SignificanceEndocytosis plays a fundamental role in Wnt signaling, leading to the sequestration of cytosolic GSK3 kinase inside multivesicular endosomes. Here we show an unexpected role for the enzyme protein arginine methyltransferase 1 (PRMT1) during Wnt signaling. In the case of the soluble tumor suppressor transcription factor Smad4, modification of a particular arginine was required before this protein could be phosphorylated by GSK3. Wnt3a addition caused rapid endocytosis that strikingly expanded the cellular liquid-phase compartment within 5–20 min. Wnt-induced vesicles contained arginine-methylated proteins, GSK3 and PRMT1 itself, within membrane-bound organelles. The translocation of these proteins into multivesicular endosomes via microautophagy was essential for canonical Wnt signaling.

Canonical Wnt signaling is emerging as a major regulator of endocytosis. Wnt treatment markedly increased the endocytosis and degradation in lysosomes of BSA. In this study, we report that in addition to receptor-mediated endocytosis, Wnt also triggers the intake of large amounts of extracellular fluid by macropinocytosis, a nonreceptor-mediated actin-driven process. Macropinocytosis induction is rapid and independent of protein synthesis. In the presence of Wnt, large amounts of nutrient-rich packages such as proteins and glycoproteins were channeled into lysosomes after fusing with smaller receptor-mediated vesicles containing glycogen synthase kinase 3 (GSK3) and protein arginine ethyltransferase 1 (PRMT1), an enzyme required for canonical Wnt signaling. Addition of Wnt3a, as well as overexpression of Disheveled (Dvl), Frizzled (Fz8), or dominant-negative Axin induced endocytosis. Depletion of the tumor suppressors adenomatous polyposis coli (APC) or Axin dramatically increased macropinocytosis, defined by incorporation of the high molecular weight marker tetramethylrhodamine (TMR)-dextran and its blockage by the Na+/H+ exchanger ethylisopropyl amiloride (EIPA). Macropinocytosis was blocked by dominant-negative vacuolar protein sorting 4 (Vps4), indicating that the Wnt pathway is dependent on multivesicular body formation, a process called microautophagy. SW480 colorectal cancer cells displayed constitutive macropinocytosis and increased extracellular protein degradation in lysosomes, which were suppressed by restoring full-length APC. Accumulation of the transcriptional activator β-catenin in the nucleus of SW480 cells was inhibited by methyltransferase inhibition, EIPA, or the diuretic amiloride. The results indicate that Wnt signaling switches metabolism toward nutrient acquisition by engulfment of extracellular fluids and suggest possible treatments for Wnt-driven cancer progression.

Phosphorylation and methylation of desmoplakin are required for proper junction assembly and adhesion strengthening, and inhibition of these modifications might contribute to skin and heart diseases.

Highlights d GSK3 and Axin1 normally repress macropinocytosis in basal cellular conditions d GSK3 inhibition or Axin1 mutation triggers macropinocytosis and metabolite changes d Macropinocytosis induces acidification and catabolic activity of lysosomes d Lysosomal activation by Wnt signaling is independent of new protein synthesis