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Xeroderma pigmentosum (XP) is characterized by defective repair of ultraviolet radiation(UVR)-induced DNA damage. Patients have UVR hypersensitivity and increased skin cancer risk. Effective photoprotection has reduced childhood cancer-related deaths, but revealed adolescence-onset neurodegeneration, arising through unknown mechanisms. Here, we investigate XP neurodegeneration using pluripotent stem cells derived from XP patients and healthy relatives, performing functional multi-omics on samples during neuronal differentiation. We find endoplasmic reticulum stress is upregulated, preceded by oxidative stress, causing substantial 5’,8-cyclopurine and 8-oxopurine DNA damage. Critically, XP neurons exhibit inappropriate downregulation of the protein clearance ubiquitin-proteasome system (UPS). Chemical enhancement of UPS activity improves phenotypes, albeit inadequately, implying that early detection/prevention strategies are necessary to produce clinically impactful outcomes. Thus, we develop an early detection assay predicting neurodegeneration in at-risk patients.

Supplementary Figures 1-5 from Inhibitors of the Proteasome Suppress Homologous DNA Recombination in Mammalian Cells

Proteasome inhibitors are novel antitumor agents against multiple myeloma and other malignancies. Despite the increasing clinical application, the molecular basis of their antitumor effect has been poorly understood due to the involvement of the ubiquitin-proteasome pathway in multiple cellular metabolisms. Here, we show that treatment of cells with proteasome inhibitors has no significant effect on nonhomologous end joining but suppresses homologous recombination (HR), which plays a key role in DNA double-strand break (DSB) repair. In this study, we treat human cells with proteasome inhibitors and show that the inhibition of the proteasome reduces the efficiency of HR-dependent repair of an artificial HR substrate. We further show that inhibition of the proteasome interferes with the activation of Rad51, a key factor for HR, although it does not affect the activation of ATM, γH2AX, or Mre11. These data show that the proteasome-mediated destruction is required for the promotion of HR at an early step. We suggest that the defect in HR-mediated DNA repair caused by proteasome inhibitors contributes to antitumor effect, as HR plays an essential role in cellular proliferation. Moreover, because HR plays key roles in the repair of DSBs caused by chemotherapeutic agents such as cisplatin and by radiotherapy, proteasome inhibitors may enhance the efficacy of these treatments through the suppression of HR-mediated DNA repair pathways. [Cancer Res 2007;67(18):8536–43]

Xeroderma pigmentosum (XP) is a genetic disorder caused by mutations in genes of the Nucleotide Excision Repair (NER) pathway (groups A-G) or in Translesion Synthesis (TLS) DNA polymerase η (V). XP is associated with an increased skin cancer risk, reaching, for some groups, several thousand-fold compared to the general population. Here, we analyzed 38 skin cancer genomes from five XP groups. We found that the activity of NER determines heterogeneity of the mutation rates across skin cancer genomes and that transcription-coupled NER extends beyond the gene boundaries reducing the intergenic mutation rate. Mutational profile in XP-V tumors and experiments withPOLH-KO cell line revealed the role of polymerase η in the error-free bypass of (i) rare TpG and TpA DNA lesions, (ii) 3’ nucleotides in pyrimidine dimers, and (iii) TpT photodimers. Our study unravels the genetic basis of skin cancer risk in XP and provides insights into the mechanisms reducing UV-induced mutagenesis in the general population.

Cockayne syndrome is a rare autosomal recessive disorder with dwarfism, mental retardation, and otherwise clinically heterogeneous features. Classically, the onset of Cockayne syndrome starts in the second year of life. The failure of RNA synthesis to recover to normal rates after UV-C irradiation provides a useful diagnostic test and the clinical feature that correlates most strongly with defective RNA synthesis is photosensitivity.To report an unusual case of Cockayne Syndrome.A case of a five-year-old girl with Cockayne with an onset in early infancy the girl and without photosensitivity is presented. The diagnosis was confirmed by the failure of RNA synthesis to recover to normal rate after UV-C irradiation. The patient died at the age of 6 of pneumonia.Although rare, Cockayne syndrome may be presented without photosensitivity and had an early onset.

During translesion synthesis past sites of damaged DNA, specialized Y-family polymerases are employed by the cell to replace the high stringency replicative polymerases and synthesize DNA past the damaged site. These polymerases are localized in replication factories during the S phase of the cell cycle. When progress of the replication fork is blocked, the polymerase accessory protein, proliferating cell nuclear antigen (PCNA), becomes ubiquitinated and the monoubiquitinated PCNA has an increased affinity for Y-family DNA polymerase eta (poleta). This chapter describes methods for visualizing the polymerases in replication factories, for analyzing the ubiquitination status of PCNA, and for measuring its interaction with poleta in chromatin extracts.

The deformation of a collider quadrupole magnet (CQM) coil under pre-compression, cool down and Lorentz forces may cause some significant changes in magnetic multipoles. The magnetic measurements of the CQM short model magnets at 4.2 K showed an average b/sub 5/ of -2.866 units compared to a numerical prediction of -0.162 units. Out of these -2.866 units, -1.5 units are due to a shim placed at the mid-plane to increase coil precompression. We also suspect that the permeability of the stainless steel collar may contribute some of the missing b/sub 5/. For example, a permeability of 1.004 for the collar adds -0.5 units of b/sub 5/. Including the collar, we are still missing -0.8 to -1.0 units. We believe that the mechanical deformation due to pre-compression of the coil is responsible for the remaining units. This study builds a 2D magneto-structural coupled model, calculates the magnetic multipoles under different loading conditions and establishes a correlation between multipole changes and coil pre-compression and Lorentz force stresses. >