WLS-dependent secretion of WNT3A requires Ser209 acylation and vacuolar acidification.
Wnt proteins are secreted post-translationally modified proteins that signal locally to regulate development and proliferation. The production of bioactive Wnts requires a number of dedicated factors in the secreting cell whose coordinated functions are not fully understood. A screen for small molecules identified inhibitors of vacuolar acidification as potent inhibitors of Wnt secretion. Inhibition of the V-ATPase or disruption of vacuolar pH gradients by diverse drugs potently inhibited Wnt/β-catenin signaling both in cultured human cells and in vivo, and impaired Wnt-regulated convergent extension movements in Xenopus embryos. WNT secretion requires its binding to the carrier protein wntless (WLS); we find that WLS is ER-resident in human cells and WNT3A binding to WLS requires PORCN-dependent lipid modification of WNT3A at serine 209. Inhibition of vacuolar acidification results in accumulation of the WNT3A-WLS complex both in cells and at the plasma membrane. Modeling predictions suggest that WLS has a lipid-binding β-barrel that is similar to the lipocalin-family fold. We propose that WLS binds Wnts in part through a lipid-binding domain, and that vacuolar acidification is required to release palmitoylated WNT3A from WLS in secretory vesicles, possibly to facilitate transfer of WNT3A to a soluble carrier protein.
Coombs
Gary S
GS
Program in Cancer and Stem Cell Biology, Duke-NUS Graduate Medical School, 8 College Road, 169857, Singapore.
Yu
Jia
J
Canning
Claire A
CA
Veltri
Charles A
CA
Covey
Tracy M
TM
Cheong
Jit K
JK
Utomo
Velani
V
Banerjee
Nikhil
N
Zhang
Zong Hong
ZH
Jadulco
Raquel C
RC
Concepcion
Gisela P
GP
Bugni
Tim S
TS
Harper
Mary Kay
MK
Mihalek
Ivana
I
Jones
C Michael
CM
Ireland
Chris M
CM
Virshup
David M
DM
D000215:Acylation; D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors*; D000818:Animals; D004625:Embryo, Nonmammalian; D047108:Embryonic Development / Q000187:drug effects; D057809:HEK293 Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D018942:Macrolides / Q000302:isolation & purification / Q000494:pharmacology*; D011485:Protein Binding; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D054852:Small Molecule Libraries / Q000302:isolation & purification; D014617:Vacuoles / Q000737:chemistry / Q000378:metabolism*; D051153:Wnt Proteins / Q000378:metabolism*; D060508:Wnt3 Protein; D060509:Wnt3A Protein; D014981:Xenopus; D029867:Xenopus Proteins
D047908:Intracellular Signaling Peptides and Proteins; D018942:Macrolides; D043562:Receptors, G-Protein-Coupled; D054852:Small Molecule Libraries; C549111:WLS protein, human; C555506:WNT3A protein, Xenopus; C555510:WNT3A protein, human; D051153:Wnt Proteins; D060508:Wnt3 Protein; D060509:Wnt3A Protein; D029867:Xenopus Proteins; C440684:chondropsin A; D012694:Serine; D000251:Adenosine Triphosphatases
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