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A decade has passed since the first reported connection between RAP80 and BRCA1 in DNA double-strand break repair. Despite the initial identification of RAP80 as a factor localizing BRCA1 to DNA double-strand breaks and potentially promoting homologous recombination, there is increasing evidence that RAP80 instead suppresses homologous recombination to fine-tune the balance of competing DNA repair processes during the S/G phase of the cell cycle. RAP80 opposes homologous recombination by inhibiting DNA end-resection and sequestering BRCA1 into the BRCA1-A complex. Ubiquitin and SUMO modifications of chromatin at DNA double-strand breaks recruit RAP80, which contains distinct sequence motifs that recognize ubiquitin and SUMO. Here, we review RAP80's role in repressing homologous recombination at DNA double-strand breaks and how this role is facilitated by its ability to bind ubiquitin and SUMO modifications. [ABSTRACT FROM AUTHOR]

The hexanucleotide repeat expansion (HRE) GGGGCC (G4C2) in C9orf72 is the most common cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Recent studies support an HRE RNA gain-of-function mechanism of neurotoxicity, and we previously identified protein interactors for the G4C2 RNA including RanGAP1. A candidate-based genetic screen in Drosophila expressing 30 G4C2 repeats identified RanGAP (Drosophila orthologue of human RanGAP1), a key regulator of nucleocytoplasmic transport, as a potent suppressor of neurodegeneration. Enhancing nuclear import or suppressing nuclear export of proteins also suppresses neurodegeneration. RanGAP physically interacts with HRE RNA and is mislocalized in HRE-expressing flies, neurons from C9orf72 ALS patient-derived induced pluripotent stem cells (iPSC-derived neurons), and in C9orf72 ALS patient brain tissue. Nuclear import is impaired as a result of HRE expression in the fly model and in C9orf72 iPSC-derived neurons, and these deficits are rescued by small molecules and antisense oligonucleotides targeting the HRE G-quadruplexes. Nucleocytoplasmic transport defects may be a fundamental pathway for ALS and FTD that is amenable to pharmacotherapeutic intervention. [ABSTRACT FROM AUTHOR]

OTUB1 is a Lys48-specific deubiquitinating enzyme that forms a complex in vivo with E2 ubiquitin (Ub)-conjugating enzymes including UBC13 and UBCH5. OTUB1 binds E2~Ub thioester intermediates and prevents ubiquitin transfer, thereby noncatalytically inhibiting accumulation of polyubiquitin. We report here that a second role of OTUB1-E2 interactions is to stimulate OTUB1 cleavage of Lys48 polyubiquitin. This stimulation is regulated by the ratio of charged to uncharged E2 and by the concentration of Lys48-linked polyubiquitin and free ubiquitin. Structural and biochemical studies of human and worm OTUB1 and UBCH5B show that the E2 enzyme stimulates binding of the Lys48 polyubiquitin substrate by stabilizing folding of the OTUB1 N-terminal ubiquitin-binding helix. Our results suggest that OTUB1-E2 complexes in the cell are poised to regulate polyubiquitin chain elongation or degradation in response to changing levels of E2 charging and available free ubiquitin. [ABSTRACT FROM AUTHOR]

Tandem mass spectrometry (MS/MS) allows for the rapid identification of many types of post-translational modifications (PTMs), especially those that can be detected by a diagnostic mass shift in one or more peptide fragment ions (for example, phosphorylation). But some PTMs (for example, SUMOs and other ubiquitin-like modifiers) themselves produce multiple fragment ions; combined with fragments from the modified target peptide, a complex overlapping fragmentation pattern is thus generated, which is uninterpretable by standard peptide sequencing software. Here we introduce SUMmOn, an automated pattern recognition tool that detects diagnostic PTM fragment ion series within complex MS/MS spectra, to identify modified peptides and modification sites within these peptides. Using SUMmOn, we demonstrate for the first time that human SUMO-1 multimerizes in vitro primarily via three N-terminal lysines, Lys7, Lys16 and Lys17. Notably, our method is theoretically applicable to any type of modification or chemical moiety generating a unique fragment ion pattern. [ABSTRACT FROM AUTHOR]

Germ cells must develop along distinct male or female paths to produce the sperm or eggs required for sexual reproduction. In both mouse and Drosophila, the sexual identity of germ cells is influenced by the sex of the surrounding somatic tissue (for example, refs 1, 2, reviewed in refs 3, 4); however, little is known about how the soma controls germline sex determination. Here we show that the janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway provides a sex-specific signal from the soma to the germ line in Drosophila embryonic gonads. The somatic gonad expresses a JAK/STAT ligand, unpaired (upd), in a male-specific manner, and activates the JAK/STAT pathway in male germ cells at the time of gonad formation. Furthermore, the JAK/STAT pathway is necessary for male-specific germ cell behaviour during early gonad development, and is sufficient to activate aspects of male germ cell behaviour in female germ cells. Our findings provide direct evidence that the JAK/STAT pathway mediates a key signal from the somatic gonad that regulates male germline sexual development. [ABSTRACT FROM AUTHOR]

The XY body is a specialized chromatin territory that forms during meiotic prophase of spermatogenesis and comprises the transcriptionally repressed sex chromosomes. Remodeling of the XY chromatin is brought about by recruitment of specific proteins to the X and Y chromosomes during meiosis, and also by post-translational modifications of histones and other chromatin-associated proteins. Here, we demonstrate that SUMO, a small ubiquitin-related modifier protein that regulates a wide variety of nuclear functions in somatic cells, dramatically localizes to the XY body. SUMO was first detected in the XY body of early pachytene spermatocytes and gradually accumulated, reaching maximal levels there during the mid to late pachytene stages. Several known SUMO substrates, including PML and DAXX, were also found to accumulate in the XY body of mid to late stage pachytene spermatocytes. These same proteins localize to PML nuclear bodies of somatic interphase nuclei. Together, these findings indicate a role for SUMO modification in regulating the structure and function of the XY body and reveal molecular similarities between the XY body and PML nuclear bodies. [ABSTRACT FROM AUTHOR]

The L1 family of long interspersed repetitive DNA in the rabbit genome (L1Oc) has been studied by determining the sequence of the five L1 repeats in the rabbit β-like globin gene cluster and by hybridization analysis of other L1 repeats in the genome. L1Oc repeats have a common 3′ end that terminates in a poly A addition signal and an A-rich tract, but individual repeats have different 5′ ends, indicating a polar truncation from the 5′ end during their synthesis or propagation. As a result of the polar truncations, the 5′ end of L1Oc is present in about 11,000 copies per haploid genome, whereas the 3′ end is present in at least 66,000 copies per haploid genome. One type of L1Oc repeat has internal direct repeats of 78 bp in the 3′ untranslated region, whereas other L1Oc repeats have only one copy of this sequence. The longest repeat sequenced, L1Oc5, is 6.5 kb long, and genomic blot-hybridization data using probes from the 5′ end of L1Oc5 indicate that a full length L1Oc repeat is about 7.5 kb long, extending about 1 kb 5′ to the sequenced region. The L1Oc5 sequence has long open reading frames (ORFs) that correspond to ORF-1 and ORF-2 described in the mouse L1 sequence. In contrast to the overlapping reading frames seen for mouse L1, ORF-1 and ORF-2 are in the same reading frame in rabbit and human L1s, resulting in a discistronic structure. The region between the likely stop codon for ORF-1 and the proposed start codon for ORF-2 is not conserved in interspecies comparisons, which is further evidence that this short region does not encode part of a protein. ORF-1 appears to be a hybrid of sequences, of which the 3′ half is unique to and conserved in mammalian L1 repeats. The 5′ half of ORF-1 is not conserved between mammalian L1 repeats, but this segment of L1Oc is related significantly to type II cytoskeletal keratin. [ABSTRACT FROM AUTHOR]

The article discusses the study conducted by S. Watanabe which identifies that the protein demethylase JMJD1C is the first selective branch-factor required for BRCA1-dependent HR-mediated repair. It examines how JMJD1C affect RAP80-BRCA1 recruitment to double-strand breaks (DSBs). It states that the earliest proteins detected at DSBs is the mediator of DNA-damage checkpoint 1 (MDC1). It also mentions that JMJD1C enhance interactions between MDC1 and RNF8.

The activation and targeted localization of ubiquitin E2 conjugating enzymes could provide a point for regulating ubiquitin-dependent cell functions. Supporting this view, the ubiquitin charging and activation of a class of E2 enzymes has been directly linked to their nuclear import. [ABSTRACT FROM AUTHOR]

The article focuses on the small ubiquitin-like modifier (SUMO) ligase. Ubiquitin-like protein modifiers (Ubls) regulate a host of processes in eukaryotic cells. Ubiquitin, the original Ubl, signals proteolysis and diverse non-proteolytic outcomes. Functional breadth is also characteristic of SUMO. SUMOylation was discovered because it is required to localize RanGAP1, a regulator of nucleocytoplasmic transport, to nuclear pore complexes. Studies of the SUMOylation of RanGAP1 and other substrates have revealed distinguishing features. SUMOylation, unlike ubiquitylation, is frequently site specific. SUMOylation of RanGAPl is frequently observable with just E1 and Ubc9 (the SUMO E2).