Evidence of an Evolutionarily Conserved LMBR1 Domain-Containing Protein That Associates with Endocytic Cups and Plays a Role in Cell Migration in Dictyostelium discoideum

TheampA gene plays a role inDictyostelium discoideum cell migration. Loss ofampA function results in reduced ability of growing cells to migrate to folic acid and causes small plaques on bacterial lawns, while overexpression of AmpA results in a rapid-migration phenotype and correspondingly larger plaques than seen with wild-type cells. To help understand how theampA gene functions, second-site suppressors were created by restriction enzyme-mediated integration (REMI) mutagenesis. These mutants were selected for their ability to reduce the large plaque size of the AmpA overexpresser strain. Thelmbd2B gene was identified as a suppressor of an AmpA-overexpressing strain. Thelmbd2B gene product belongs to the evolutionarily conserved LMBR1 protein family, some of whose known members are endocytic receptors associated with human diseases, such as anemia. In order to understandlmbd2B function, mRFP fusion proteins were created andlmbd2B knockout cell lines were established. Our findings indicate that the LMBD2B protein is found associated with endocytic cups. It colocalizes with proteins that play key roles in endocytic events and is localized to ruffles on the dorsal surfaces of growing cells. Vegetativelmbd2B -null cells display defects in cell migration. These cells have difficulty sensing the chemoattractant folic acid, as indicated by a decrease in their chemotactic index.lmbd2B -null cells also appear to have difficulty establishing a front/back orientation to facilitate migration. A role forlmbd2B in development is also suggested. Our research gives insight into the function of a previously uncharacterized branch of the LMBR1 family of proteins. We provide evidence of an LMBR1 family plasma membrane protein that associates with endocytic cups and plays a role in chemotaxis.