Stimulation with carbachol alters endomembrane distribution and plasma membrane expression of intracellular proteins in lacrimal acinar cells

The events that lead to Sjogren's autoimmune processes in the lacrimal glan d remain poorly understood. The acinar cell's responses to acute cholinergi c stimulation include release of secretory products across the apical plasm a membrane (apm) and a number of processes related to traffic between endom embrane compartments and the basal-lateral plasma membranes (blm), such as recruitment of Na,K-ATPase, accelerated recycling, and accelerated transcyt osis of secretory IgA. We tested the hypothesis that stimulation-induced ac celeration of endomembrane traffic is accompanied by changes in compartment ation and increased blm expression of proteins that are normally sequestere d in endomembrane compartments. Isolated rabbit lacrimal gland acinar cells were cultured in serum-free media for 2 days. After harvesting, cells were incubated with or without 10 mu M carbachol at 37 degrees C for 20 min. Ce lls were lysed, and lysates were analysed by isopycnic centrifugation on so rbitol gradients. Galactosyltransferase catalytic activity was determined b iochemically. Different forms of cathepsin B were detected by Western blott ing. Carbachol stimulation decreased the contents of beta-hexosaminidase, a lpha-glucosidase, and protein in secretory vesicles and increased them in s pecific compartments of the trans-Golgi network (ld-tgns). Stimulation also caused levels of galactosyltransferase, preprocathepsin B, and procathepsi n B to increase two- to three-fold in the blm as well as increasing in the ld-tgns. Other changes caused by sustained stimulation included: (a) increa sed levels of protein and procathepsin B in compartments of the lysosomal p athway; (b) changes in the distributions of Rab5 within the endomembrane sy stem; (c) changes in the distribution of Rab6 within the Golgi complex and tgn; (d) decreased expression of acid phosphatase and MHC class II molecule s in the blm: and (e) decreased total content of Na,K-ATPase, which appeare d to have been selectively depleted from the tgn and blmre. We propose that the normal compartmentation of certain proteins may allow them to remain c ryptic, such that they are not subject to central tolerance. Stimulation-in duced increases in the levers expressed at the blm or secreted to the inter stitium may, therefore, contribute to initiation of local autoimmune respon ses. (C) 1999 Academic Press.