LRH-1/NR5A2 targets mitochondrial dynamics to reprogram type 1 diabetes macrophages and dendritic cells into an immune tolerance phenotype.

Background: The complex aetiology of type 1 diabetes (T1D), characterised by a detrimental cross-talk between the immune system and insulin-producing beta cells, has hindered the development of effective disease-modifying therapies. The discovery that the pharmacological activation of LRH-1/NR5A2 can reverse hyperglycaemia in mouse models of T1D by attenuating the autoimmune attack coupled to beta cell survival/regeneration prompted us to investigate whether immune tolerisation could be translated to individuals with T1D by LRH-1/NR5A2 activation and improve islet survival.

Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from individuals with and without T1D and derived into various immune cells, including macrophages and dendritic cells.

Lrif1 modulates Trim28-mediated repression of the locus in mouse embryonic stem cells.

Germline mutations in SMCHD1, DNMT3B and LRIF1 can cause facioscapulohumeral muscular dystrophy type 2 (FSHD2). FSHD is an epigenetic skeletal muscle disorder in which partial failure in heterochromatinization of the D4Z4 macrosatellite repeat causes spurious expression of the repeat-embedded gene in skeletal muscle, ultimately leading to muscle weakness and wasting. All three proteins play a role in chromatin organization and gene silencing; however, their functional relationship has not been fully elucidated.

Targeting epigenetic regulation and post-translational modification with 5-Aza-2' deoxycytidine and SUMO E1 inhibition augments T-cell receptor therapy.

Background: Cellular immunotherapy using modified T cells offers new avenues for cancer treatment. T-cell receptor (TCR) engineering of CD8 T cells enables these cells to recognize tumor-associated antigens and tumor-specific neoantigens. Improving TCR T-cell therapy through increased potency and in vivo persistence will be critical for clinical success.

The small CRL4 ubiquitin ligase component DDA1 regulates transcription-coupled repair dynamics.

Transcription-blocking DNA lesions are specifically targeted by transcription-coupled nucleotide excision repair (TC-NER), which removes a broad spectrum of DNA lesions to preserve transcriptional output and thereby cellular homeostasis to counteract aging. TC-NER is initiated by the stalling of RNA polymerase II at DNA lesions, which triggers the assembly of the TC-NER-specific proteins CSA, CSB and UVSSA. CSA, a WD40-repeat containing protein, is the substrate receptor subunit of a cullin-RING ubiquitin ligase complex composed of DDB1, CUL4A/B and RBX1 (CRL4).

Physical interactions between specifically regulated subpopulations of the MCM and RNR complexes prevent genetic instability.

The helicase MCM and the ribonucleotide reductase RNR are the complexes that provide the substrates (ssDNA templates and dNTPs, respectively) for DNA replication. Here, we demonstrate that MCM interacts physically with RNR and some of its regulators, including the kinase Dun1. These physical interactions encompass small subpopulations of MCM and RNR, are independent of the major subcellular locations of these two complexes, augment in response to DNA damage and, in the case of the Rnr2 and Rnr4 subunits of RNR, depend on Dun1.

BRCA1/BARD1 ubiquitinates PCNA in unperturbed conditions to promote continuous DNA synthesis.

Deficiencies in the BRCA1 tumor suppressor gene are the main cause of hereditary breast and ovarian cancer. BRCA1 is involved in the Homologous Recombination DNA repair pathway and, together with BARD1, forms a heterodimer with ubiquitin E3 activity. The relevance of the BRCA1/BARD1 ubiquitin E3 activity for tumor suppression and DNA repair remains controversial.

Deciphering non-canonical ubiquitin signaling: biology and methodology.

Ubiquitination is a dynamic post-translational modification that regulates virtually all cellular processes by modulating function, localization, interactions and turnover of thousands of substrates. Canonical ubiquitination involves the enzymatic cascade of E1, E2 and E3 enzymes that conjugate ubiquitin to lysine residues giving rise to monomeric ubiquitination and polymeric ubiquitination. Emerging research has established expansion of the ubiquitin code by non-canonical ubiquitination of N-termini and cysteine, serine and threonine residues.

SUMO proteases: from cellular functions to disease.

Posttranslational modification by small ubiquitin-like modifiers (SUMOs) is critical in regulating diverse cellular processes including gene expression, cell cycle progression, genome integrity, cellular metabolism, and inflammation and immunity. The covalent attachment of SUMOs to target proteins is highly dynamic and reversible through the concerted action of SUMO conjugating and deconjugating enzymes. In mammalian cells, sentrin-specific proteases (SENPs) are the most abundant family of deconjugating enzymes.

BioE3 identifies specific substrates of ubiquitin E3 ligases.

Hundreds of E3 ligases play a critical role in recognizing specific substrates for modification by ubiquitin (Ub). Separating genuine targets of E3s from E3-interactors remains a challenge. We present BioE3, a powerful approach for matching substrates to Ub E3 ligases of interest.

DDA1, a novel factor in transcription-coupled repair, modulates CRL4 dynamics at DNA damage-stalled RNA polymerase II.

Transcription-blocking DNA lesions are specifically targeted by transcription-coupled nucleotide excision repair (TC-NER), which removes a broad spectrum of DNA lesions to preserve transcriptional output and thereby cellular homeostasis to counteract aging. TC-NER is initiated by the stalling of RNA polymerase II at DNA lesions, which triggers the assembly of the TC-NER-specific proteins CSA, CSB and UVSSA. CSA, a WD40-repeat containing protein, is the substrate receptor subunit of a cullin-RING ubiquitin ligase complex composed of DDB1, CUL4A/B and RBX1 (CRL4).

SENP6 regulates localization and nuclear condensation of DNA damage response proteins by group deSUMOylation.

The SUMO protease SENP6 maintains genomic stability, but mechanistic understanding of this process remains limited. We find that SENP6 deconjugates SUMO2/3 polymers on a group of DNA damage response proteins, including BRCA1-BARD1, 53BP1, BLM and ERCC1-XPF. SENP6 maintains these proteins in a hypo-SUMOylated state under unstressed conditions and counteracts their polySUMOylation after hydroxyurea-induced stress.

Exome sequencing links the SUMO protease SENP7 with fatal arthrogryposis multiplex congenita, early respiratory failure and neutropenia.

Background: SUMOylation involves the attachment of small ubiquitin-like modifier (SUMO) proteins to specific lysine residues on thousands of substrates with target-specific effects on protein function. Sentrin-specific proteases (SENPs) are proteins involved in the maturation and deconjugation of SUMO. Specifically, SENP7 is responsible for processing polySUMO chains on targeted substrates including the heterochromatin protein 1α (HP1α).

The lncRNA LETS1 promotes TGF-β-induced EMT and cancer cell migration by transcriptionally activating a TβR1-stabilizing mechanism.

Transforming growth factor-β (TGF-β) signaling is a critical driver of epithelial-to-mesenchymal transition (EMT) and cancer progression. In SMAD-dependent TGF-β signaling, activation of the TGF-β receptor complex stimulates the phosphorylation of the intracellular receptor-associated SMADs (SMAD2 and SMAD3), which translocate to the nucleus to promote target gene expression. SMAD7 inhibits signaling through the pathway by promoting the polyubiquitination of the TGF-β type I receptor (TβRI).

A disease-associated XPA allele interferes with TFIIH binding and primarily affects transcription-coupled nucleotide excision repair.

XPA is a central scaffold protein that coordinates the assembly of repair complexes in the global genome (GG-NER) and transcription-coupled nucleotide excision repair (TC-NER) subpathways. Inactivating mutations in XPA cause xeroderma pigmentosum (XP), which is characterized by extreme UV sensitivity and a highly elevated skin cancer risk. Here, we describe two Dutch siblings in their late forties carrying a homozygous H244R substitution in the C-terminus of XPA.

Inhibition of SUMOylation enhances DNA hypomethylating drug efficacy to reduce outgrowth of hematopoietic malignancies.

Combination therapies targeting malignancies aim to increase treatment efficacy and reduce toxicity. Hypomethylating drug 5-Aza-2'-deoxycytidine (5-Aza-2') enhances transcription of tumor suppressor genes and induces replication errors via entrapment of DNMT1, yielding DNA-protein crosslinks. Post-translational modification by SUMO plays major roles in the DNA damage response and is required for degradation of entrapped DNMT1.

Divergent Molecular and Cellular Responses to Low and High-Dose Ionizing Radiation.

Cancer risk after ionizing radiation (IR) is assumed to be linear with the dose; however, for low doses, definite evidence is lacking. Here, using temporal multi-omic systems analyses after a low (LD; 0.1 Gy) or a high (HD; 1 Gy) dose of X-rays, we show that, although the DNA damage response (DDR) displayed dose proportionality, many other molecular and cellular responses did not.

In-Plate Chemical Synthesis of Isopeptide-Linked SUMOylated Peptide Fluorescence Polarization Reagents for High-Throughput Screening of SENP Preferences.

Small ubiquitin-like modifiers (SUMOs) are conjugated to protein substrates in cells to regulate their function. The attachment of SUMO family members SUMO1-3 to substrate proteins is reversed by specific isopeptidases called SENPs (sentrin-specific protease). Whereas SENPs are SUMO-isoform or linkage type specific, comprehensive analysis is missing.

Exploration of nuclear body-enhanced sumoylation reveals that PML represses 2-cell features of embryonic stem cells.

Membrane-less organelles are condensates formed by phase separation whose functions often remain enigmatic. Upon oxidative stress, PML scaffolds Nuclear Bodies (NBs) to regulate senescence or metabolic adaptation. PML NBs recruit many partner proteins, but the actual biochemical mechanism underlying their pleiotropic functions remains elusive.

XPC-PARP complexes engage the chromatin remodeler ALC1 to catalyze global genome DNA damage repair.

Cells employ global genome nucleotide excision repair (GGR) to eliminate a broad spectrum of DNA lesions, including those induced by UV light. The lesion-recognition factor XPC initiates repair of helix-destabilizing DNA lesions, but binds poorly to lesions such as CPDs that do not destabilize DNA. How difficult-to-repair lesions are detected in chromatin is unknown.

Signalling mechanisms and cellular functions of SUMO.

Sumoylation is an essential post-translational modification that is catalysed by a small number of modifying enzymes but regulates thousands of target proteins in a dynamic manner. Small ubiquitin-like modifiers (SUMOs) can be attached to target proteins as one or more monomers or in the form of polymers of different types. Non-covalent readers recognize SUMO-modified proteins via SUMO interaction motifs.

THO complex deficiency impairs DNA double-strand break repair via the RNA surveillance kinase SMG-1.

The integrity and proper expression of genomes are safeguarded by DNA and RNA surveillance pathways. While many RNA surveillance factors have additional functions in the nucleus, little is known about the incidence and physiological impact of converging RNA and DNA signals. Here, using genetic screens and genome-wide analyses, we identified unforeseen SMG-1-dependent crosstalk between RNA surveillance and DNA repair in living animals.

Deubiquitinating enzymes and the proteasome regulate preferential sets of ubiquitin substrates.

The ubiquitin-proteasome axis has been extensively explored at a system-wide level, but the impact of deubiquitinating enzymes (DUBs) on the ubiquitinome remains largely unknown. Here, we compare the contributions of the proteasome and DUBs on the global ubiquitinome, using UbiSite technology, inhibitors and mass spectrometry. We uncover large dynamic ubiquitin signalling networks with substrates and sites preferentially regulated by DUBs or by the proteasome, highlighting the role of DUBs in degradation-independent ubiquitination.

Targeting pancreatic cancer by TAK-981: a SUMOylation inhibitor that activates the immune system and blocks cancer cell cycle progression in a preclinical model.

Objective: Pancreatic ductal adenocarcinoma (PDAC) has the characteristics of high-density desmoplastic stroma, a distinctive immunosuppressive microenvironment and is profoundly resistant to all forms of chemotherapy and immunotherapy, leading to a 5-year survival rate of 9%. Our study aims to add novel small molecule therapeutics for the treatment of PDAC.

Design: We have studied whether TAK-981, a novel highly selective and potent small molecule inhibitor of the small ubiquitin like modifier (SUMO) activating enzyme E1 could be used to treat a preclinical syngeneic PDAC mouse model and we have studied the mode of action of TAK-981.