Smads are important intracellular signaling effectors for transforming growth factor-␤ (TGF-␤) and related factors. Proper TGF-␤ signaling requires precise control of Smad functions. In this study, we have identified a novel HECT class ubiquitin E3 ligase, designated Smurf2, that negatively regulates Smad2 signaling. In both yeast two-hybrid and in vitro binding assays, we found that Smurf2 could interact with receptor-activated Smads (R-Smads), including Smad1, Smad2, and Smad3 but not Smad4. Ectopic expression of Smurf2 was sufficient to reduce the steady-state levels of Smad1 and Smad2 but not Smad3 or Smad4. Significantly, Smurf2 displayed preference to Smad2 as its target for degradation. Furthermore, Smurf2 exhibited higher binding affinity to activated Smad2 upon TGF-␤ stimulation. The ability of Smurf2 to promote Smad2 destruction required the HECT catalytic activity of Smurf2 and depended on the proteasome-dependent pathway. Consistent with these results, Smurf2 potently reduced the transcriptional activity of Smad2. These data suggest that a ubiquitin/proteasome-dependent mechanism is important for proper regulation of TGF-␤ signaling. Smads are important intracellular signaling effectors for transforming growth factor-␤ (TGF-␤)1 and related factors (1-5). There are three subgroups of Smads, receptor-activated Smads (or R-Smads, e.g. mammalian Smad1, -2, -3, -5, and -8), the common Smads (e.g. mammalian Smad4), and the inhibitory Smads (e.g. mammalian Smad6 and Smad7). Smads have two highly conserved domains at the N terminus (the MH1 domain) and the C terminus (the MH2 domain). The MH1 domain is responsible for DNA binding, whereas the MH2 domain mediates the transcriptional activity of R-Smads, Smad oligomerization, and Smad-receptor interaction (1-5). The key regulation of R-Smad activity is the ligand-induced type I receptor-mediated phosphorylation, which leads to a series of downstream events in TGF-␤ signal transduction. The phosphorylation in the C-terminal SXS motif results in the release of mutual inhibitory effects of MH1 and MH2 domains (6), followed by R-Smad association with Smad4 and nuclear import of the Smad complex (7-9). In the nucleus, the Smad complex cooperates with transcription factors such as AP-1,

The transcription corepressor CtBP is often recruited to the target promoter via interaction with a conserved PxDLS motif in the interacting repressor. In this study, we demonstrate that CtBP1 was SUMOylated and that its SUMOylation profoundly affected its subcellular localization. SUMOylation occurred at a single Lys residue, Lys428, of CtBP1. CtBP2, a close homolog of CtBP1, lacked the SUMOylation site and was not modified by SUMO-1. Mutation of Lys428 into Arg (K428R) shifted CtBP1 from the nucleus to the cytoplasm, while it had little effect on its interaction with the PxDLS motif. Consistent with a change in localization, the K428R mutation abolished the ability of CtBP1 to repress the E-cadherin promoter activity. Notably, SUMOylation of CtBP1 was inhibited by the PDZ domain of nNOS, correlating with the known inhibitory effect of nNOS on the nuclear accumulation of CtBP1. This study identifies SUMOylation as a regulatory mechanism underlying CtBP1-dependent transcriptional repression.

Tumor suppressor Smad4/DPC4 is a central intracellular signal transducer for transforming growth factor-␤ (TGF-␤) signaling. We recently reported that transcriptional potential of Smad4 was regulated by SUMOylation in transfected HeLa cells (1), but the precise mechanism and function of Smad4 SUMOylation in TGF-␤ signaling remain to be elucidated. Here, we describe the regulation of TGF-␤ signaling by SUMOylation through the control of Smad4 metabolic stability and subcellular localization. We found that SUMO-1 overexpression strongly increases Smad4 levels, while inhibition of SUMOylation by small interfering RNA (siRNA)-mediated knockdown of the E2 enzyme Ubc9 reduces endogenous Smad4 levels. Concomitantly, SUMO-1 overexpression enhances and Ubc9 knockdown reduces levels of intranuclear Smad4, growth inhibitory response, as well as transcriptional responses to TGF-␤. Comparison of wild type and mutant forms of Smad4 for SUMOylation, ubiquitination, and half-life allows the conclusion that SUMO-1 modification serves to protect Smad4 from ubiquitin-dependent degradation and consequently enhances the growth inhibitory and transcriptional responses of Smad4.

The oncolytic viruses now hold a promise of new therapeutic strategy for cancer. Its concept has inspired a wave of commercial research and development activities for the products of this category in China since 1998. The first commercialized oncolytic virus product in the world, Oncorine (H101), developed by Shanghai Sunway Biotech Co., Ltd since 1999, was approved by Chinese SFDA in November, 2005 for nasopharyngeal carcinoma in combination with chemotherapy after the phase III clinical trial, and finally acquired GMP certificate in August, 2006. This review introduces how Oncorine was successfully developed in China, and how the Chinese market responded after it was launched into the market in 2006.

Smads are important intracellular effectors in signaling pathways of the transforming growth factor-␤ (TGF-␤) superfamily. Upon activation by TGF-␤, receptor-phosphorylated Smads form a complex with tumor suppressor Smad4/DPC4, and the Smad complexes then are imported into the nucleus. Although diverse pathways regulate the activity and expression of receptorphosphorylated and inhibitory Smads, cellular factors modulating the activity of the common Smad4 remain unidentified. Here we describe the involvement of the small ubiquitin-like modifier-1 (SUMO-1) conjugation pathway in regulating the growth inhibitory and transcriptional responses of Smad4. The MH1 domain of Smad4 was shown to associate physically with Ubc9, the ubiquitin carrier protein (E2) conjugating enzyme in sumoylation. In cultured cells, Smad4 is modified by SUMO-1 at the endogenous level. The sumoylation sites were identified as two evolutionarily conserved lysine residues, Lys-113 and Lys-159, in the MH1 domain. We found that the mutations at Lys-113 and Lys-159 did not alter the ability of Smad4 to form a complex with Smad2 and FAST on the Mix.2 promoter. Importantly, SUMO-1 overexpression enhanced TGF-␤-induced transcriptional responses. These findings identify sumoylation as a unique mechanism to modulate Smad4-dependent cellular responses.Transforming growth factor-␤ (TGF-␤) 1 is a secreted multifunctional protein that exhibits a diverse set of cellular responses including cell proliferation and differentiation. TGF-␤ functions as a potent growth inhibitor and induces the expression of a variety of genes during growth and development (1-4).TGF-␤ signals are transduced by transmembrane serine/threonine kinase receptors and intracellular effectors called Smads (2,5,6). Upon TGF-␤ stimulation, Smad2 and/or Smad3 are phosphorylated by the activated type I receptor and then form complexes with Smad4. The heteromeric complexes of R-Smads and Smad4 then are translocated into the nucleus, where they exert ligand-induced changes in the transcription of a variety of genes (2, 6, 7). The heteromeric Smad complex activates transcription through its ability to cooperate functionally with several promoter-specific transcription factors and/or to bind specific DNA sequences (7,8).Proper TGF-␤ signaling requires precise control of Smad functions. Recent studies have shown that R-Smads are modified post-translationally by ubiquitin and can be removed irreversibly by the proteasome-mediated degradation system (9 -13). A number of ubiquitin-related proteins also are present in eukaryotic cells. These proteins, including the small ubiquitinlike modifier-1 (SUMO-1), utilize a conjugation system that is similar to ubiquitination (14 -16). SUMO-1 uses Aos1/Uba2 as E1 activating enzymes (17, 18) and Ubc9 as an E2 conjugating enzyme (19,20). Unlike ubiquitination, SUMO-1 modifications of target proteins do not promote their degradation. In contrast, SUMO-1 modifications of I B preclude its ubiquitination (21). Functional consequences of SUMO-1 modification vary dep...