FBXW11 deletion alleviates Alzheimer's disease by reducing neuroinflammation and amyloid-ß plaque formation via repression of ASK1 signaling.

Alzheimer's disease (AD) is a neurodegenerative disease with a complicated pathogenesis. F-box and WD-40 domain protein 11 (FBXW11), as a component of the SCF (Skp1-Cul1-F-box) E3 ubiquitin ligase complex, regulates multiple different signaling pathways. However, the effects of FBXW11 on AD progression and the underlying mechanisms have not been studied. In this study, we found that FBXW11 expression was markedly increased in microglial cells stimulated by amyloid-ß (Aß). Immunofluorescence staining showed that FBXW11 was co-localized with Iba-1 in microglial cells, suggesting its potential in regulating neuroinflammation. Meanwhile, significantly elevated expression of FBXW11 was detected in hippocampus of AD mouse models. Then, our in vitro studies showed that FBXW11 deletion considerably ameliorated inflammatory response in Aß-incubated microglial cells through suppressing nuclear transcription factor κB (NF-κB) signaling. We further found that FBXW11 physically interacted with apoptosis signal-regulating kinase 1 (ASK1) and promoted its ubiquitination, which led to the aberrant activation of NF-κB and mitogen-activated protein kinase (MAPK) signaling pathways. Importantly, promoting ASK1 significantly abolished the effects of FBXW11 knockdown to repress inflammation and MAPKs/NF-κB activation in Aß-treated microglial cells. Subsequently, our in vivo experiments demonstrated that hippocampus-specific knockout of FBXW11 dramatically alleviated Aß plaque load, neuronal death, and microglial activation in AD mice. Furthermore, hippocampal deficiency of FBXW11 markedly mitigated neuroinflammation in AD mice through restraining ASK1/MAPKs/NF-κB signaling, along with alleviated cognitive deficits. Together, our findings demonstrated that FBXW11 may be a functionally important mediator of ASK1 activation, which could be a novel molecular target for AD treatment.

The ubiquitin ligase subunit ß-TrCP in Sertoli cells is essential for spermatogenesis in mice.

ß-TrCP is the substrate recognition subunit of an SCF-type ubiquitin ligase. We recently showed that deletion of the genes for both ß-TrCP1 and ß-TrCP2 paralogs in germ cells of male mice resulted in accumulation of the transcription factor DMRT1 and spermatogenic failure, whereas systemic ß-TrCP1 knockout combined with ß-TrCP2 knockdown had previously been shown to lead to disruption of testicular organization and accumulation of the transcription factor SNAIL. Here we investigated ß-TrCP function in Sertoli cells by generating mice with targeted deletion of the ß-TrCP2 gene in Sertoli cells on a background of whole-body ß-TrCP1 knockout. Loss of ß-TrCP in Sertoli cells caused infertility due to a reduction in the number of mature sperm. Whereas spermatogonia were not affected, male germ cells entered meiosis prematurely and the number of round spermatids was reduced in the mutant mice. Extracts of Sertoli cells and of the testis from the mutant mice manifested accumulation of SNAIL, and expression of the SNAIL target gene for E-cadherin was down-regulated in Sertoli cells from these animals. Our results indicate that ß-TrCP in Sertoli cells regulates Sertoli cell-germ cell interaction through degradation of SNAIL, with such regulation being critical for sperm development.

NSs Virulence Factor of Rift Valley Fever Virus Engages the F-Box Proteins FBXW11 and ß-TRCP1 To Degrade the Antiviral Protein Kinase PKR.

UNLABELLED: Rift Valley fever virus (RVFV, family Bunyaviridae, genus Phlebovirus) is a relevant pathogen of both humans and livestock in Africa. The nonstructural protein NSs is a major virulence factor known to suppress the type I interferon (IFN) response by inhibiting host cell transcription and by proteasomal degradation of a major antiviral IFN effector, the translation-inhibiting protein kinase PKR. Here, we identified components of the modular SCF (Skp1, Cul1, F-box protein)-type E3 ubiquitin ligases as mediators of PKR destruction by NSs. Small interfering RNAs (siRNAs) against the conserved SCF subunit Skp1 protected PKR from NSs-mediated degradation. Consequently, RVFV replication was severely reduced in Skp1-depleted cells when PKR was present. SCF complexes have a variable F-box protein subunit that determines substrate specificity for ubiquitination. We performed an siRNA screen for all (about 70) human F-box proteins and found FBXW11 to be involved in PKR degradation. The partial stabilization of PKR by FBXW11 depletion upregulated PKR autophosphorylation and phosphorylation of the PKR substrate eIF2α and caused a shutoff of host cell protein synthesis in RVFV-infected cells. To maximally protect PKR from the action of NSs, knockdown of structurally and functionally related FBXW1 (also known as ß-TRCP1), in addition to FBXW11 deletion, was necessary. Consequently, NSs was found to interact with both FBXW11 and ß-TRCP1. Thus, NSs eliminates the antiviral kinase PKR by recruitment of SCF-type E3 ubiquitin ligases containing FBXW11 and ß-TRCP1 as substrate recognition subunits. This antagonism of PKR by NSs is essential for efficient RVFV replication in mammalian cells. IMPORTANCE: Rift Valley fever virus is a pathogen of humans and animals that has the potential to spread from Africa and the Arabian Peninsula to other regions. A major virulence mechanism is the proteasomal degradation of the antiviral kinase PKR by the viral protein NSs. Here, we demonstrate that NSs requires E3 ubiquitin ligase complexes of the SCF (Skp1, Cul1, F-box protein) type to destroy PKR. SCF-type complexes can engage variant ubiquitination substrate recognition subunits, and we found the F-box proteins FBXW11 and ß-TRCP1 to be relevant for the action of NSs against PKR. Thus, we identified the host cell factors that are critically needed by Rift Valley fever virus to uphold its replication against the potent antiviral kinase PKR.

Control of chromosome stability by the beta-TrCP-REST-Mad2 axis.

REST/NRSF (repressor-element-1-silencing transcription factor/neuron-restrictive silencing factor) negatively regulates the transcription of genes containing RE1 sites. REST is expressed in non-neuronal cells and stem/progenitor neuronal cells, in which it inhibits the expression of neuron-specific genes. Overexpression of REST is frequently found in human medulloblastomas and neuroblastomas, in which it is thought to maintain the stem character of tumour cells. Neural stem cells forced to express REST and c-Myc fail to differentiate and give rise to tumours in the mouse cerebellum. Expression of a splice variant of REST that lacks the carboxy terminus has been associated with neuronal tumours and small-cell lung carcinomas, and a frameshift mutant (REST-FS), which is also truncated at the C terminus, has oncogenic properties. Here we show, by using an unbiased screen, that REST is an interactor of the F-box protein beta-TrCP. REST is degraded by means of the ubiquitin ligase SCF(beta-TrCP) during the G2 phase of the cell cycle to allow transcriptional derepression of Mad2, an essential component of the spindle assembly checkpoint. The expression in cultured cells of a stable REST mutant, which is unable to bind beta-TrCP, inhibited Mad2 expression and resulted in a phenotype analogous to that observed in Mad2(+/-) cells. In particular, we observed defects that were consistent with faulty activation of the spindle checkpoint, such as shortened mitosis, premature sister-chromatid separation, chromosome bridges and mis-segregation in anaphase, tetraploidy, and faster mitotic slippage in the presence of a spindle inhibitor. An indistinguishable phenotype was observed by expressing the oncogenic REST-FS mutant, which does not bind beta-TrCP. Thus, SCF(beta-TrCP)-dependent degradation of REST during G2 permits the optimal activation of the spindle checkpoint, and consequently it is required for the fidelity of mitosis. The high levels of REST or its truncated variants found in certain human tumours may contribute to cellular transformation by promoting genomic instability.

Silencing of both beta-TrCP1 and HOS (beta-TrCP2) is required to suppress human immunodeficiency virus type 1 Vpu-mediated CD4 down-modulation.

The human immunodeficiency virus type 1 (HIV-1) Vpu protein interacts with CD4 within the endoplasmic reticula of infected cells and targets CD4 for degradation through interaction with beta-TrCP1. Mammals possess a homologue of beta-TrCP1, HOS, which is also named beta-TrCP2. We show by coimmunoprecipitation experiments that beta-TrCP2 binds Vpu and is able to induce CD4 down-modulation as efficiently as beta-TrCP1. In two different cell lines, HeLa CD4+ and Jurkat, Vpu-mediated CD4 down-modulation could not be reversed through the individual silencing of endogenous beta-TrCP1 or beta-TrCP2 but instead required the two genes to be silenced simultaneously.

Role of F-box protein betaTrcp1 in mammary gland development and tumorigenesis.

The F-box protein betaTrcp1 controls the stability of several crucial regulators of proliferation and apoptosis, including certain inhibitors of the NF-kappaB family of transcription factors. Here we show that mammary glands of betaTrcp1(-/-) female mice display a hypoplastic phenotype, whereas no effects on cell proliferation are observed in other somatic cells. To investigate further the role of betaTrcp1 in mammary gland development, we generated transgenic mice expressing human betaTrcp1 targeted to epithelial cells under the control of the mouse mammary tumor virus (MMTV) long terminal repeat promoter. Compared to controls, MMTV betaTrcp1 mammary glands display an increase in lateral ductal branching and extensive arrays of alveolus-like protuberances. The mammary epithelia of MMTV betaTrcp1 mice proliferate more and show increased NF-kappaB DNA binding activity and higher levels of nuclear NF-kappaB p65/RelA. In addition, 38% of transgenic mice develop tumors, including mammary, ovarian, and uterine carcinomas. The targeting of betaTrcp1 to lymphoid organs produces no effects on these tissues. In summary, our results support the notion that betaTrcp1 positively controls the proliferation of breast epithelium and indicate that alteration of betaTrcp1 function and expression may contribute to malignant behavior of breast tumors, at least in part through NF-kappaB transactivation.