Sterol-regulated release of SREBP-2 from cell membranes requires two sequential cleavages, one within a transmembrane segment.
Sterol regulatory element binding proteins (SREBPs) are transcription factors attached to the endoplasmic reticulum. The NH2-segment, which activates transcription, is connected to membranes by a hairpin anchor formed by two transmembrane sequences and a short lumenal loop. Using H-Ras-SREBP-2 fusion proteins, we show that the NH2-segment is released from membranes by two sequential cleavages. The first, regulated by sterols, occurs in the lumenal loop. The second, not regulated by sterols, occurs within the first transmembrane domain. The liberated NH2-segment enters the nucleus and activates genes controlling cholesterol synthesis and uptake. Certain mutant Chinese hamster ovary cells are auxotrophic for cholesterol because they fail to carry out the second cleavage; the NH2-segment remains membrane-bound and transcription is not activated.
Sakai
J
J
Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas, Texas 75235, USA.
Duncan
E A
EA
Rawson
R B
RB
Hua
X
X
Brown
M S
MS
Goldstein
J L
JL
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D002460:Cell Line / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D002784:Cholesterol / Q000096:biosynthesis / Q000235:genetics; D006224:Cricetinae; D018076:DNA, Complementary / Q000235:genetics; D004268:DNA-Binding Proteins / Q000187:drug effects* / Q000378:metabolism; D010450:Endopeptidases / Q000378:metabolism; D000939:Epitopes / Q000235:genetics; D006801:Humans; D015151:Immunoblotting; D007668:Kidney / Q000166:cytology; D016350:Leucine Zippers / Q000187:drug effects; D008565:Membrane Proteins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000502:physiology; D017693:Sodium Bicarbonate; D051782:Sterol Regulatory Element Binding Protein 2; D013261:Sterols / Q000494:pharmacology*; D014157:Transcription Factors / Q000187:drug effects* / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics; D014162:Transfection; D018631:ras Proteins / Q000235:genetics
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D000939:Epitopes; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; C497390:SREBF2 protein, human; D051782:Sterol Regulatory Element Binding Protein 2; D013261:Sterols; D014157:Transcription Factors; D017693:Sodium Bicarbonate; D002784:Cholesterol; D010450:Endopeptidases; D018631:ras Proteins