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Disruption of the gene encoding the cell adhesion molecule DdCAD-1 leads to aberrant cell sorting and cell-type
Estella Wong1, Chunzhong Yang, Jun Wang
1Banting and Best Department of Medical Research and Department of Biochemistry, University of Toronto, Toronto, Ontario M5G 1L6, Canada.
Abstract:
The cadA gene in Dictyostelium encodes the Ca2+-dependent cell adhesion molecule DdCAD-1, which is expressed soon after the initiation of development. To investigate the biological role of DdCAD-1, the cadA gene was disrupted by homologous recombination. The cadA-null cells showed a 50% reduction in EDTA-sensitive cell adhesion. The remaining EDTA-sensitive adhesion sites were resistant to dissociation by anti-DdCAD-1 antibody, suggesting that they were distinct adhesion sites. Cells that lacked DdCAD-1 were able to complete development and form fruiting bodies. However, they displayed abnormal slug morphology and culmination was delayed by approximately 6 hours. The yield of spores was reduced by approximately 50%. The proportion of prestalk cells in cadA(-) slugs showed a 2.5-fold increase over the parental strain. When cadA(-) cells were transfected with pcotB::GFP to label prespore cells, aberrant cell-sorting patterns in slugs became apparent. When mutant prestalk cells were mixed with wild-type prespore cells, mutant prestalk cells were unable to return to the anterior position of chimeric slugs, suggesting defects in the sorting mechanism. The wild-type phenotype was restored when cadA(-) cells were transfected with a cadA-expression vector. These results indicate that, in addition to cell-cell adhesion, DdCAD-1 plays a role in cell type proportioning and pattern formation.
Insights
The cadA gene disruption in Dictyostelium reduced cell adhesion and altered slug morphology. DdCAD-1 is crucial for cell sorting, proportioning, and pattern formation during development.
Area of Science:
- Cell biology
- Developmental biology
- Molecular genetics
Background:
- The cadA gene in Dictyostelium discoideum encodes the Ca2+-dependent cell adhesion molecule DdCAD-1.
- DdCAD-1 is expressed early in development and plays a role in cell adhesion.
Purpose of the Study:
- To investigate the biological role of DdCAD-1 in Dictyostelium development.
- To understand DdCAD-1's function beyond cell adhesion, particularly in cell sorting and pattern formation.
Main Methods:
- Homologous recombination was used to disrupt the cadA gene, creating cadA-null cells.
- Cell adhesion assays, morphological observations, and genetic manipulation (transfection) were employed.
- Reporter gene (GFP) expression was used to visualize cell types and sorting patterns.
Main Results:
- cadA-null cells exhibited a 50% reduction in EDTA-sensitive cell adhesion.
- Development was completed, but slug morphology was abnormal, culmination was delayed, and spore yield decreased by 50%.
- Aberrant cell-sorting patterns were observed, with prestalk cells showing defects in returning to anterior positions.
Conclusions:
- DdCAD-1 is essential for normal cell-cell adhesion in Dictyostelium.
- Beyond adhesion, DdCAD-1 plays a critical role in cell type proportioning and pattern formation during development.
- Defects in DdCAD-1 function lead to significant disruptions in developmental processes, including cell sorting and tissue patterning.