Rab11a-dependent exocytosis of discoidal/fusiform vesicles in bladder umbrella cells

Abstract

The discoidal/fusiform vesicles (DFV) of bladder umbrella cells undergo regulated exocytosis in response to stretch, but little is known about their biogenesis or the molecular machinery that modulates this process. We observed that Rab11a was expressed in umbrella cells (but not Rab11b or Rab25) and was associated with DFV. Using adenovirus-mediated delivery we transduced umbrella cells in situ with either dominant active (DA) or dominant negative (DN) mutants of Rab11a. DA-Rab11a stimulated an increase in apical surface area in the absence of stretch, whereas DN-Rab11a inhibited stretch-induced changes. Endocytosed fluid and membrane markers had little access to Rab11a-positive DFV, but virally expressed human growth hormone (hGH), a secretory protein, was packaged into DFV. Whereas expression of DARab11a stimulated release of hGH into the bladder lumen, expression of DN-Rab11a had the opposite effect. Our results indicate that DFV may be biosynthetic in nature and that their exocytosis depends on the activity of the Rab11a GTPase.uroepithelium ͉ urothelium ͉ GTPases

Distinct Apical and Basolateral Membrane Requirements for Stretch-induced Membrane Traffic at the Apical Surface of Bladder Umbrella Cells

“…In control experiments the tissue was equilibrated for 30 min ( Figure 1A) and then left unperturbed for up to 3 h, during which time the umbrella cells maintained a relatively stable TEV of Ϫ15 to Ϫ20 mV, a transepithelial conductance of ϳ0.05 mS/cm 2 (where conductance is the inverse of transepithelial resistance), a low short-circuit current (I sc ) of ϳ2 A/cm 2 , and a transepithelial membrane capacitance (C T ) of ϳ2.0 F, where 1 F Ϸ 1 cm 2 of surface area (Figure 2A). We previously showed that increases in capacitance are a result of increased apical membrane exocytosis of subapical discoidal/fusiform vesicles (Truschel et al, 2002;Khandelwal et al, 2008). When hydrostatic pressure in the mucosal chamber was raised to 4 cm H 2 O ( Figure 1B), the tissue visibly bowed outward (data not shown), and electrophysiological changes consistent with ion transport were observed including a rapid decrease in TEV and transient increases in conductance and I sc ( Figure 2A).…”

“…In addition, we previously examined the ultrastructure of serially sectioned tissue to show that when uroepithelial tissue is mounted on tissue rings, the apical surface of umbrella cells contains no detectable pockets/adhesions and the subapical pool of discoidal vesicles are not continuous with the plasma membrane (Truschel et al, 2002). Furthermore we have performed numerous biochemical studies that show that stretch-induced increases in surface area are a result of fusion of discoidal/ fusiform-shaped vesicles with the apical plasma membrane of the umbrella cell and not simply unfolding of apical membrane (Truschel et al, 2002;Apodaca, 2004;Khandelwal et al, 2008).…”

At the Intersection of the Pathways for Exocytosis and Autophagy

“…For example, lipids and proteins from the plasma membrane are recovered by the cell for cytosolic recycling via compensatory endocytosis, which also allows for the maintenance of membrane homeostasis at the sites of active exocytosis; directing endocytosed membrane back into the endosomal network or Golgi for degradation or recycling (Sramkova et al, 2009). This type of endocytosis is of particular importance in specialized secretory cells, such as, bladder umbrella cells (Khandelwal et al, 2008;Khandelwal et al, 2010), neurons (Kim & von Gersdorff, 2009;Llobet et al, 2011;Logiudice et al, 2009) and neuroendocrine cells (Engisch & Nowycky, 1998;Barg & Machado, 2008). This allows for the rapid recycling of secretory vesicles back into the reserve pool.…”

A Rab11a-enriched subapical membrane compartment regulates a cytoskeleton-dependent transcytotic pathway in secretory epithelial cells of the lacrimal gland

“…However, Rab11a also appears to be involved in regulated exocrine secretory processes in specialized epithelial cells. In bladder umbrella cells, Rab11a is associated with discoidal/fusiform exocytotic vesicles, and expression of Rab11a DN inhibits the regulated secretion from these vesicles (Khandelwal et al, 2008).…”