Vitamin D 3 up-regulated protein 1 (VDUP1) is a stressresponse gene that is up-regulated by 1,25(OH) 2 D 3 in many cells. It has been reported that VDUP1 expression is reduced in many tumor cells and the enforced expression of VDUP1 inhibits cell proliferation by arresting cell cycle progression. Here, we found that VDUP1 À/À fibroblast cells proliferated more rapidly compared with wild-type cells with reduced expression of p27 kip1 , a cyclin-dependent kinase inhibitor. JAB1 is known to interact with p27 kip1 and to decrease the stability of p27 kip1 . VDUP1 interacted with JAB1 and restored JAB1-induced suppression of p27 kip1 stability. In this process, VDUP1 blocked the JAB1-mediated translocation of p27 kip1 from the nucleus to the cytoplasm. In addition, VDUP1 inhibited JAB1-mediated activator protein-1 activation and cell proliferation. Taken together, these results indicate that VDUP1 is a novel factor of p27 kip1 stability via regulating JAB1. (Cancer Res 2005; 65(11): 4485-9)

Mammalian sterile 20-like kinases 1 and 2 (Mst1 and Mst2, respectively) are potent serine/threonine kinases that are involved in cell proliferation and cell death. To investigate the physiological functions of Mst1 and Mst2, we generated Mst1 and Mst2 mutant mice. Mst1 ؊/؊ and Mst2 ؊/؊ mice were viable and fertile and developed normally, suggesting possible functional overlaps between the two genes. A characterization of heterozygous and homozygous combinations of Mst1 and Mst2 mutant mice showed that mice containing a single copy of either gene underwent normal organ development; however, Mst1 ؊/؊ ; Mst2 ؊/؊ mice lacking both Mst1 and Mst2 genes started dying in utero at approximately embryonic day 8.5. Mst1 ؊/؊ ; Mst2 ؊/؊ mice exhibited severe growth retardation, failed placental development, impaired yolk sac/embryo vascular patterning and primitive hematopoiesis, increased apoptosis in placentas and embryos, and disorganized proliferating cells in the embryo proper. These findings indicate that both Mst1 and Mst2 kinases play essential roles in early mouse development, regulating placental development, vascular patterning, primitive hematopoiesis, and cell proliferation and survival.

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p21Cip1 is an inhibitor of cell cycle progression that promotes G 1 -phase arrest by direct binding to cyclindependent kinase and proliferating cell nuclear antigen. Here we demonstrate that mitogenic stimuli, such as epidermal growth factor treatment and oncogenic Ras transformation, induce p21 Cip1 downregulation at the posttranslational level. This downregulation requires the sustained activation of extracellular signal-regulated kinase 2 (ERK2), which directly interacts with and phosphorylates p21 Cip1 , promoting p21 Cip1 nucleocytoplasmic translocation and ubiquitin-dependent degradation, thereby resulting in cell cycle progression. ERK1 is not likely involved in this process. Phosphopeptide analysis of in vitro ERK2-phosphorylated p21Cip1 revealed two phosphorylation sites, Thr57 and Ser130. Double mutation of these sites abolished ERK2-mediated p21 Cip1 translocation and degradation, thereby impairing ERK2-dependent cell cycle progression at the G 1 /S transition. These results indicate that ERK2 activation transduces mitogenic signals, at least in part, by downregulating the cell cycle inhibitory protein p21Cip1 .

TRIM32, which belongs to the tripartite motif (TRIM) protein family, has the RING finger, B-box, and coiled-coil domain structures common to this protein family, along with an additional NHL domain at the C terminus. TRIM32 reportedly functions as an E3 ligase for actin, a protein inhibitor of activated STAT y (PIASy), dysbindin, and c-Myc, and it has been associated with diseases such as muscular dystrophy and epithelial carcinogenesis. Here, we identify a new substrate of TRIM32 and propose a mechanism through which TRIM32 might regulate apoptosis. Our overexpression and knockdown experiments demonstrate that TRIM32 sensitizes cells to TNF␣-induced apoptosis. The RING domain is necessary for this pro-apoptotic function of TRM32 as well as being responsible for its E3 ligase activity. TRIM32 colocalizes and directly interacts with X-linked inhibitor of apoptosis (XIAP), a well known cancer therapeutic target, through its coiled-coil and NHL domains. TRIM32 overexpression enhances XIAP ubiquitination and subsequent proteasomemediated degradation, whereas TRIM32 knockdown has the opposite effect, indicating that XIAP is a substrate of TRIM32. In vitro reconstitution assay reveals that XIAP is directly ubiquitinated by TRIM32. Our novel results collectively suggest that TRIM32 sensitizes TNF␣-induced apoptosis by antagonizing XIAP, an anti-apoptotic downstream effector of TNF␣ signaling. This function may be associated with TRIM32-mediated tumor suppressive mechanism.TRIM32/HT2A is a member of the tripartite motif (TRIM) 5 protein family. To date, 72 TRIM-encoding genes have been identified in the human genome, all sharing the same overall arrangement of RING, B-box, and coiled-coil domains. Their main divergences are in the number of B-boxes and the natures of their C-terminal domains, with most TRIMs possessing one or two additional C-terminal domains, such as B30.2-like/RFP/ SPRY, NHL, ARF, PHD, and BROMO domains (1). TRIM32 has one B-box and six repeats of the NHL motif (2).Several TRIM genes have been associated with disease. For example, mutations in TRIM18/MID1, TRIM20/PYRIN, and TRIM37/MUL have been associated with X-linked Optiz G/BBB syndrome (3), familial Mediterranean fever (4), and mulibrey nanism (5), respectively. Mutations in TRIM32 were recently linked to limb girdle muscular dystrophy type 2H and sarcotubular myopathy, both of which may be caused by the same mutation (D487N) at the third NHL repeat (6). Three additional mutations in the first (R394H), fourth (T520fsX13), and fifth (D588del) NHL repeats of TRIM32 have also been found to cause limb girdle muscular dystrophy type 2H (7). Several TRIM genes have been associated with tumorigenesis; for example, TRIM19/PML (8), TRIM24/TIF1␣ (9, 10), TRIM25/EFP (11), and TRIM27/RFP (12) have been linked to tumor initiation and progression, and the level of TRIM32 mRNA was found to be up-regulated in epidermal cancers (13). However, the precise biological functions of the majority of TRIM family members have not yet been characterized.The RING domain is ...