Betaglycan, a cell surface heparan sulphate proteoglycan, is traditionally thought to function by binding transforming growth factor type beta (TGF-beta) via its core protein and then transferring the growth factor to its signaling receptor, the type II receptor. However, there is increasing evidence that the function of betaglycan is more complex. Here, we have evaluated the role of betaglycan through adenoviral expression (Adv-BG) in myoblasts and fibroblasts and found that in Adv-BG-infected cells, the activity of p3TP-Lux and pCTGF-Luc reporter after transient transfection, as well as fibronectin synthesis, all of which are target processes for TGF-beta, were highly increased in the absence of TGF-beta. It is known that this cytokine strongly inhibits myogenin induction in myoblasts. In Adv-BG-infected myoblasts, the activity of pMyo-Luc reporter after transient transfection was strongly inhibited in the absence of TGF-beta. These effects were not precluded by applying TGF-beta-blocking antibodies, the soluble TGF-beta type II receptor, or soluble betaglycan to sequester TGF-beta present in the cell medium. Furthermore, the data suggest that the cytoplasmic domain of betaglycan is required for this TGF-beta-independent response, giving further support to a ligand-independent signaling effect for betaglycan. The process also seemed independent of Smad-2 phosphorylation, although Adv-BG infection induced p38 phosphorylation, and SB239063, an inhibitor of the p38 pathway, inhibited p3TP-Lux-driven activity. These results suggest a novel signaling mechanism for betaglycan, which is independent of the canonical TGF-beta signal pathway although it involves TGF-beta receptors and takes place through p38 pathways.