Mitogen-Activated Protein Kinase (MAPK) Pathway Regulates Branching by Remodeling Epithelial Cell Adhesion

Abstract

Although the growth factor (GF) signaling guiding renal branching is well characterized, the intracellular cascades mediating GF functions are poorly understood. We studied mitogen-activated protein kinase (MAPK) pathway specifically in the branching epithelia of developing kidney by genetically abrogating the pathway activity in mice lacking simultaneously dual-specificity protein kinases Mek1 and Mek2. Our data show that MAPK pathway is heterogeneously activated in the subset of G1- and S-phase epithelial cells, and its tissue-specific deletion results in severe renal hypodysplasia. Consequently to the deletion of Mek1/2, the activation of ERK1/2 in the epithelium is lost and normal branching pattern in mutant kidneys is substituted with elongation-only phenotype, in which the epithelium is largely unable to form novel branches and complex three-dimensional patterns, but able to grow without primary defects in mitosis. Cellular characterization of double mutant epithelium showed increased E-cadherin at the cell surfaces with its particular accumulation at baso-lateral locations. This indicates changes in cellular adhesion, which were revealed by electron microscopic analysis demonstrating intercellular gaps and increased extracellular space in double mutant epithelium. When challenged to form monolayer cultures, the mutant epithelial cells were impaired in spreading and displayed strong focal adhesions in addition to spiky E-cadherin. Inhibition of MAPK activity reduced paxillin phosphorylation on serine 83 while remnants of phospho-paxillin, together with another focal adhesion (FA) protein vinculin, were augmented at cell surface contacts. We show that MAPK activity is required for branching morphogenesis, and propose that it promotes cell cycle progression and higher cellular motility through remodeling of cellular adhesions.

Development of the Mammalian Kidney

“…Of these PI3 kinase action is essential for Etv4 and Etv5 expression and branching growth (Tang et al, 2002; Lu et al, 2009). Genetic removal of MAP kinase pathway components blocks branching, but the ureteric epithelium continues to elongate, highlighting a direct role for MAP-kinase signaling in the branching process (Ihermann-Hella et al, 2014). Analysis of Ctnnd1 (β-catenin) mutants suggests canonical Wnt signaling maintains the epithelial progenitor pool, as loss of Ctnnd1 results in premature differentiation of progenitor cells (Marose et al, 2008; Bridgewater et al, 2008).…”

“…Genetic modification of actin depolymerizing factors indicates that actin remodeling is critical for the branching process (Kuure et al, 2010). Further, enhanced epithelial adhesion following removal of MAP kinase signaling suggests adhesive remodeling is necessary for normal branching (Ihermann-Hella et al, 2014). Altering adhesive properties may also impinge on proliferative programs.…”

“…The observation that different Fgf signals induce elongation versus branching growth mediated through the MAP kinase pathway is especially intriguing in this regard (Qiao et al, 2001; Ihermann-Hella et al, 2014). Arcading is a prevalent mode of hooking up nephrons in the human kidney following the period of branching growth.…”

Generating tissue topology through remodeling of cell-cell adhesions

“…Kidney branching morphogenesis proceeds via outgrowth and bifurcation of the ureteric bud. Epithelial cells of the kidney tubules adhere to each other via E-cadherin; in the tip, E-cadherin is enriched apically, while in the stalk or medullary regions it is evenly distributed along the entire lateral membrane [41]. Bifurcation of the ureteric bud requires signaling through mitogen-activated protein kinase (MAPK); when MAPK activity is perturbed, E-cadherin is enriched in tip cells and localizes more basally than in the native kidney [41].…”

“…Epithelial cells of the kidney tubules adhere to each other via E-cadherin; in the tip, E-cadherin is enriched apically, while in the stalk or medullary regions it is evenly distributed along the entire lateral membrane [41]. Bifurcation of the ureteric bud requires signaling through mitogen-activated protein kinase (MAPK); when MAPK activity is perturbed, E-cadherin is enriched in tip cells and localizes more basally than in the native kidney [41]. Loss of MAPK activity results in decreased phosphorylation of Paxillin but also redistribution of pPaxillin to and accumulation of vinculin at cell-cell adhesions, which could theoretically stabilize E-cadherin-mediated adhesion [41].…”