapidly than Insig-2, owing to ubiquitination of Lys-156 and Lys-158 in Insig-1. In this study, we use domain-swapping methods to localize amino acid residues responsible for this differential degradation. In the case of Insig-2, Glu-214 stabilizes the protein by preventing ubiquitination. When Glu-214 is changed to alanine, Insig-2 becomes ubiquitinated, but it is still not degraded as rapidly as ubiquitinated Insig-1. The difference in the degradation rates is traced to two amino acids: Ser-149 in Insig-1 and Ser-106 in Insig-2. Ser-149, which lies NH(2)-terminal to the ubiquitination sites, accelerates the degradation of ubiquitinated Insig-1. Ser-106, which is COOH-terminal to the ubiquitination sites, retards the degradation of ubiquitinated Insig-2. The current studies indicate that the degradation of ubiquitinated Insigs is controlled by serine residues flanking the sites of ubiquitination.
Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
Joon No
JN
Lee
Yi
Y
Gong
Xiangyu
X
Zhang
Jin
J
Ye
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D017124:Conserved Sequence; D006224:Cricetinae; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008239:Lysine / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D011485:Protein Binding; D016415:Sequence Alignment; D012694:Serine / Q000235:genetics / Q000378:metabolism*; D025801:Ubiquitin / Q000378:metabolism*
C108626:INSIG1 protein, human; C467727:INSIG2 protein, human; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D025801:Ubiquitin; D012694:Serine; D046988:Proteasome Endopeptidase Complex; D008239:Lysine