Recent genetic studies have documented a pivotal growth-regulatory role played by the Cullin 7 (CUL7) E3 ubiquitin ligase complex containing the Fbw8-substrate-targeting subunit, Skp1, and the ROC1 RING finger protein. In this report, we identified insulin receptor substrate 1 (IRS-1), a critical mediator of the insulin/insulin-like growth factor 1 signaling, as a proteolytic target of the CUL7 E3 ligase in a manner that depends on mammalian target of rapamycin and the p70 S6 kinase activities. Interestingly, while embryonic fibroblasts of Cul7-/- mice were found to accumulate IRS-1 and exhibit increased activation of IRS-1's downstream Akt and MEK/ERK pathways, these null cells grew poorly and displayed phenotypes reminiscent of those associated with oncogene-induced senescence. Taken together, our findings demonstrate a key role for the CUL7 E3 in targeting IRS-1 for degradation, a process that may contribute to the regulation of cellular senescence.

Intrauterine growth retardation is caused by maternal, fetal or placental factors that result in impaired endovascular trophoblast invasion and reduced placental perfusion. Although various causes of intrauterine growth retardation have been identified, most cases remain unexplained. Studying 29 families with 3-M syndrome (OMIM 273750), an autosomal recessive condition characterized by severe pre- and postnatal growth retardation, we first mapped the underlying gene to chromosome 6p21.1 and then identified 25 distinct mutations in the gene cullin 7 (CUL7). CUL7 assembles an E3 ubiquitin ligase complex containing Skp1, Fbx29 (also called Fbw8) and ROC1 and promotes ubiquitination. Using deletion analysis, we found that CUL7 uses its central region to interact with the Skp1-Fbx29 heterodimer. Functional studies indicated that the 3-M-associated CUL7 nonsense and missense mutations R1445X and H1464P, respectively, render CUL7 deficient in recruiting ROC1. These results suggest that impaired ubiquitination may have a role in the pathogenesis of intrauterine growth retardation in humans.

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Background: Negative feedback regulation of insulin signaling involves ubiquitin-dependent degradation of insulin receptor substrate 1 (IRS1). Results: Cullin-RING E3 ubiquitin ligase 7 (CRL7) mediates the ubiquitination of IRS1 in hyperphosphorylated form. Conclusion: Multisite IRS1 phosphorylation triggers interactions with CRL7 for ubiquitin modification. Significance: Insulin signaling is self-restrained when its downstream effector kinases are hyperactivated to trigger the negative feedback inhibition.

Skp2B, an F-box protein of unknown function, is frequently overexpressed in breast cancer. In order to determine the function of Skp2B and whether it has a role in breast cancer, we performed a two-hybrid screen and established transgenic mice expressing Skp2B in the mammary glands. We found that Skp2B interacts with the repressor of estrogen receptor activity (REA) and that overexpression of Skp2B leads to a reduction in REA levels. In the mammary glands of MMTV-Skp2B mice, REA levels are also low. Our results show that in virgin transgenic females, Skp2B induces lobuloalveolar development and differentiation of the mammary glands normally observed during pregnancy. As this phenotype is identical to what was observed for REA heterozygote mice, our observations suggest that the Skp2B-REA interaction is physiologically relevant. However, in contrast to REA ؉/؊ mice, MMTV-Skp2B mice develop mammary tumors, suggesting that Skp2B affects additional proteins. These results indicate that the observed expression of Skp2B in breast cancer does contribute to tumorigenesis at least in part by modulating the activity of the estrogen receptor.

Simian virus 40 (SV40) large tumor antigen (LT) triggers oncogenic transformation by inhibition of key tumor suppressor proteins, including p53 and members of the retinoblastoma family. In addition, SV40 transformation requires binding of LT to Cullin 7 (CUL7), a core component of Cullin-RING E3 ubiquitin ligase 7 (CRL7). However, the pathomechanistic effects of LT-CUL7 interaction are mostly unknown. Here we report both in vitro and in vivo experimental evidence that SV40 LT suppresses the ubiquitin ligase function of CRL7. We show that SV40 LT, but not a CUL7 binding-deficient mutant (LT Δ69-83 ), impaired 26S proteasomedependent proteolysis of the CRL7 target protein insulin receptor substrate 1 (IRS1), a component of the insulin and insulin-like growth factor 1 signaling pathway. SV40 LT expression resulted in the accumulation and prolonged half-life of IRS1. In vitro, purified SV40 LT reduced CRL7-dependent IRS1 ubiquitination in a concentration-dependent manner. Expression of SV40 LT, or depletion of CUL7 by RNA interference, resulted in the enhanced activation of IRS1 downstream signaling pathways phosphatidylinositol-3-kinase/AKT and Erk mitogen-activated pathway kinase, as well as up-regulation of the downstream target gene c-fos. Finally, SV40 LT-positive carcinoma of carcinoembryonic antigen 424/SV40 LT transgenic mice displayed elevated IRS1 protein levels and activation of downstream signaling. Taken together, these data suggest that SV40 LT protects IRS1 from CRL7-mediated degradation, thereby sustaining high levels of promitogenic IRS1 downstream signaling pathways. S tudies with simian virus 40 (SV40), a member of the Polyomaviridae family of tumor viruses, have led to fundamental insights into molecular processes of cell transformation and oncogenesis (1, 2). SV40 encodes the large tumor antigen (LT) with the potential to transform cells in culture and induce tumors in rodents. The tumorigenic features of SV40 have been attributed to binding and deactivation of key tumor suppressor proteins of the host cell including p53 and members of the retinoblastoma (pRB) family (1-3). In addition, SV40 LT was shown to be physically associated with Cullin 7 (CUL7; also named p185 or p193) (4, 5) as well as insulin receptor substrate 1 (IRS1) (6). It has been proposed that the association of SV40 LT with either CUL7 or IRS1 is critical to SV40 oncogenic transformation (7-9). However, the functional effect of LT interaction with CUL7/IRS1 and their pathophysiological interrelation remains mostly unknown.CUL7 is a scaffold protein responsible for assembling the multisubunit Cullin-RING E3 ubiquitin ligase 7 (CRL7) that consists of the RING-finger protein ROC1 and the Skp1-Fbw8 substrate-targeting subunit (10, 11). Genetic studies documented a pivotal growth-regulatory role of CRL7. Both cul7 (12) and fbw8 (13) null mice exhibit intrauterine growth retardation. In addition, CUL7 germ-line mutations were linked to 3-M syndrome, a hereditary disorder characterized by pre-and postnatal growth retardation in hum...

Convolutional neural networks (CNN) have been widely used in the field of synthetic aperture radar (SAR) image classification for their high classification accuracy. However, because CNNs learn a fairly discontinuous input-output mapping, they are vulnerable to adversarial examples. Unlike most existing attack manners that fool CNN models with complex global perturbations, this study provides an idea for generating more dexterous adversarial perturbations. It demonstrates that minor local perturbations are also effective for attacking. We propose a new attack method called local aggregative attack (LAA), which is a black-box method based on probability label information, to reduce the range and amplitude of adversarial perturbations. Our attack introduces the differential evolution (DE) algorithm to search for the optimal perturbations and applies the maximum between-class variance method (OTSU algorithm) to accomplish pixel-level labelling of the target and background areas, enabling attackers to generate adversarial examples of SAR images (AESIs) by adding small-scale perturbations to specific areas. Meanwhile, the structural dissimilarity (DSSIM) metric optimises the cost function to limit image distortion and improve attack stealthiness. Experiments show that our method achieves a high attack success rate against these CNN-based classifiers, and the generated AESIs are equipped with non-negligible transferability between different models.