title: Furin processing dictates ectodomain shedding of human FAT1 cadherin.
abstract: Fat1 is a single pass transmembrane protein and the largest member of the cadherin superfamily. Mouse knockout models and in vitro studies have suggested that Fat1 influences cell polarity and motility. Fat1 is also an upstream regulator of the Hippo pathway, at least in lower vertebrates, and hence may play a role in growth control. In previous work we have established that FAT1 cadherin is initially cleaved by proprotein convertases to form a noncovalently linked heterodimer prior to expression on the cell surface. Such processing was not a requirement for cell surface expression, since melanoma cells expressed both unprocessed FAT1 and the heterodimer on the cell surface. Here we further establish that the site 1 (S1) cleavage step to promote FAT1 heterodimerisation is catalysed by furin and we identify the cleavage site utilised. For a number of other transmembrane receptors that undergo heterodimerisation the S1 processing step is thought to occur constitutively but the functional significance of heterodimerisation has been controversial. It has also been generally unclear as to the significance of receptor heterodimerisation with respect to subsequent post-translational proteolysis that often occurs in transmembrane proteins. Exploiting the partial deficiency of FAT1 processing in melanoma cells together with furin-deficient LoVo cells, we manipulated furin expression to demonstrate that only the heterodimer form of FAT1 is subject to cleavage and subsequent release of the extracellular domain. This work establishes S1-processing as a clear functional prerequisite for ectodomain shedding of FAT1 with general implications for the shedding of other transmembrane receptors.
authors.affiliation: School of Biomedical Sciences and Pharmacy, University of Newcastle, Callaghan, NSW 2308, Australia; Hunter Medical Research Institute, New Lambton Heights, NSW 2305, Australia.
authors.forename: Elham
authors.initials: E
authors.lastname: Sadeqzadeh
authors.affiliation: School of Biomedical Sciences and Pharmacy, University of Newcastle, Callaghan, NSW 2308, Australia; Hunter Medical Research Institute, New Lambton Heights, NSW 2305, Australia.
authors.forename: Charles E
authors.initials: CE
authors.lastname: de Bock
authors.affiliation: Department of Internal Medicine, Knappschaftskrankenhaus, Ruhr-University Bochum, Bochum, Germany.
authors.forename: Natalie
authors.initials: N
authors.lastname: Wojtalewicz
authors.affiliation: School of Biomedical Sciences and Pharmacy, University of Newcastle, Callaghan, NSW 2308, Australia.
authors.forename: Janet E
authors.initials: JE
authors.lastname: Holt
authors.affiliation: ABRF, Research Services, University of Newcastle, Callaghan, NSW 2308, Australia.
authors.forename: Nathan D
authors.initials: ND
authors.lastname: Smith
authors.affiliation: School of Biomedical Sciences and Pharmacy, University of Newcastle, Callaghan, NSW 2308, Australia; Hunter Medical Research Institute, New Lambton Heights, NSW 2305, Australia; Hunter Translational Cancer Research Unit, School of Biomedical Sciences and Pharmacy, University of Newcastle, Callaghan, NSW 2308, Australia.
authors.forename: Matthew D
authors.initials: MD
authors.lastname: Dun
authors.affiliation: Department of Internal Medicine, Knappschaftskrankenhaus, Ruhr-University Bochum, Bochum, Germany.
authors.forename: Irmgard
authors.initials: I
authors.lastname: Schwarte-Waldhoff
authors.affiliation: Hunter Medical Research Institute, New Lambton Heights, NSW 2305, Australia; Hunter Translational Cancer Research Unit, School of Biomedical Sciences and Pharmacy, University of Newcastle, Callaghan, NSW 2308, Australia; School of Environmental & Life Sciences, University of Newcastle, Callaghan, New South Wales, Australia. Electronic address: Rick.Thorne@newcastle.edu.au.
authors.forename: Rick F
authors.initials: RF
authors.lastname: Thorne
mesh_terms: D015820:Cadherins / Q000096:biosynthesis / Q000378:metabolism*; D045744:Cell Line, Tumor; D045683:Furin / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D015603:Keratinocytes; D008545:Melanoma / Q000378:metabolism; D043484:Proprotein Convertases / Q000235:genetics; D055503:Protein Multimerization; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D059748:Proteolysis; D034622:RNA Interference; D034741:RNA, Small Interfering; D012697:Serine Endopeptidases / Q000235:genetics; D013381:Subtilisins / Q000235:genetics
chemical_list: D015820:Cadherins; C502403:FAT1 protein, human; D034741:RNA, Small Interfering; C080654:PCSK6 protein, human; C099101:PCSK7 protein, human; D043484:Proprotein Convertases; D012697:Serine Endopeptidases; D013381:Subtilisins; C115575:membrane-bound transcription factor peptidase, site 1; D045683:Furin
keywords: Ectodomain shedding; FAT1 cadherin; Furin; Heterodimerisation; Proprotein convertase; S1 cleavage