title: Rumi functions as both a protein O-glucosyltransferase and a protein O-xylosyltransferase.
abstract: Mutations in rumi result in a temperature-sensitive loss of Notch signaling in Drosophila. Drosophila Rumi is a soluble, endoplasmic reticulum-retained protein with a CAP10 domain that functions as a protein O-glucosyltransferase. In human and mouse genomes, three potential Rumi homologues exist: one with a high degree of identity to Drosophila Rumi (52%), and two others with lower degrees of identity but including a CAP10 domain (KDELC1 and KDELC2). Here we show that both mouse and human Rumi, but not KDELC1 or KDELC2, catalyze transfer of glucose from UDP-glucose to an EGF repeat from human factor VII. Similarly, human Rumi, but not KDELC1 or KDELC2, rescues the Notch phenotypes in Drosophila rumi clones. During characterization of the Rumi enzymes, we noted that, in addition to protein O-glucosyltransferase activity, both mammalian and Drosophila Rumi also showed significant protein O-xylosyltransferase activity. Rumi transfers Xyl or glucose to serine 52 in the O-glucose consensus sequence ( ) of factor VII EGF repeat. Surprisingly, the second serine (S53) facilitates transfer of Xyl, but not glucose, to the EGF repeat by Rumi. EGF16 of mouse Notch2, which has a diserine motif in the consensus sequence ( ), is also modified with either O-Xyl or O-glucose glycans in cells. Mutation of the second serine (S590A) causes a loss of O-Xyl but not O-glucose at this site. Altogether, our data establish dual substrate specificity for the glycosyltransferase Rumi and provide evidence that amino acid sequences of the recipient EGF repeat significantly influence which donor substrate (UDP-glucose or UDP-Xyl) is used.
authors.affiliation: Department of Biochemistry and Cell Biology, Institute of Cell and Developmental Biology, Stony Brook University, Stony Brook, NY 11794, USA.
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Takeuchi
authors.forename: Rodrigo C
authors.initials: RC
authors.lastname: Fernández-Valdivia
authors.forename: Devin S
authors.initials: DS
authors.lastname: Caswell
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Nita-Lazar
authors.forename: Nadia A
authors.initials: NA
authors.lastname: Rana
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Garner
authors.forename: Thomas K
authors.initials: TK
authors.lastname: Weldeghiorghis
authors.forename: Megan A
authors.initials: MA
authors.lastname: Macnaughtan
authors.forename: Hamed
authors.initials: H
authors.lastname: Jafar-Nejad
authors.forename: Robert S
authors.initials: RS
authors.lastname: Haltiwanger
mesh_terms: D000818:Animals; D004330:Drosophila / Q000235:genetics / Q000502:physiology*; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism*; D005167:Factor VII / Q000235:genetics / Q000378:metabolism*; D005964:Glucosyltransferases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D013058:Mass Spectrometry; D051379:Mice; D009154:Mutation / Q000235:genetics; D010430:Pentosyltransferases / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology*; D013379:Substrate Specificity; D000097608:UDP Xylose-Protein Xylosyltransferase
chemical_list: D029721:Drosophila Proteins; D005167:Factor VII; D005964:Glucosyltransferases; D010430:Pentosyltransferases; D000097608:UDP Xylose-Protein Xylosyltransferase; C527374:Rumi protein, Drosophila