Topology of the C-terminal fragment of human presenilin 1.
Mutations of human presenilin 1 (PS1) have been genetically linked to early-onset familial Alzheimer's disease. PS1 contains 10 hydrophobic regions (HRs) sufficiently long to be alpha-helical membrane spanning segments. Most previous topology studies agree that the N-terminus of PS1 is cytosolic and HRs 1-6 span the membrane but HR 7 does not. However, whether HRs 8 and 9 are membrane spanning segments remains controversial. Here we study the topology and biogenesis of this region of PS1 using a reporter gene fusion approach, where portions of the PS1 sequence containing possible membrane spanning segments were fused up- or downstream of a reporter sequence whose translocation into the endoplasmic reticulum could be monitored via its glycosylation. We provide strong evidence, supported by cysteine accessibility studies in full-length PS1, that HRs 8 and 9 are indeed membrane spanning and that the integration of HR 8 into the membrane is dependent on the presence of HR 9. We also explain how our results reconcile previous apparently divergent conclusions regarding the topology of HRs 8 and 9.
Oh
Young S
YS
Membrane Biology Section, Gene Therapy and Therapeutics Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, DHHS, Bethesda, Maryland 20892-1190, USA. yoh@mail.nih.gov
Turner
R James
RJ
D000595:Amino Acid Sequence; D019921:Biotinylation; D002462:Cell Membrane / Q000737:chemistry*; D003545:Cysteine / Q000737:chemistry; D003600:Cytosol / Q000737:chemistry; D050939:Gene Fusion / Q000379:methods; D017930:Genes, Reporter; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D053764:Presenilin-1; D011993:Recombinant Fusion Proteins / Q000737:chemistry
D008565:Membrane Proteins; C508853:PSEN1 protein, human; D053764:Presenilin-1; D011993:Recombinant Fusion Proteins; D003545:Cysteine