Nuclear localization of SYT, SSX and the synovial sarcoma-associated SYT-SSX fusion proteins

Abstract

Synovial sarcoma is characterized by a prevalent chromosomal translocation, t(X;18)(p11;q11). As a result of this translocation the SYT gene on chromosome 18 fuses to either the SSX1 or the SSX2 gene on the X chromosome. In this study, we generated polyclonal antibodies against the SYT and SSX2 proteins. These antibodies specifically detected both these proteins and the SYT-SSX fusion proteins in transfected COS-1 cell extracts. Indirect immunofluorescence analysis of COS-1 cells expressing tagged or untagged SYT, SSX2, SYT-SSX1 or SYT-SSX2 indicated that all these proteins are localized in the nucleus, excluding the nucleoli. The SSX2 protein exhibited a diffuse staining pattern whereas both the SYT and SYT-SSX proteins appeared in several nuclear dots. Similar nuclear dots were also detected in primary synovial sarcoma cells growing in a short-term in vitro culture. Double immunofluorescence in conjunction with confocal laser-scanning microscopy revealed that the SYT and SYT-SSX nuclear dots do not cc-localize with known nuclear structures as e.g. coiled bodies, SC35 interchromatin granules or PML bodies. The similar nuclear localization patterns of SYT and SYT-SSX suggest that the SYT-SSX fusion proteins are directed to SYT-associated nuclear domains where an abnormal function may be exerted.