Targeted disruption of p185/Cul7 gene results in abnormal vascular morphogenesis

Abstract

Cul1, a member of the cullin ubiquitin ligase family, forms a multiprotein complex known as SCF and plays an essential role in numerous cellular and biological activities. A Cul1 homologue, p185 (Cul7), has been isolated as an simian virus 40 large T antigen-binding protein. To understand the physiological role of p185, we generated mice lacking p185. p185 ؊/؊ embryos are runted and die immediately after birth because of respiratory distress. Dermal and hypodermal hemorrhage is detected in mutant embryos at late gestational stage. p185 ؊/؊ placentas show defects in the differentiation of the trophoblast lineage with an abnormal vascular structure. We demonstrate that p185 forms an SCF-like complex with Skp1, Rbx1, Fbw6 (Fbx29), and FAP68 (FAP48, glomulin). FAP68 has recently been identified as a gene responsible for familial glomuvenous malformation. These results suggest that p185 forms a multiprotein complex and plays an important role in vascular morphogenesis.T he rapid destruction of regulatory proteins by ubiquitinmediated proteolysis plays an important role in various biological processes. Protein ubiquitination is carried out by the sequential action of three enzymes, namely E1, E2, and E3 (1). E3 ubiquitin ligases determine the specificity of the substrate protein, and it is likely that protein ubiquitination in vivo is controlled primarily by regulating E3 activity or E3-substrate interaction (2). The Skp1-Cul1-F-box (SCF) complex is a well characterized E3 ubiquitin ligase that plays an essential role in a wide variety of activities, including cell cycle control (3, 4), muscle atrophy (5, 6), and protein quality control (7). To date, the human cullin protein family consists of six members: CUL1, CUL2, CUL3, CUL4A, CUL4B, and CUL5 (8). Although the physiological function of each cullin protein is largely unknown, mouse knockout evidence has demonstrated their biological significance. Cul1-, Cul3-, and Cul4A-deficient mice show early embryonic lethality (9-12), indicating their crucial role in early mammalian development.F box proteins are the variable subunits of the SCF complex, and a large number of human F box proteins have been reported (13,14). F box proteins contain an F box motif that binds to Skp1 and a carboxyl-terminal domain that specifies association with substrate proteins (15,16). SCF targets a number of different substrate proteins by associating with specific F box proteins (16,17). Among the human cullin proteins, SKP1 protein has been reported to bind specifically to CUL1 (18). Degradation of some F box proteins, such as Cdc4, Grr1, and SKP2 (22) in humans, are mediated by the SCF complex itself.Simian virus 40 large T antigen transforms a variety of rodent cells in vitro and in vivo. p185 (KIAA0076, p193, Cul7) was isolated as an simian virus 40 large T antigen-associated protein in murine cells (23,24). Although it has been reported that overexpression of p185 induces apoptosis in NIH 3T3 cells (24), the biological and molecular functions of p185 remain unclear. Here we demonstrate ...

A large-scale mutation search reveals genetic heterogeneity in 3M syndrome

“…12 On the other hand, in normal cells, the growth-promoting activity of CUL7 is attributed to its ability to bind p53, 12 and the increased p53 activity could partly explain the delayed growth of Cul7À/À mouse embryo fibroblasts. 17 Finally, in cells expressing T antigen, CUL7 acts as a tumour suppressor, 7,13 and T antigen may inhibit CUL7 activity by inhibiting the ubiquitination of a substrate or its binding to CUL7. 14 We have shown earlier that nonsense and missense CUL7 mutations (R1445X and H1464P) impaired the ability of CUL7 to recruit ROC1, suggesting that impaired ubiquitination may play a role in the pathogenesis of prenatal growth retardation in humans.…”

Nedd8 on cullin: building an expressway to protein destruction

“…g (Nayak et al, 2002). h (Kohrman and Imperiale, 1992;Tsai et al, 2000;Dias et al, 2002;Arai et al, 2003;Dias and Pan, unpublished results). i (Michel et al, 2003).…”

“…i (Michel et al, 2003). j (Nikolaev et al, 2003;Gu, personal communication) Role of Nedd8 in proteolysis Z-Q Pan et al CUL7 assembles an SCF-ROC1-like E3 ligase complex comprising Skp1, CUL7, the F-box protein Fbx29 and ROC1/Rbx1 (Dias et al, 2002;Arai et al, 2003). However, unlike CUL1, CUL7 specifically interacts with Skp1 Fbx29, but not with Skp1 alone nor with Skp1-bTrCP2 or Skp1-Skp2 (Dias et al, 2002).…”

“…Mouse CUL7 exhibited a potent proapoptotic activity when overexpressed in NIH 3T3 cells, and coexpression of T-ag inhibited the CUL7-mediated proapoptotic activity. Recently, work by Arai et al (2003) showed that deletion of the mouse CUL7 gene resulted in neonatal death, presumably due to abnormal vascular development. In addition, cul7 À/À mouse embryonic fibroblasts exhibited slow growth.…”

DDB1 functions as a linker to recruit receptor WD40 proteins to CUL4–ROC1 ubiquitin ligases

“…2). FBXW5 and WSB1 have been shown to bind with CUL1 and CUL2/5, respectively (Kamura et al 2001;Arai et al 2003). Whether these receptor proteins bind simultaneously with both CUL1 and CUL4 or CUL2/5 and CUL4, and whether cullin heterodimers more efficiently promote substrate polyubiquitination is an interesting possibility that is yet to be tested.…”