The cancer‐related protein SSX2 interacts with the human homologue of a Ras‐like GTPase interactor, RAB3IP, and a novel nuclear protein, SSX2IP

Abstract

The SSX gene family is composed of at least five functional and highly homologous members, SSX1 to SSX5, that are normally expressed in only the testis and thyroid. SSX1, SSX2, or SSX4 may be fused to the SYT gene as a result of the t(X;18) translocation in synovial sarcoma. In addition, the SSX1, SSX2, SSX4, and SSX5 genes were found to be aberrantly expressed in several other malignancies, including melanoma. The SSX proteins are localized in the nucleus and are diffusely distributed. In addition, they may be included in polycomb-group nuclear bodies. Other studies have indicated that the SSX proteins may act as transcriptional repressors. As a first step toward the elucidation of the cellular signaling networks in which the SSX proteins may act, we used the yeast two-hybrid system to identify SSX2-interacting proteins. By doing so, two novel human proteins were detected: RAB3IP, the human homolog of an interactor of the Ras-like GTPase Rab3A; and a novel protein, SSX2IP. RAB3IP did not interact with either SSX1, SSX3, or SSX4 in the yeast two-hybrid system, whereas SSX2IP interacted with SSX3 but not with either SSX1 or SSX4. Further analysis of deletion mutants showed that both RAB3IP and SSX2IP interact with the N-terminal moiety of the SSX2 protein. Immunofluorescence analyses of transfected cells revealed that the RAB3IP protein is normally localized in the cytoplasm. However, coexpression of both RAB3IP and SSX2 led to colocalization of both proteins in the nucleus. Likewise, the SSX2IP protein was found to be colocalizing with SSX2 in the nucleus. By performing glutathione-S-transferase pull-down assays, we found that both RAB3IP and SSX2IP interact directly with SSX2 in vitro. These newly observed protein/protein interactions may have important implications for the mechanisms underlying normal and malignant cellular growth.

Cancer/testis antigen SSX2 enhances invasiveness in MCF-7 cells by repressing ERα signaling

“…In addition, the conserved C-terminus of SSX2 named as SSXRD (SSX repression domain) is associated with transcriptional repressor regulation (19,20). The SSX KRAB domain interacts with the RAB3IP (RAB3A interacting protein) and SSX2IP (SSX2 interacting protein), while the SSXRD domain interacts with LHX4 (LIM homeobox 4) and PcG protein (21,22 Vector construction. Wild-type SSX2 ORF was isolated by PCR from human testis cDNA (forward primer is 5'-GGGGAATTC ATGAACGGAGACGACGCCTT-3' and reverse primer is 5'-CGCGGATCCTCACCAGCTGTTTTCTCTCA-3').…”

The Tumor Antigen Repertoire Identified in Tumor-Bearing Neu Transgenic Mice Predicts Human Tumor Antigens

“…Many of the antigenic proteins play an important role in tumorigenesis. For example, Rab3ip is a proto-oncogene that interacts with the cancer-testis antigen SSX2 (8). Gsn, an actinsevering protein, has tumor suppressor function and has been reported to be down-regulated in human ovarian and breast cancer (9)(10)(11).…”

Role of Scaffold Protein Afadin Dilute Domain-interacting Protein (ADIP) in Platelet-derived Growth Factor-induced Cell Movement by Activating Rac Protein through Vav2 Protein

“…Another line of evidence indicates that the human orthologue of ADIP (human ADIP) is the synovial sarcoma X breakpoint 2-interacting protein based on its interaction with SSX2 (46). The expression of SSX2 is usually very low in normal tissues except for the testis but is up-regulated in various types of cancer, including melanoma (47).…”

“…The expression of SSX2 is usually very low in normal tissues except for the testis but is up-regulated in various types of cancer, including melanoma (47). In contrast to SSX2, human ADIP (synovial sarcoma X breakpoint 2-interacting protein) as well as mouse ADIP is expressed in a wide variety of normal tissues (19,46). Human ADIP was also identified as a leukemiaassociated antigen and was detected in 33% of patients with acute myeloid leukemia but not in normal donors (48).…”