Human HRD1 is an E3 ubiquitin ligase involved in degradation of proteins from the endoplasmic reticulum.
The ubiquitin system plays an important role in endoplasmic reticulum (ER)-associated degradation of proteins that are misfolded, that fail to associate with their oligomerization partners, or whose levels are metabolically regulated. E3 ubiquitin ligases are key enzymes in the ubiquitination process as they recognize the substrate and facilitate coupling of multiple ubiquitin units to the protein that is to be degraded. The Saccharomyces cerevisiae ER-resident E3 ligase Hrd1p/Der3p functions in the metabolically regulated degradation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase and additionally facilitates the degradation of a number of misfolded proteins from the ER. In this study we characterized the structure and function of the putative human orthologue of yeast Hrd1p/Der3p, designated human HRD1. We show that human HRD1 is a non-glycosylated, stable ER protein with a cytosolic RING-H2 finger domain. In the presence of the ubiquitin-conjugating enzyme UBC7, the RING-H2 finger has in vitro ubiquitination activity for Lys(48)-specific polyubiquitin linkage, suggesting that human HRD1 is an E3 ubiquitin ligase involved in protein degradation. Human HRD1 appears to be involved in the basal degradation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase but not in the degradation that is regulated by sterols. Additionally we show that human HRD1 is involved in the elimination of two model ER-associated degradation substrates, TCR-alpha and CD3-delta.
Department of Medical Microbiology, Leiden University Medical Center, P. O. Box 9600, 2300 RC Leiden, The Netherlands.
Marjolein
M
Kikkert
Ram
R
Doolman
Min
M
Dai
Rachel
R
Avner
Gerco
G
Hassink
Sjaak
S
van Voorden
Swapna
S
Thanedar
Joseph
J
Roitelman
Vincent
V
Chau
Emmanuel
E
Wiertz
D000595:Amino Acid Sequence; D000818:Animals; D017252:CD3 Complex / Q000096:biosynthesis; D019286:Endopeptidase K / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000201:enzymology*; D019673:Genes, T-Cell Receptor alpha / Q000235:genetics; D006031:Glycosylation; D006367:HeLa Cells; D006801:Humans; D006903:Hydroxymethylglutaryl CoA Reductases / Q000378:metabolism; D008239:Lysine / Q000737:chemistry; D051379:Mice; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D041681:NIH 3T3 Cells; D014176:Protein Biosynthesis; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013261:Sterols / Q000378:metabolism; D013329:Structure-Activity Relationship; D013997:Time Factors; D014162:Transfection; D025801:Ubiquitin / Q000378:metabolism; D044763:Ubiquitin-Conjugating Enzymes / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry* / Q000502:physiology*
D017252:CD3 Complex; D013261:Sterols; D025801:Ubiquitin; D006903:Hydroxymethylglutaryl CoA Reductases; C471617:UBE2G2 protein, human; D044763:Ubiquitin-Conjugating Enzymes; C482345:SYVN1 protein, human; D044767:Ubiquitin-Protein Ligases; D019286:Endopeptidase K; D008239:Lysine