CD99, a transmembrane protein encoded by MIC2 gene is involved in multiple 
cellular events including cell adhesion and migration, apoptosis, cell 
differentiation and regulation of protein trafficking either in physiological or 
pathological conditions. In osteosarcoma, CD99 is expressed at low levels and 
functions as a tumour suppressor. The full-length protein (CD99wt) and the 
short-form harbouring a deletion in the intracytoplasmic domain (CD99sh) have 
been associated with distinct functional outcomes with respect to tumour 
malignancy. In this study, we especially evaluated modulation of cell-cell 
contacts, reorganisation of the actin cytoskeleton and modulation of signalling 
pathways by comparing osteosarcoma cells characterised by different metastasis 
capabilities and CD99 expression, to identify molecular mechanisms responsible 
for metastasis. Our data indicate that forced expression of CD99wt induces 
recruitment of N-cadherin and β-catenin to adherens junctions. In addition, 
transfection of CD99wt inhibits the expression of several molecules crucial to 
the remodelling of the actin cytoskeleton, such as ACTR2, ARPC1A, 
Rho-associated, coiled-coil containing protein kinase 2 (ROCK2) as well as 
ezrin, an ezrin/radixin/moesin family member that has been clearly associated 
with tumour progression and metastatic spread in osteosarcoma. Functional 
studies point to ROCK2 as a crucial intracellular mediator regulating 
osteosarcoma migration. By maintaining c-Src in an inactive conformation, CD99wt 
inhibits ROCK2 signalling and this leads to ezrin decrease at cell membrane 
while N-cadherin and β-catenin translocate to the plasma membrane and function 
as main molecular bridges for actin cytoskeleton. Taken together, we propose 
that the re-expression of CD99wt, which is generally present in osteoblasts but 
lost in osteosarcoma, through inhibition of c-Src and ROCK2 activity, manages to 
increase contact strength and reactivate stop-migration signals that counteract 
the otherwise dominant promigratory action of ezrin in osteosarcoma cells.