DNA Damage Response Regulation by Histone Ubiquitination.

Cells are constantly exposed to numerous genotoxic stresses that induce DNA damage. DNA double-strand breaks (DSBs) are among the most serious damages and should be systematically repaired to preserve genomic integrity. The efficiency of repair is closely associated with chromatin structure, which is regulated by posttranslational modifications of histones, including ubiquitination.

Retusone A, a Guaiane-Type Sesquiterpene Dimer from and Its Inhibitory Effects on Histone Acetyltransferase HBO1 Expression.

Retusone A (), a new sesquiterpene dimer consisting of two guaiane-type sesquiterpenoids, and oleodaphnal () were isolated from heartwood of (Thymelaeaceae). The planar structure of was elucidated on the basis of HRESIMS and NMR spectroscopic data, and the relative stereochemistry was established by X-ray diffraction analysis. The absolute configuration of was determined by electronic circular dichroism.

Regulation of Hypoxic Signaling and Oxidative Stress via the MicroRNA-SIRT2 Axis and Its Relationship with Aging-Related Diseases.

The sirtuin family of nicotinamide adenine dinucleotide-dependent deacetylase and ADP-ribosyl transferases plays key roles in aging, metabolism, stress response, and aging-related diseases. SIRT2 is a unique sirtuin that is expressed in the cytosol and is abundant in neuronal cells. Various microRNAs were recently reported to regulate SIRT2 expression via its 3'-untranslated region (UTR), and single nucleotide polymorphisms in the miRNA-binding sites of SIRT2 3'-UTR were identified in patients with neurodegenerative diseases.

SIRT2 inhibition activates hypoxia-inducible factor 1α signaling and mediates neuronal survival.

Sirtuins are deacetylases dependent on nicotine adenine dinucleotide (NAD) and take an important role in metabolism and aging. In mammals, there are seven sirtuins (SlRTl-7), and only SIRT2 is predominantly localized in cytoplasm. Under hypoxic environments, metazoan organisms must maintain oxygen homeostasis to survive.

MITOL dysfunction causes dwarfism with anterior pituitary hypoplasia.

In mitochondrial disorders, short stature and growth failure are common symptoms, but their underlying mechanism remains unknown. In this study, we examined the cause of growth failure of mice induced by nestin promoter-driven knockout of the mitochondrial ubiquitin ligase MITOL (MARCH5), a key regulator of mitochondrial function. MITOL-knockout mice have congenital hypoplasia of the anterior pituitary caused by decreased expression of pituitary transcript factor 1 (Pit1).

Mitochondrial Dynamics Regulation in Skin Fibroblasts from Mitochondrial Disease Patients.

Mitochondria are highly dynamic organelles that constantly fuse, divide, and move, and their function is regulated and maintained by their morphologic changes. Mitochondrial disease (MD) comprises a group of disorders involving mitochondrial dysfunction. However, it is not clear whether changes in mitochondrial morphology are related to MD.

Critical role of CRAG, a splicing variant of centaurin-γ3/AGAP3, in ELK1-dependent SRF activation at PML bodies.

CRMP-5-associated GTPase (CRAG), a short splicing variant of centaurin-γ3/AGAP3, is predominantly expressed in the developing brain. We previously demonstrated that CRAG, but not centaurin-γ3, translocates to the nucleus and activates the serum response factor (SRF)-c-Fos pathway in cultured neuronal cells. However, the physiological relevance of CRAG in vivo is unknown.

MITOL deletion in the brain impairs mitochondrial structure and ER tethering leading to oxidative stress.

Mitochondrial abnormalities are associated with developmental disorders, although a causal relationship remains largely unknown. Here, we report that increased oxidative stress in neurons by deletion of mitochondrial ubiquitin ligase MITOL causes a potential neuroinflammation including aberrant astrogliosis and microglial activation, indicating that mitochondrial abnormalities might confer a risk for inflammatory diseases in brain such as psychiatric disorders. A role of MITOL in both mitochondrial dynamics and ER-mitochondria tethering prompted us to characterize three-dimensional structures of mitochondria in vivo.

MITOL prevents ER stress-induced apoptosis by IRE1α ubiquitylation at ER-mitochondria contact sites.

Unresolved endoplasmic reticulum (ER) stress shifts the unfolded protein response signaling from cell survival to cell death, although the switching mechanism remains unclear. Here, we report that mitochondrial ubiquitin ligase (MITOL/MARCH5) inhibits ER stress-induced apoptosis through ubiquitylation of IRE1α at the mitochondria-associated ER membrane (MAM). MITOL promotes K63-linked chain ubiquitination of IRE1α at lysine 481 (K481), thereby preventing hyper-oligomerization of IRE1α and regulated IRE1α-dependent decay (RIDD).

Induction of Short NFATc1/αA Isoform Interferes with Peripheral B Cell Differentiation.

In lymphocytes, immune receptor signals induce the rapid nuclear translocation of preformed cytosolic NFAT proteins. Along with co-stimulatory signals, persistent immune receptor signals lead to high levels of NFATc1/αA, a short NFATc1 isoform, in effector lymphocytes. Whereas NFATc1 is not expressed in plasma cells, in germinal centers numerous centrocytic B cells express nuclear NFATc1/αA.

Regulation of B cell differentiation by the ubiquitin-binding protein TAX1BP1.

Tax1-binding protein 1 (TAX1BP1) is a ubiquitin-binding protein that restricts nuclear factor-κB (NF-κB) activation and facilitates the termination of aberrant inflammation. However, its roles in B-cell activation and differentiation are poorly understood. To evaluate the function of TAX1BP1 in B cells, we established TAX1BP1-deficient DT40 B cells that are hyper-responsive to CD40-induced extracellular signal-regulated kinase (ERK) activation signaling, exhibit prolonged and exaggerated ERK phosphorylation and show enhanced B lymphocyte-induced maturation protein 1 (Blimp-1; a transcription factor inducing plasma cell differentiation) expression that is ERK-dependent.

Modularized functions of the Fanconi anemia core complex.

The Fanconi anemia (FA) core complex provides the essential E3 ligase function for spatially defined FANCD2 ubiquitination and FA pathway activation. Of the seven FA gene products forming the core complex, FANCL possesses a RING domain with demonstrated E3 ligase activity. The other six components do not have clearly defined roles.

MITOL regulates endoplasmic reticulum-mitochondria contacts via Mitofusin2.

The mitochondrial ubiquitin ligase MITOL regulates mitochondrial dynamics. We report here that MITOL regulates mitochondria-associated endoplasmic reticulum (ER) membrane (MAM) domain formation through mitofusin2 (Mfn2). MITOL interacts with and ubiquitinates mitochondrial Mfn2, but not ER-associated Mfn2.

Direct inhibition of TNF-α promoter activity by Fanconi anemia protein FANCD2.

Fanconi anemia (FA), an inherited disease, is associated with progressive bone marrow failure, predisposition to cancer, and genomic instability. Genes corresponding to 15 identified FA complementation groups have been cloned, and each gene product functions in the response to DNA damage induced by cross-linking agents and/or in protection against genome instability. Interestingly, overproduction of inflammatory cytokines such as tumor necrosis factor alpha (TNF-α) and aberrant activation of NF-κB-dependent transcriptional activity have been observed in FA cells.

Distinct regulation of mitochondrial localization and stability of two human Sirt5 isoforms.

Seven human Sir2 homologues (sirtuin) have been identified to date. In this study, we clarified the mechanism of subcellular localization of two SIRT5 isoforms (i.e.

A mitochondrial ubiquitin ligase MITOL controls cell toxicity of polyglutamine-expanded protein.

Expansion of a polyglutamine tract in ataxin-3 (polyQ) causes Machado-Joseph disease, a late-onset neurodegenerative disorder characterized by ubiquitin-positive aggregate formation. Several lines of evidence demonstrate that polyQ also accumulates in mitochondria and causes mitochondrial dysfunction. To uncover the mechanism of mitochondrial quality-control via the ubiquitin-proteasome pathway, we investigated whether MITOL, a novel mitochondrial ubiquitin ligase localized in the mitochondrial outer membrane, is involved in the degradation of pathogenic ataxin-3 in mitochondria.

Mitochondrial ubiquitin ligase MITOL ubiquitinates mutant SOD1 and attenuates mutant SOD1-induced reactive oxygen species generation.

We have previously identified a novel mitochondrial ubiquitin ligase, MITOL, which is localized in the mitochondrial outer membrane and is involved in the control of mitochondrial dynamics. In this study, we examined whether MITOL eliminates misfolded proteins localized to mitochondria. Mutant superoxide dismutase1 (mSOD1), one of misfolded proteins, has been shown to localize in mitochondria and induce mitochondrial dysfunction, possibly involving in the onset and progression of amyotrophic lateral sclerosis.

The Fanconi anemia pathway promotes homologous recombination repair in DT40 cell line.

Fanconi anemia (FA) is a rare hereditary disorder characterized by bone marrow failure, compromised genome stability, and increased incidence of cancer. FA is caused by abnormalities that occur in components of the FA core complex, a key factor FancD2, breast cancer susceptibility protein BRCA2/FancD1, or BRIP1/FancJ. These proteins are proposed to function in a common biochemical process (FA pathway), however, its precise role is still unclear.

A requirement of FancL and FancD2 monoubiquitination in DNA repair.

The rare hereditary disorder Fanconi anemia (FA) can be caused by mutations in components of the FA core complex (FancA/B/C/E/F/G/L/M), a key regulator FancD2, the breast cancer susceptibility protein BRCA2/FancD1, or the newly identified FancJ/BRIP1 helicase. By performing yeast two-hybrid (Y2H) screens using N-terminal chicken (ch) FancD2 as a bait, we have identified chFancL, the likely ubiquitin E3 ligase subunit of the FA core complex. We also found that ectopically expressed FancD2 and FancL co-immunoprecipitated in 293T cells, and this interaction was dependent on the PHD domain of FancL.

Functional interplay between BRCA2/FancD1 and FancC in DNA repair.

A rare hereditary disorder, Fanconi anemia (FA), is caused by mutations in an array of genes, which interact in a common FA pathway/network. These genes encode components of the FA "core" complex, a key factor FancD2, the familial breast cancer suppressor BRCA2/FancD1, and Brip1/FancJ helicase. Although BRCA2 is known to play a pivotal role in homologous recombination repair by regulating Rad51 recombinase, the precise functional relationship between BRCA2 and the other FA genes is unclear.

A FancD2-monoubiquitin fusion reveals hidden functions of Fanconi anemia core complex in DNA repair.

In DNA damage responses, the Fanconi anemia (FA) protein, FancD2, is targeted to chromatin and forms nuclear foci following its monoubiquitination, a process likely catalyzed by the FA core complex. Here, we show that a chicken FancD2-ubiquitin fusion protein, carrying a Lys-Arg substitution removing the natural monoubiquitination site (D2KR-Ub), could reverse cisplatin hypersensitivity and localize to chromatin in FANCD2-deficient DT40 cells. Importantly, the chromatin targeting was dependent on three core complex components as well as the hydrophobic surface of ubiquitin that may direct protein-protein interactions.

Role of NAD-dependent deacetylases SIRT1 and SIRT2 in radiation and cisplatin-induced cell death in vertebrate cells.

Yeast Sir2 is a nicotinamide adenine dinucleotide (NAD)-dependent histone deacetylase that plays a central role in transcriptional silencing, chromosomal stability, DNA damage response and aging. In mammals, Sir2-like genes constitute a seven-member family whose function is largely unknown. To investigate the role of the Sir2 family in vertebrates, we have disrupted Sir2 homologues SIRT1 and SIRT2 in the p53-deficient chicken cell line DT40.

Functional relationships of FANCC to homologous recombination, translesion synthesis, and BLM.

Some of the restarting events of stalled replication forks lead to sister chromatid exchange (SCE) as a result of homologous recombination (HR) repair with crossing over. The rate of SCE is elevated by the loss of BLM helicase or by a defect in translesion synthesis (TLS). We found that spontaneous SCE levels were elevated approximately 2-fold in chicken DT40 cells deficient in Fanconi anemia (FA) gene FANCC.

Fanconi anemia protein FANCD2 promotes immunoglobulin gene conversion and DNA repair through a mechanism related to homologous recombination.

Recent studies show overlap between Fanconi anemia (FA) proteins and those involved in DNA repair mediated by homologous recombination (HR). However, the mechanism by which FA proteins affect HR is unclear. FA proteins (FancA/C/E/F/G/L) form a multiprotein complex, which is responsible for DNA damage-induced FancD2 monoubiquitination, a key event for cellular resistance to DNA damage.