F-Box Only Protein 31 (FBXO31) Negatively Regulates p38 Mitogen-activated Protein Kinase (MAPK) Signaling by Mediating Lysine 48-linked Ubiquitination and Degradation of Mitogen-activated Protein Kinase Kinase 6 (MKK6)

The p38 MAPK signal transduction pathway plays an important role in inflammatory and stress responses. MAPKK6 (MKK6), a dual specificity protein kinase, is a p38 activator. Activation of the MKK6-p38 pathway is kept in check by multiple layers of regulations, including autoinhibition, dimerization, scaffold proteins, and Lys-63-linked polyubiquitination. However, the mechanisms underlying deactivation of MKK6-p38, which is crucial for maintaining the magnitude and duration of signal transduction, are not well understood. Lys-48-linked ubiquitination, which marks substrates for proteasomal degradation, is an important negative posttranslational regulatory machinery for signal pathway transduction. Here we report that the accumulation of F-box only protein 31 (FBXO31), a component of Skp1·Cul1·F-box protein E3 ligase, negatively regulated p38 activation in cancer cells upon genotoxic stresses. Our results show that FBXO31 binds to MKK6 and mediates its Lys-48-linked polyubiquitination and degradation, thereby functioning as a negative regulator of MKK6-p38 signaling and protecting cells from stress-induced cell apoptosis. Taken together, our findings uncover a new mechanism of deactivation of MKK6-p38 and substantiate a novel regulatory role of FBXO31 in stress response.     

去乙酰化转移酶SIRT7的作用及机制研究进展

SIRT7是哺乳动物Sirtuins家族中的一员,定位于核仁,是一种高度特异性的H3K18Ac(组蛋白H3的乙酰化18位赖氨酸残基)去乙酰化酶。近年来的研究发现SIRT7可通过多种途径参与调控核糖体RNA转录、细胞代谢、细胞应激以及DNA损伤修复等生理过程。此外,SIRT7还与衰老、心脏疾病及脂肪肝等密切相关。特别是SIRT7在多种肿瘤如肝癌、胃癌、乳腺癌、膀胱癌、结直肠癌、胰腺癌和头颈鳞状细胞癌等发生发展中起着重要的调节作用。文中综述了SIRT7的细胞及分子生物学作用,并系统总结了其在人类疾病中的研究现状。     

FBXO7 Y52C Polymorphism as a Potential Protective Factor in Parkinson's Disease

Mutations in the F-box only protein 7 gene (FBXO7), the substrate-specifying subunit of SCF E3 ubiquitin ligase complex, cause Parkinson''s disease (PD)-15 (PARK15). To identify new variants, we sequenced FBXO7 cDNA in 80 Taiwanese early onset PD patients (age at onset ≤50) and only two known variants, Y52C (c.155A>G) and M115I (c.345G>A), were found. To assess the association of Y52C and M115I with the risk of PD, we conducted a case–control study in a cohort of PD and ethnically matched controls. There was a nominal difference in the Y52C G allele frequency between PD and controls (p = 0.045). After combining data from China [1], significant difference in the Y52C G allele frequency between PD and controls (p = 0.012) and significant association of G allele with decreased PD risk (p = 0.017) can be demonstrated. Upon expressing EGFP-tagged Cys52 FBXO7 in cells, a significantly reduced rate of FBXO7 protein decay was observed when compared with cells expressing Tyr52 FBXO7. In silico modeling of Cys52 exhibited a more stable feature than Tyr52. In cells expressing Cys52 FBXO7, the level of TNF receptor-associated factor 2 (TRAF2) was significantly reduced. Moreover, Cys52 FBXO7 showed stronger interaction with TRAF2 and promoted TRAF2 ubiquitination, which may be responsible for the reduced TRAF2 expression in Cys52 cells. After induced differentiation, SH-SY5Y cells expressing Cys52 FBXO7 displayed increased neuronal outgrowth. We therefore hypothesize that Cys52 variant of FBXO7 may contribute to reduced PD susceptibility in Chinese.     

The Rice NAD+-Dependent Histone Deacetylase OsSRT1 Targets Preferentially to Stress- and Metabolism-Related Genes and Transposable Elements

Histone acetylation/deacetylation is an important chromatin modification for epigenetic regulation of gene expression. Silent information regulation2 (Sir2)-related sirtuins are nicotinamide-adenine dinucleotide (NAD⁺)-dependent histone deacetylases (HDAC). The mammalian sirtuin family comprises 7 members (SIRT1-7) that act in different cellular compartments to regulate metabolism and aging. The rice genome contains only two Sir2-related genes: OsSRT1 (or SRT701) and OsSRT2 (orSRT702). OsSRT1 is closely related to the mammalian SIRT6, while OsSRT2 is homologous to SIRT4. Previous work has shown that OsSRT1 is required for the safeguard against genome instability and cell damage in rice plant. In this work we investigated the role of OsSRT1 on genome-wide acetylation of histone H3 lysine 9 (H3K9ac) and studied the genome-wide binding targets of OsSRT1. The study reveals that OsSRT1 binds to loci with relatively low levels of H3K9ac and directly regulates H3K9ac and expression of many genes that are related to stress and metabolism, indicating that OsSRT1 is an important site-specific histone deacetylase for gene regulation in rice. In addition, OsSRT1 is found to also target to several families of transposable elements, suggesting that OsSRT1 is directly involved in transposable element repression.     

Emerging roles of SIRT6 on telomere maintenance,DNA repair,metabolism and mammalian aging

With the characterization of Sir2 gene in yeast aging, its mammalian homologs Sirtuins 1–7 have been attracting attention from scientists with various research backgrounds. Among Sirtuins, SIRT1 is the most extensively studied. Recent progress on mammalian Sirtuins has shown that SIRT6 as a histone deacetylase may also play a critical role in regulating mammalian aging. This review summarizes recent advances on SIRT6 as a key modulator of telomere structure, DNA repair, metabolism, and NF-kappa B pathway in aging. In addition, we discuss the challenges that remain to be studied in SIRT6 biology.     

Loss of FBXO7 (PARK15) results in reduced proteasome activity and models a parkinsonism‐like phenotype in mice

Mutations in the FBXO7 (PARK15) gene have been implicated in a juvenile form of parkinsonism termed parkinsonian pyramidal syndrome (PPS), characterized by Parkinsonian symptoms and pyramidal tract signs. FBXO7 (F‐box protein only 7) is a subunit of the SCF (SKP1/cullin‐1/F‐box protein) E3 ubiquitin ligase complex, but its relevance and function in neurons remain to be elucidated. Here, we report that the E3 ligase FBXO7‐SCF binds to and ubiquitinates the proteasomal subunit PSMA2. In addition, we show that FBXO7 is a proteasome‐associated protein involved in proteasome assembly. In FBXO7 knockout mice, we find reduced proteasome activity and early‐onset motor deficits together with premature death. In addition, we demonstrate that NEX (neuronal helix–loop–helix protein‐1)‐Cre‐induced deletion of the FBXO7 gene in forebrain neurons or the loss of FBXO7 in tyrosine hydroxylase (TH)‐positive neurons results in motor defects, reminiscent of the phenotype in PARK15 patients. Taken together, our study establishes a vital role for FBXO7 in neurons, which is required for proper motor control and accentuates the importance of FBXO7 in proteasome function.     

SIRT1 inhibits transforming growth factor beta-induced apoptosis in glomerular mesangial cells via Smad7 deacetylation

SIRT1, a class III histone deacetylase, is considered a key regulator of cell survival and apoptosis through its interaction with nuclear proteins. In this study, we have examined the likelihood and role of the interaction between SIRT1 and Smad7, which mediates transforming growth factor beta (TGFbeta)-induced apoptosis in renal glomerular mesangial cells. Immunoprecipitation analysis revealed that SIRT1 directly interacts with the N terminus of Smad7. Furthermore, SIRT1 reversed acetyl-transferase (p300)-mediated acetylation of two lysine residues (Lys-64 and -70) on Smad7. In mesangial cells, the Smad7 expression level was reduced by SIRT1 overexpression and increased by SIRT1 knockdown. SIRT1-mediated deacetylation of Smad7 enhanced Smad ubiquitination regulatory factor 1 (Smurf1)-mediated ubiquitin proteasome degradation, which contributed to the low expression of Smad7 in SIRT1-overexpressing mesangial cells. Stimulation by TGFbeta or overexpression of Smad7 induced mesangial cell apoptosis, as assessed by morphological apoptotic changes (nuclear condensation) and biological apoptotic markers (cleavages of caspase3 and poly(ADP-ribose) polymerase). However, TGFbeta failed to induce apoptosis in Smad7 knockdown mesangial cells, indicating that Smad7 mainly mediates TGFbeta-induced apoptosis of mesangial cells. Finally, SIRT1 overexpression attenuated both Smad7- and TGFbeta-induced mesangial cell apoptosis, whereas SIRT1 knockdown enhanced this apoptosis. We have concluded that Smad7 is a new target molecule for SIRT1 and SIRT1 attenuates TGFbeta-induced mesangial cell apoptosis through acceleration of Smad7 degradation. Our results suggest that up-regulation of SIRT1 deacetylase activity is a potentially useful therapeutic strategy for prevention of TGFbeta-related kidney disease through its effect on cell survival.     

TRAF7 sequesters c-Myb to the cytoplasm by stimulating its sumoylation

Small ubiquitin-related modifiers (SUMOs) are proteins that are posttranslationally conjugated to diverse proteins. The c-myb proto-oncogene product (c-Myb) regulates proliferation and differentiation of hematopoietic cells. PIASy is the only known SUMO E3 ligase for c-Myb. Here, we report that TRAF7 binds to c-Myb and stimulates its sumoylation. TRAF7 bound to the DNA-binding domain of c-Myb via its WD40 repeats. TRAF7 has an E3 ubiquitin ligase activity for self-ubiquitination, but TRAF7 also stimulated the sumoylation of c-Myb at Lys-523 and Lys-499, which are the same sites as those used for PIASy-induced sumoylation. TRAF7 inhibited trans-activation induced by wild-type c-Myb, but not by the sumoylation site mutant of c-Myb. The expression of both c-myb and TRAF7 was down-regulated during differentiation of M1 cells. Endogenous TRAF7 localized to both the cytoplasm and nucleus of M1 cells. Consistent with this, significant amounts of sumoylated c-Myb were found in the cytoplasm of M1 cells, whereas nonsumoylated c-Myb was found predominantly in the nucleus. Overexpressed TRAF7 was localized in the cytoplasm of CV-1 cells, and sequestered c-Myb and SUMO1 in the cytosol, whereas PIASy was localized in the nucleus. Thus, TRAF7 negatively regulates c-Myb activity by sequestering c-Myb to the cytosol via sumoylation.     

HDAC6 mediates the acetylation of TRIM50

The E3 Ubiquitin ligase TRIM50 promotes the formation and clearance of aggresome-associated polyubiquitinated proteins through HDAC6 interaction, a tubulin specific deacetylase that regulates microtubule-dependent aggresome formation. In this report we showed that TRIM50 is a target of HDAC6 with Lys-372 as a critical residue for acetylation. We identified p300 and PCAF as two TRIM50 acetyltransferases and we further showed that a balance between ubiquitination and acetylation regulates TRIM50 degradation.     

SCFFBXO28-mediated self-ubiquitination of FBXO28 promotes its degradation

The F-box protein is the substrate recognition subunit of SCF (SKP1/CUL1/F-box) E3 ubiquitin ligase complex, a multicomponent RING-type E3 ligase involved in the regulation of numerous cellular processes by targeting critical regulatory proteins for ubiquitination. However, whether and how F-box proteins are regulated is largely unknown. Here we report that FBXO28, a poorly characterized F-box protein, is a novel substrate of SCF E3 ligase. Pharmaceutical or genetic inhibition of neddylation pathway that is required for the activation of SCF stabilizes FBXO28 and prolongs its half-life. Meanwhile, FBXO28 is subjected to ubiquitination and cullin1-based SCF complex promotes FBXO28 degradation. Moreover, deletion of F-box domain stabilizes FBXO28 and knockdown of endogenous FBXO28 strongly upregulates exogenous FBXO28 expression. Taken together, these data reveal that SCF^FBXO28 is the E3 ligase responsible for the self-ubiquitination and proteasomal degradation of FBXO28, providing a new clue for the upstream signaling regulation for F-box proteins.     

The Proapoptotic F-box Protein Fbxl7 Regulates Mitochondrial Function by Mediating the Ubiquitylation and Proteasomal Degradation of Survivin

Fbxl7, a component of the Skp1·Cul1·F-box protein type ubiquitin E3 ligase, regulates mitotic cell cycle progression. Here we demonstrate that overexpression of Fbxl7 in lung epithelia decreases the protein abundance of survivin, a member of the inhibitor of apoptosis family. Fbxl7 mediates polyubiquitylation and proteasomal degradation of survivin by interacting with Glu-126 within its carboxyl-terminal α helix. Furthermore, both Lys-90 and Lys-91 within survivin serve as ubiquitin acceptor sites. Ectopically expressed Fbxl7 impairs mitochondrial function, whereas depletion of Fbxl7 protects mitochondria from actions of carbonyl cyanide m-chlorophenylhydrazone, an inhibitor of oxidative phosphorylation. Compared with wild-type survivin, cellular expression of a survivin mutant protein deficient in its ability to interact with Fbxl7 (E126A) and a ubiquitylation-resistant double point mutant (KK90RR/KK91RR) rescued mitochondria to a larger extent from damage induced by overexpression of Fbxl7. Therefore, these data suggest that the Skp1·Cul1·F-box protein complex subunit Fbxl7 modulates mitochondrial function by controlling the cellular abundance of survivin. The results raise opportunities for F-box protein targeting to preserve mitochondrial function.     

SIRT2 reduces actin polymerization and cell migration through deacetylation and degradation of HSP90

SIRT2, a member of the class III histone deacetylase family, has been identified as a tumor suppressor, which is associated with various cellular processes including metabolism and proliferation. However, the effects of SIRT2 on cancer cell migration caused by cytoskeletal rearrangement remain uncertain. Here we show that SIRT2 inhibits cell motility by suppressing actin polymerization. SIRT2 regulates actin dynamics through HSP90 destabilization and subsequent repression of LIM kinase (LIMK) 1/cofilin pathway. SIRT2 directly interacts with HSP90 and regulates its acetylation and ubiquitination. In addition, the deacetylase activity of SIRT2 is required for the regulation of actin polymerization and the ubiquitin-mediated proteasomal degradation of HSP90 induced by SIRT2.     

Lipopolysaccharide Primes the NALP3 Inflammasome by Inhibiting Its Ubiquitination and Degradation Mediated by the SCFFBXL2 E3 Ligase

The inflammasome is a multiprotein complex that augments the proinflammatory response by increasing the generation and cellular release of key cytokines. Specifically, the NALP3 inflammasome requires two-step signaling, priming and activation, to be functional to release the proinflammatory cytokines IL-1β and IL-18. The priming process, through unknown mechanisms, increases the protein levels of NALP3 and pro-IL-1β in cells. Here we show that LPS increases the NALP3 protein lifespan without significantly altering steady-state mRNA in human cells. LPS exposure reduces the ubiquitin-mediated proteasomal processing of NALP3 by inducing levels of an E3 ligase component, FBXO3, which targets FBXL2. The latter is an endogenous mediator of NALP3 degradation. FBXL2 recognizes Trp-73 within NALP3 for interaction and targets Lys-689 within NALP3 for ubiquitin ligation and degradation. A unique small molecule inhibitor of FBXO3 restores FBXL2 levels, resulting in decreased NALP3 protein levels in cells and, thereby, reducing the release of IL-1β and IL-18 in human inflammatory cells after NALP3 activation. Our findings uncover NALP3 as a molecular target for FBXL2 and suggest that therapeutic targeting of the inflammasome may serve as a platform for preclinical intervention.