Cellular segregation in co-cultures driven by differential adhesion and contractility on distinct time scales

Abstract

Cellular sorting and pattern formation is crucial for many biological processes such as development, tissue regeneration, and cancer progression. Prominent physical driving forces for cellular sorting are differential adhesion and contractility. Here, we studied the segregation of epithelial co-cultures containing highly contractile, ZO1/2-depleted MDCKII cells (dKD) and their wildtype (WT) counterparts using multiple quantitative, high-throughput methods to monitor their dynamical and mechanical properties. We observe a time-dependent segregation process, governed mainly by differential contractility on short (< 5 h) and differential adhesion on long (> 5 h) time scales, respectively. The overly contractile dKD cells exert strong lateral forces on their WT neighbors, thereby apically depleting their surface area. This is reflected in a six-fold difference in excess surface area between both cell types. The lateral forces lead to a four- to six-fold increase in tension at all junctions that are in contact with the contractile cells including the interface between heterotypic cell-cell contacts. Concomitantly, the tight junction-depleted, contractile cells exhibit weaker cell-cell adhesion. Drug-induced contractility reduction delays the initial segregation but ceases to change the final demixed state, rendering differential adhesion the dominant segregation force at longer time scales. This well-controlled model system shows how cell sorting is accomplished through a complex interplay between differential adhesion and contractility and can be explained largely by generic physical driving forces.

Cell-substrate distance fluctuations of confluent cells enable fast and coherent collective migration

“…Further studies showed that the balance between cellular contractility and differential adhesion between cells leads to a time-dependent demixing in WT/dKD confluent co-cultures. 26 Here, we show that ZO-1/ZO-2 depleted cells display less coordinated movement of confluent cell monolayers than WT cells, even in a pre-jammed state. We attribute the loss of collective cell rearrangement to reduced actively driven cell-substrate-fluctuations (micromotion) along with lower traction forces to overcome cell-adhesion forces that are responsible for friction between substrate and cell monolayer.…”

“…We found pronounced actin and myosin gradients in dKD cells, suggesting reduced basal contractility as recently reported for glassy substrates. 26 The altered basal actomyosin skeleton in dKD cells explains the reduced average traction forces in dKD monolayers. The monolayers with higher density exerted lower traction forces on average regardless of the cell type, as also found by Saraswathibhatla et al 10 We could rule out that proliferation is responsible for the observed alteration of traction forces but noticed that the average traction forces in the cell layer depend on the composition of the cell monolayer.…”

“…Further studies showed that the balance between cellular contractility and differential adhesion between cells leads to a time-dependent demixing in MDCKII cell (wild-type) WT and dKD confluent co-cultures. 26 These findings show some discrepancies not only because of the different cell types used and the proteins chosen for tight junction interference, but also the differences in migration assays (scratch-assays, patterning, or stencils) used.…”

Tight junctions regulate lumen morphology via hydrostatic pressure and junctional tension

“…The TJ scaffold proteins ZO1 and ZO2 have been shown to be important regulators of the apical actin cortex [9][10][11] . In addition, ZO1 is known to inhibit myosin activity via inhibition of the RhoA exchange factor GEF-H1 25 , and depletion of ZO1 leads to increased junctional tension 26,27 . Our quantification of endogenous junctional myosin-IIa levels in combination with junction tension measurements confirm the key role of ZO proteins in regulating apicaljunctional contractility via myosin inhibition (Fig.…”