Molecular cloning of a novel hyaluronan receptor that mediates tumor cell motility

Abstract

Abstract. A eDNA encoding a unique hyaluronan receptor has been molecularly cloned from a ~GTll 3T3 eDNA expression library. Immunoblot analyses of cell lysates, using antibodies to peptides encoded in the eDNA, specifically react with a 58-kD protein. This protein is regulated by the mutant H-ras gene in cells containing a metallothionein promoter H-ras hybrid gene. Further, antibodies to peptide sequences encoded in the eDNA block the increase in locomotion resulting from induction of the mutant H-ras gene in this cell line. In a transblot assay, the bacterially expressed protein binds to biotinylated hyaluronan. Antibodies to peptides encoded in the cDNA react in immunoblot assays with the 58-and 52-kD proteins of a novel hyaluronan receptor complex previously implicated in cell locomotion. Furthermore, antibodies specific to the 58-and 52-kD proteins, which block ms-induced locomotion, also cross-react with the expressed, encoded protein. The gene product described here appears to be a new type of hyaluronan receptor that is involved in cell locomotion. It is named RHAMM, an acronym for receptor for hyaluronanmediated motility. THE transforming oncogene H-ras has been reported to promote cell locomotion (17), although the regulatory mechanisms remain unknown. Several observations suggest that when this oncogene promotes locomotion, the mechanisms are complex and involve, at least, the release of autocrine motility factor(s) (14,20), growth factors (14), and the glycosaminoglycan hyaluronan (HA) ~ (20, 34). In particular, HA appears to function as an autocrine mechanism for stimulating maximal locomotion in ms-transformed cells (34). Further, it is also required for the ability of an autocrine motility-stimulating factor to promote breast carcinoma cell locomotion (20). We have shown that HA-promoted, rastransformed cell locomotion requires the presence of a novel hyaluronan receptor complex termed I/ARC (34). This complex of proteins occurs at the cell surface or is released as soluble proteins of 72, 68, 58, and 52 kD (32). The complex is tightly regulated in vitro (30) and expressed on the leading lamellae and perinuclear region only on rapidly locomoting cells (29, 31). Both polyclonal and monoclonal antibodies (pAbs and mAbs, respectively) prepared against this complex block cell locomotion regulated by mutant ms (34). In Address all correspondence and requests for reprints to E. A. Turley, Manitoba Institute of Cell Biology, 100 Olivia St., Winnipeg, Manitoba R3E OV9 Canada.1. Abbreviations used in this paper: HA, hyaluronan; HARC, hyaluronan receptor complex; pAb, polyclonal antibody; RHAMM, receptor for HAmediated motility, a recent study, we have shown that these blocking mAbs are specific to the 58-and 52-kD proteins, that these proteins are isoforms of each other, and, further, that these proteins are the HA-binding component of HARC (Turley, E. A., K. Home, and V. Cripps, manuscript submitted for publication).We have used the blocking antibodies specific to the 58-and 52-kD HARC proteins to screen...

Two-marker protein profile predicts poor prognosis in patients with early rectal cancer

“…We have recently identified RHAMM in combination with p21 as highly conducive towards a severely adverse prognosis in microsatellite instability-high (MSI-H) colorectal cancer (Zlobec et al, 2008a). When expressed at the cell surface, RHAMM serves as a receptor for hyaluronic acid and has been implicated in both tumour cell motility and invasiveness (Hardwick et al, 1992) . In N0 rectal cancers, RHAMM positivity may be indicative of a particularly invasive tumour phenotype prone to the release of micrometastases from the primary tumour.…”

RHAMM expression and isoform balance predict aggressive disease and poor survival in multiple myeloma

“…7 RHAMM was cloned in 1992 and described as a hyaluronan-binding protein. 8,9 A variant of RHAMM, which is predominantly cytoplasmic and involved in multiple signaling pathways, 10,11 is transforming and essential for ras transformation. 12 Human full-length RHAMM (RHAMM FL ) was cloned in 1996 as a predicted 85-to 95-kDa protein encoded by 18 exons.…”

Hyaluronidases - a group of neglected enzymes

“…H A interacts with a variety of receptors and binding proteins on the surface of cells, of which CD-44 and RHAMM have been characterized most extensively (Aruffo et al, 1990;Hardwick et al, 1992;Yang et al, 1994). It therefore seems likely that the formation and degradation of HA and the concomitant modulation of the physicochemical properties of the ECM have to be tightly controlled.…”