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Immortalized connexin43 knockout cell lines display a subset of biological properties associated with the transformed
K D Martyn1, W E Kurata, B J Warn-Cramer
1Molecular Carcinogenesis Program, Cancer Research Center, University of Hawaii at Manoa 96813, USA.
Abstract:
Immortalized cells from embryonic connexin43 knockout mice (Cx43-/-) and homozygous littermates (Cx43+/+) were cloned and characterized to determine whether the absence of Cx43 function would induce observable phenotypic changes. Cells of the Cx43+/+ clones expressed Cx43 and engaged in gap junctional communication with 10-12 neighboring cells. The Cx43-/- cells were devoid of Cx43 and communicated to less than 1 cell. Electrophysiological analysis indicated that the Cx43-/- cells communicated through Cx45 channels from 8-80-fold less than did the Cx43+/+ subclones, which seemed to communicate through Cx43 and Cx45 channels. The Cx43-/- clones grew at faster rates and to higher saturation densities, had a more spindly morphology, were more refractile, and adhered less well to the substratum than did the Cx43+/+ clones. Reintroducing the Cx43 gene into the Cx43-/- clones resulted in three subclones that communicated to 3-4 cells. Partial restoration of gap junctional communication in the three subclones was accompanied by reduced growth rates and saturation densities (2-fold compared to that of parental Cx43-/- clones) but no reversions in morphology or cell-substratum adhesion. The increased growth rates and saturation densities, altered morphology, and decreased cell adhesion displayed by the Cx43-/- clones reflect a subset of the properties of transformed cells. These studies advance the hypothesis that loss of Cx43 function during development may cause cells to acquire a preneoplastic condition.
Insights
Loss of connexin43 (Cx43) function in mice cells led to faster growth, altered morphology, and reduced adhesion. Restoring Cx43 partially reversed these changes, suggesting Cx43 loss may predispose cells to a preneoplastic state.
Area of Science:
- Cell Biology
- Developmental Biology
- Molecular Biology
Background:
- Connexin43 (Cx43) is a key protein in gap junctional communication, essential for intercellular signaling.
- Understanding Cx43's role is crucial for comprehending cell growth regulation and tissue development.
Purpose of the Study:
- To investigate the phenotypic consequences of connexin43 (Cx43) absence in immortalized mouse embryonic cells.
- To determine if Cx43 deficiency induces changes indicative of cellular transformation or preneoplasia.
Main Methods:
- Cloning and characterization of Cx43 knockout (Cx43-/-) and wild-type (Cx43+/+) mouse embryonic cells.
- Electrophysiological analysis to assess gap junctional communication.
- Comparative analysis of cell growth rates, morphology, adhesion, and saturation densities.
Main Results:
- Cx43-/- cells exhibited significantly reduced gap junctional communication compared to Cx43+/+ cells.
- Cx43-/- cells showed increased growth rates, higher saturation densities, altered morphology, and decreased substratum adhesion.
- Reintroduction of Cx43 partially restored gap junctional communication and reduced growth rates and saturation densities.
Conclusions:
- Absence of Cx43 function induces phenotypic changes resembling those of transformed cells.
- Loss of Cx43 may contribute to a preneoplastic condition during cellular development.
- Cx43 plays a critical role in regulating cell proliferation, morphology, and adhesion.