title: Exploring the membrane topology of prohormone convertase 1 in AtT20 Cells: in situ analysis by immunofluorescence microscopy.
abstract: Prohormone convertase 1 (PC1) was previously characterized as a partially transmembrane protein in purified chromaffin granules of bovine adrenal medulla1. This was challenged with experiments on transfected PC1 in COS1 cells, a non-endocrine cell line2. To address this issue, we undertook to analyze its extraction properties  in vitro and its immunocytochemical localization  in situ in AtT20 cells, an endocrine cell line that expresses PC1. Most of the 87 kDa form of PC1 was resistant to carbonate extraction suggesting that it had properties of a transmembrane protein. Under semi-permeabilized conditions whereby only the plasma membrane was permeabilized, the carboxy-terminus of PC1 was specifically immunostained whereas the amino-terminus was not. These results indicate that the amino-terminus of PC1 was within the lumen of the Golgi and granules, and some of the C-terminus was exposed to the cytosol. Thus, endogenous PC1 can assume a transmembrane orientation  in situ in AtT20 cells.
authors.affiliation: Section on Cellular Neurobiology, Program in Developmental Neuroscience, Eunice Shriver Kennedy National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, 20892, USA ; National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, 20892, USA.
authors.forename: Niamh X
authors.initials: NX
authors.lastname: Cawley
authors.forename: Meera
authors.initials: M
authors.lastname: Sridhar
authors.forename: Hong
authors.initials: H
authors.lastname: Hong
authors.forename: Peng
authors.initials: P
authors.lastname: Loh