Total tetra knockout of GP138 multigene family implicated in cell interactions in Dictyostelium discoideum

The cellular slime mold Dictyostelium discoideum reproduces sexually under submerged and dark conditions. A cell surface glycoprotein gp138 has been identified as a target molecule for cell fusion-blocking antibodies, and is considered to be indispensable for the sexual cell fusion in this organism. Currently, four isoforms of gp138, DdFRP1alpha, DdFRP1beta, DdFRP2, and DdFRP3, are known. Genes encoding the latter three isoforms, GP138C, GP138A, and GP138B, have been isolated, comprising a GP138 multigene family. Here we isolated the fourth GP138 gene, GP138D, encoding DdFRP1alpha. These GP138 genes were found to cluster in a tandem array on chromosome 5, being bordered by two GP138-like sequences highly homologous to them but truncated. To clarify functional relationships among the GP138 family members, the entire GP138 region was deleted by a single knockout. Northern hybridization and western immuno-blotting analyses confirmed complete losses of GP138 mRNA and DdFRPs in the knockout strains, indicating that there are no more GP138 genes. Unexpectedly, however, the GP138-null mutants were fully potent for both sexual cell fusion and subsequent development. In addition, the original fusion-blocking antibodies detected a cell surface protein of close electrophoretic mobility to gp138 in the knockouts, suggesting the possibility that the actual target molecule of the fusion-blocking antibodies was not DdFRPs but this unidentified component. Since GP138-null mutants exhibited no obvious defects either in growth or asexual development, the real function of the GP138 family is unknown. Nevertheless, the expression levels of other developmental genes such as acaA, csaA, cotA-C, and spiA appeared to be altered in the GP138-null mutants. Therefore, it seems to have a non-critical but some role(s) during asexual development.