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Planar cell polarity (PCP) signaling controls polarized behaviors in diverse tissues, including the collective cell movements of gastrulation and the planar polarized beating of motile cilia. A major question in PCP signaling concerns the mechanisms linking this signaling cascade with more general cytoskeletal elements to drive polarized behavior. Previously, we reported that the PCP effector protein Wdpcp (formerly known as Fritz) interacts with septins and is critical for collective cell migration and cilia formation. Here, we report that Wdpcp is broadly involved in maintaining cortical tension in epithelial cells. In vivo 3D time-lapse imaging revealed that Wdpcp is necessary for basolateral plasma membrane stability in epithelial tissues, and we further show that Wdpcp controls cortical septin localization to maintain cortical rigidity in mucociliary epithelial cells. Finally, we show that Wdpcp acts via actomyosin to maintain balanced cortical tension in the epithelium. These data suggest that, in addition to its role in controlling plasma membrane dynamics in collective mesenchymal cell movements, Wdpcp is also essential for normal cell cortex stability during epithelial homeostasis.
Fig. 1. Wdpcp controls plasma membrane stability in epithelial cells. (A) Control
tissue expressing membrane-RFP, cellâcell contact indicated by the blue box is
shown in (a0 ). (B) Morphant tissue showing severe membrane blebbing. The boxed
region is shown in (b0) with higher magnification. Asterisks indicate blebs. (C and c0)
Co-injection of Wdpcp-MO and Wdpcp mRNA (300 pg) rescued the membrane
blebbing. (D) The quantification of membrane blebbing. Blebbing index was
determined as the ratio of total blebbing area within a single cell-contact compared
to the length of that cellâcell contact. Control n = 125; Wdpcp-MO n = 196; Rescue
n = 131. The scale bars are 10 lm. (E) Still frames from a time-laps movie of control
epithelium (Supplemental time laps Movie 1). (F) Still frames from a time-laps
movie of Wdpcp morphant epithelium (Supplemental time laps Movie 2).
Fig. 2. Wdpcp knockdown caused severe membrane blebbing exclusively at the
basolateral membrane. (A) The apical plasma membranes in control Xenopus
epidermal epithelium, showing tight cellâcell contacts. (a0) The basolateral membranes
of cells shown in panel (A) are also remain stably connected (B). The apical
membrane of Wdpcp morphant tissue was indistinguishable from the control apical
membrane. (b0) Wdpcp morphants displayed severe blebbing of the basolateral
membranes. (C) XâZ projection of control epithelium. Green arrowheads indicate
apical junctions. Blue arrowheads indicate basolateral membrane. (D) XâZ projection
of Wdpcp morphant epithelium. Blue arrowheads indicate blebbing basolateral
membrane while the apical junction is quite stable (green arrowheads). The scale
bars are 10 lm. (c0 and d0) The drawing indicating the optical sectioning points for
confocal imaging of (A and B) (Green lines) or (a0 and b0) (Blue lines).
Fig. 3. Wdpcp is necessary for the cortical localization of septins to provide cortical
rigidity. (A and a0) GFP-Septin2 localized to the cell membrane marked by memRFP.
(B and b0) Knockdown of Wdpcp delocalized GFP-septin2 from the cell cortex. (C
and D) Knockdown of Wdpcp expression did not disrupt cortical actin accumulation.
Actin filaments were stained by phalloidin-Alexa488. The scale bars are 10 lm.
Fig. 4. Wdpcp maintains membrane stability via Rho Kinase and Myosin activity.
(A) Epidermal tissue injected with Wdpcp-MO displaying severe membrane
blebbing. (B) Y27632, a Rho kinase inhibitor, blocked the blebbing phenotype in
Wdpcp morphants. (C) The same epidermis shown in B started blebbing again after
1 hour washout of Y27632. (D) Quantification of blebbing index; Wdpcp-MO n = 24;
Y27632 n = 44; Wash n = 23; Blebbistatin n = 83. The scale bars are 10 lm.
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