Cell Systems Bioelectricity: How Different Intercellular Gap Junctions Could Regionalize a Multicellular Aggregate

Electric potential distributions can act as instructive pre-patterns for development, regeneration, and tumorigenesis in cell systems. The biophysical states influence transcription, proliferation, cell shape, migration, and differentiation through biochemical and biomechanical downstream transduction processes. A major knowledge gap is the origin of spatial patterns in vivo, and their relationship to the ion channels and the electrical synapses known as gap junctions. Understanding this is critical for basic evolutionary developmental biology as well as for regenerative medicine. We computationally show that cells may express connexin proteins with different voltage-gated gap junction conductances as a way to maintain multicellular regions at distinct membrane potentials. We show that increasing the multicellular connectivity via enhanced junction function does not always contribute to the bioelectrical normalization of abnormally depolarized multicellular patches. From a purely electrical junction view, this result suggests that the reduction rather than the increase of specific connexin levels can also be a suitable bioelectrical approach in some cases and time stages. We offer a minimum model that incorporates effective conductances ultimately related to specific ion channel and junction proteins that are amenable to external regulation. We suggest that the bioelectrical patterns and their encoded instructive information can be externally modulated by acting on the mean fields of cell systems, a complementary approach to that of acting on the molecular characteristics of individual cells. We believe that despite the limitations of a biophysically focused model, our approach can offer useful qualitative insights into the collective dynamics of cell system bioelectricity.

AQP3 Increases Intercellular Cohesion in NSCLC A549 Cell Spheroids through Exploratory Cell Protrusions

Tumor cell aggregation is critical for cell survival following the loss of extracellular matrix attachment and dissemination. However, the underlying mechanotransduction of clustering solitary tumor cells is poorly understood, especially in non-small cell lung cancers (NSCLC). Here, we examined whether cell surface protrusions played an important role in facilitating the physical contact between floating cells detached from a substrate. We employed poly-2-hydroxyethyl methacrylate-based 3D culture methods to mimic in vivo tumor cell cluster formation. The suprastructural analysis of human NSCLC A549 cell spheroids showed that finger-like protrusions clung together via the actin cytoskeleton. Time-lapse holotomography demonstrated that the finger-like protrusions of free-floating cells in 3D culture displayed exploratory coalescence. Global gene expression analysis demonstrated that the genes in the organic hydroxyl transport were particularly enriched in the A549 cell spheroids. Particularly, the knockdown of the water channel aquaporin 3 gene (AQP3) impaired multicellular aggregate formation in 3D culture through the rearrangement of the actomyosin cytoskeleton. Moreover, the cells with reduced levels of AQP3 decreased their transmigration. Overall, these data indicate that cell detachment-upregulated AQP3 contributes to cell surface protrusions through actomyosin cytoskeleton remodeling, causing the aggressive aggregation of free-floating cells dependent on the property of the substratum and collective metastasis.

Unification of aggregate growth models by emergence from cellular and intracellular mechanisms

Multicellular aggregate growth is regulated by nutrient availability and removal of metabolites, but the specifics of growth dynamics are dependent on cell type and environment. Classical models of growth are based on differential equations. While in some cases these classical models match experimental observations, they can only predict growth of a limited number of cell types and so can only be selectively applied. Currently, no classical model provides a general mathematical representation of growth for any cell type and environment. This discrepancy limits their range of applications, which a general modelling framework can enhance. In this work, a hybrid cellular Potts model is used to explain the discrepancy between classical models as emergent behaviours from the same mathematical system. Intracellular processes are described using probability distributions of local chemical conditions for proliferation and death and simulated. By fitting simulation results to a generalization of the classical models, their emergence is demonstrated. Parameter variations elucidate how aggregate growth may behave like one classical growth model or another. Three classical growth model fits were tested, and emergence of the Gompertz equation was demonstrated. Effects of shape changes are demonstrated, which are significant for final aggregate size and growth rate, and occur stochastically.

Rapid analysis of cell-generated forces within a multicellular aggregate using microsphere-based traction force microscopy

We measure cell-generated forces from the deformations of elastic microspheres embedded within multicellular aggregates. Using a computationally efficient analytical model, we directly obtain the full 3D mapping of surface stresses within minutes.

A three dimensional model of multicellular aggregate compression

We present a three-dimensional elasto-plastic model describing the mechanical behaviour of living multicellular aggregates and we numerically solve it to reproduce the biological observations of its response to a uniaxial compression test.

Individual and collective behaviour in cellular slime mould development: contributions of John Bonner (1920-2019)

John Bonner used the cellular slime moulds to address issues that lie at the heart of evolutionary and developmental biology. He did so mostly by combining acute observation and a knack for asking the right questions with the methods of classical embryology. The present paper focusses on his contributions to understanding two phenomena that are characteristic of development in general: chemotaxis of single cells to an external attractant, and spatial patterning and proportioning of cell types in the multicellular aggregate. Brief mention is also made of other areas of slime mould biology where he made significant inputs. He saw cellular slime moulds as exemplars of development and worthy of study in their own right. His ideas continue to inspire researchers.

Quorum sensing controls Vibrio cholerae multicellular aggregate formation

Bacteria communicate and collectively regulate gene expression using a process called quorum sensing (QS). QS relies on group-wide responses to signal molecules called autoinducers. Here, we show that QS activates a new program of multicellularity in Vibrio cholerae. This program, which we term aggregation, is distinct from the canonical surface-biofilm formation program, which QS represses. Aggregation is induced by autoinducers, occurs rapidly in cell suspensions, and does not require cell division, features strikingly dissimilar from those characteristic of V. cholerae biofilm formation. Extracellular DNA limits aggregate size, but is not sufficient to drive aggregation. A mutagenesis screen identifies genes required for aggregate formation, revealing proteins involved in V. cholerae intestinal colonization, stress response, and a protein that distinguishes the current V. cholerae pandemic strain from earlier pandemic strains. We suggest that QS-controlled aggregate formation is important for V. cholerae to successfully transit between the marine niche and the human host.