The intracellular hyaluronan receptor RHAMM/IHABP interacts with microtubules and actin filaments

We reported recently on the intracellular localisation of the hyaluronan re ceptor RHAMM/IHABP in human cancer cells. Here we describe the colocalisati on of RHAMM/IHABP proteins with microtubules, both in interphase and dividi ng cells, suggesting that RHAMM/IHABP represents a novel member of the fami ly of microtubule-associated proteins (MAPs), We have identified four diffe rent splice variants of RHAMM/IHABP, all of which colocalise, at least tran siently; with microtubules when expressed as GFP fusion proteins in HeLa ce lls. Using microtubule-binding assays and transient transfection experiment s of deletion-bearing RHAMM/IHABP mutants, we localised the microtubule-bin ding region to the extreme N terminus of RHAMM/IHABP. This interaction doma in is composed of two distinct subdomains, one of which is sufficient to me diate binding to the mitotic spindle while both domains are required for bi nding of RHAMM/IHABP proteins to interphase microtubules, Sequence analysis revealed that the projection domain of RHAMM/IHABP is predicted to form co iled-coils, implying that RHAMM/IHABP represents a filamentous protein capa ble of interacting with other proteins and we found that RHAMM/IHABP intera cts with actin filaments in vivo and in vitro, Moreover, in vitro translate d RHAMM/IHABP isoforms efficiently bind to immobilised calmodulin in a Ca2 dependent manner via a calmodulin-binding site within the projection domai n of RHAMM/IHABP (residues 574-602), Taken together, our results strongly suggest that RHAMM/IHABP is a ubiquiti ously expressed, filamentous protein capable of interacting with microtubul es and microfilaments and not, as numerous previous reports suggest, a cell surface receptor for the extracellular matrix component hyaluronan.