Newly synthesized polycystin‐1 takes different trafficking pathways to the apical and ciliary membranes

Abstract

Mutations in the genes encoding polycystin-1 (PC1) and polycystin 2 (PC2) cause autosomal dominant polycystic kidney disease. These transmembrane proteins colocalize in the primary cilia of renal epithelial cells, where they may participate in sensory processes. PC1 is also found in the apical membrane when expressed in cultured epithelial cells. PC1 undergoes autocatalytic cleavage, producing an extracellular N-terminal fragment that remains noncovalently attached to the transmembrane C-terminus. Exposing cells to alkaline solutions elutes the N-terminal fragment while the C-terminal fragment is retained in the cell membrane. Utilizing this observation, we developed a "strip-recovery" synchronization protocol to study PC1 trafficking in polarized LLC-PK1 renal epithelial cells. Following alkaline strip, a new cohort of PC1 repopulates the cilia within 30 minutes, while apical delivery of PC1 was not detectable until 3 hours. Brefeldin A (BFA) blocked apical PC1 delivery, while ciliary delivery of PC1 was BFA insensitive. Incubating cells at 20°C to block trafficking out of the trans-Golgi network also inhibits apical but not ciliary delivery. These results suggest that newly synthesized PC1 takes distinct pathways to the ciliary and apical membranes. Ciliary PC1 appears to by-pass BFA sensitive Golgi compartments, while apical delivery of PC1 traverses these compartments.

A New Look at the Functional Organization of the Golgi Ribbon

“…In polarized epithelial cells a primary cilium protruding from the apical membrane is anchored at the basal body, a structure consisting of the mother and daughter centrioles of the centrosome that during cell polarization relocated to the apical membrane, losing most of its pericentriolar material and ability to nucleate MTs (Figure 5C). Notably, newly synthesized proteins that are delivered to the ciliary membrane in a Rab11-, Rab8- and exocyst-dependent manner frequently follow pathways that bypass the Golgi stacks (Tian et al, 2014; Bernabé-Rubio and Alonso, 2017; Gilder et al, 2018; Witzgall, 2018). This transport is likely to involve a circular membrane compartment surrounding the base of the cilium that contains Rab11 and the IC/ cis -Golgi protein GM130 (Kim et al, 2010; He et al, 2012; Stoops et al, 2015).…”

Pannexin‐1 mediates fluid shear stress‐sensitive purinergic signaling and cyst growth in polycystic kidney disease

“…The Pkd1 gene encodes for the protein polycystin‐1 (PC1), which locates to the primary cilium and plasma membrane of renal tubular epithelial cells. PC1 forms a complex with polycystin‐2 (PC2, encoded by Pkd2 ), which is a nonselective cation channel . PC1 is suggested to act as a mechanosensor of fluid shear stress (FSS) generated by pro‐urine flow, regulating the physiological responses in renal tubular epithelial cells …”

GRAF2, WDR44, and MICAL1 mediate Rab8/10/11–dependent export of E-cadherin, MMP14, and CFTR ΔF508

“…These cargos have been proposed to bypass the Golgi and are said to follow an unconventional pathway of secretion. Among the cargos that have been reported to reach the plasma membrane in the presence of BFA are E-cadherin (Low et al, 1992), MMP14 (Deryugina et al, 2004), CFTR (Rennolds et al, 2008;Gee et al, 2011), and the ciliary protein Polycystin-1 (Gilder et al, 2018). Whether these cargos actually bypass the Golgi and follow the same route out of the ER is unclear, but what they also have in common is that their export depends on a small group of Rabs.…”