The TFIIH components p44/p62 act as a damage sensor during nucleotide excision repair

Nucleotide excision repair (NER) protects the genome following exposure to diverse types of DNA damage, including UV light and chemotherapeutics. Mutations in human NER genes lead to diseases such as xeroderma pigmentosum and Cockayne syndrome. In eukaryotes, the major transcription factor TFIIH is the central hub of NER. The core components of TFIIH include the helicases XPB, XPD, and the five core structural subunits. Two of these core-TFIIH proteins, p44 and p62 remain relatively unstudied; although p44 is known to regulate the helicase activity of XPD during NER. p62's role is thought to be structural; however, a recent cryo-EM structure shows p44, p62, and XPD making contacts with each other, implying a more extensive role in DNA repair beyond the structural integrity of TFIIH. Here, we show that p44 stimulates XPD's ATPase, but upon encountering DNA damage further stimulation is only observed when p62 is in the ternary complex. More significantly, we show that the p44/p62 complex binds DNA independently of XPD and diffuses along its backbone, indicating a novel DNA-binding entity in TFIIH. These data support a role for p44/p62 in TFIIH's mechanism of damage detection. This revises our understanding of TFIIH and prompts more extensive investigation of all of the core subunits, for an active role during both DNA repair and transcription

Harmonizing Lipidomics: NIST Interlaboratory Comparison Exercise for Lipidomics Using SRM 1950-metabolites in Frozen Human Plasma

As the lipidomics field continues to advance, self-evaluation within the community is critical. Here, we performed an interlaboratory comparison exercise for lipidomics using Standard Reference Material (SRM) 1950-Metabolites in Frozen Human Plasma, a commercially available reference material. The interlaboratory study comprised 31 diverse laboratories, with each laboratory using a different lipidomics workflow. A total of 1,527 unique lipids were measured across all laboratories and consensus location estimates and associated uncertainties were determined for 339 of these lipids measured at the sum composition level by five or more participating laboratories. These evaluated lipids detected in SRM 1950 serve as community-wide benchmarks for intra-and interlaboratory quality control and method validation. These analyses were performed using nonstandardized laboratory-independent workflows. The consensus locations were also compared with a previous examination of SRM 1950 by the LIPID MAPS consortium.jlr While the central theme of the interlaboratory study was to provide values to help harmonize lipids, lipid mediators, and precursor measurements across the community, it was also initiated to stimulate a discussion regarding areas in need of improvement

OmoMYC blunts promoter invasion by oncogenic MYC to inhibit gene expression characteristic of MYC-dependent tumors.

MYC genes have both essential roles during normal development and exert oncogenic functions during tumorigenesis. Expression of a dominant-negative allele of MYC, termed OmoMYC, can induce rapid tumor regression in mouse models with little toxicity for normal tissues. How OmoMYC discriminates between physiological and oncogenic functions of MYC is unclear. We have solved the crystal structure of OmoMYC and show that it forms a stable homodimer and as such recognizes DNA in the same manner as the MYC/MAX heterodimer. OmoMYC attenuates both MYC-dependent activation and repression by competing with MYC/MAX for binding to chromatin, effectively lowering MYC/MAX occupancy at its cognate binding sites. OmoMYC causes the largest decreases in promoter occupancy and changes in expression on genes that are invaded by oncogenic MYC levels. A signature of OmoMYC-regulated genes defines subgroups with high MYC levels in multiple tumor entities and identifies novel targets for the eradication of MYC-driven tumors

Quality control requirements for the correct annotation of lipidomics data.

10.1038/s41467-021-24984-yNature Communications121477

High-Resolution Mass Spectrometry for Human Exposomics: Expanding Chemical Space Coverage

In the modern "omics" era, measurement of the human exposome is a critical missing link between genetic drivers and disease outcomes. High-resolution mass spectrometry (HRMS), routinely used in proteomics and metabolomics, has emerged as a leading technology to broadly profile chemical exposure agents and related biomolecules for accurate mass measurement, high sensitivity, rapid data acquisition, and increased resolution of chemical space. Non-targeted approaches are increasingly accessible, supporting a shift from conventional hypothesis-driven, quantitation-centric targeted analyses toward data-driven, hypothesis-generating chemical exposome-wide profiling. However, HRMS-based exposomics encounters unique challenges. New analytical and computational infrastructures are needed to expand the analysis coverage through streamlined, scalable, and harmonized workflows and data pipelines that permit longitudinal chemical exposome tracking, retrospective validation, and multi-omics integration for meaningful health-oriented inferences. In this article, we survey the literature on state-of-the-art HRMS-based technologies, review current analytical workflows and informatic pipelines, and provide an up-to-date reference on exposomic approaches for chemists, toxicologists, epidemiologists, care providers, and stakeholders in health sciences and medicine. We propose efforts to benchmark fit-for-purpose platforms for expanding coverage of chemical space, including gas/liquid chromatography-HRMS (GC-HRMS and LC-HRMS), and discuss opportunities, challenges, and strategies to advance the burgeoning field of the exposome

Lipidomics and Redox Lipidomics Indicate Early Stage Alcohol-Induced Liver Damage.

Alcoholic fatty liver disease (AFLD) is characterized by lipid accumulation and inflammation and can progress to cirrhosis and cancer in the liver. AFLD diagnosis currently relies on histological analysis of liver biopsies. Early detection permits interventions that would prevent progression to cirrhosis or later stages of the disease. Herein, we have conducted the first comprehensive time-course study of lipids using novel state-of-the art lipidomics methods in plasma and liver in the early stages of a mouse model of AFLD, i.e., Lieber-DeCarli diet model. In ethanol-treated mice, changes in liver tissue included up-regulation of triglycerides (TGs) and oxidized TGs and down-regulation of phosphatidylcholine, lysophosphatidylcholine, and 20-22-carbon-containing lipid-mediator precursors. An increase in oxidized TGs preceded histological signs of early AFLD, i.e., steatosis, with these changes observed in both the liver and plasma. The major lipid classes dysregulated by ethanol play important roles in hepatic inflammation, steatosis, and oxidative damage. Conclusion: Alcohol consumption alters the liver lipidome before overt histological markers of early AFLD. This introduces the exciting possibility that specific lipids may serve as earlier biomarkers of AFLD than those currently being used

Increased throughput and ultra-high mass resolution in DESI FT-ICR MS imaging through new-generation external data acquisition system and advanced data processing approaches

Desorption electrospray ionisation-mass spectrometry imaging (DESI-MSI) is a powerful imaging technique for the analysis of complex surfaces. However, the often highly complex nature of biological samples is particularly challenging for MSI approaches, as options to appropriately address mass spectral complexity are limited. Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) offers superior mass accuracy and mass resolving power, but its moderate throughput inhibits broader application. Here we demonstrate the dramatic gains in mass resolution and/or throughput of DESI-MSI on an FT-ICR MS by developing and implementing a sophisticated data acquisition and data processing pipeline. The presented pipeline integrates, for the first time, parallel ion accumulation and detection, post-processing absorption mode Fourier transform and pixel-by-pixel internal re-calibration. To achieve that, first, we developed and coupled an external high-performance data acquisition system to an FT-ICR MS instrument to record the time-domain signals (transients) in parallel with the instrument’s built-in electronics. The recorded transients were then processed by the in-house developed computationally-efficient data processing and data analysis software. Importantly, the described pipeline is shown to be applicable even to extremely large, up to 1 TB, imaging datasets. Overall, this approach provides improved analytical figures of merits such as: (i) enhanced mass resolution at no cost in experimental time; and (ii) up to 4-fold higher throughput while maintaining a constant mass resolution. Using this approach, we not only demonstrate the record 1 million mass resolution for lipid imaging from brain tissue, but explicitly demonstrate such mass resolution is required to resolve the complexity of the lipidome