Rumi functions as both a protein O-glucosyltransferase and a protein O-xylosyltransferase
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 UDPglucose 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 ( 50CASSPC 55) 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 ( 587CYSSPC 592), 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.
Publication Source (Journal or Book title)
Proceedings of the National Academy of Sciences of the United States of America
Takeuchi, H., Fernańdez-Valdivia, R., Caswell, D., Nita-Lazar, A., Rana, N., Garner, T., Weldeghiorghis, T., Macnaughtan, M., Jafar-Nejad, H., & Haltiwanger, R. (2011). Rumi functions as both a protein O-glucosyltransferase and a protein O-xylosyltransferase. Proceedings of the National Academy of Sciences of the United States of America, 108 (40), 16600-16605. https://doi.org/10.1073/pnas.1109696108