Biomedical subjects

David E Root

Publications and source records attributed to David E Root.

9 recordsLinked to original sources

PubMed2026

FANCM is required for the PAX3::FOXO1-driven oncogenic program in rhabdomyosarcoma.

Many cancers are driven by mutationally altered transcription factors (TFs) that rewire cells to an oncogenic state. Cells must activate specific mechanisms to tolerate the burden of oncogenic TF activity. To define such mechanisms, we focused on a canonical oncogenic fusion protein-driven cancer, alveolar rhabdomyosarcoma (ARMS), where the PAX3::FOXO1 fusion protein hyperactivates and mislocalizes PAX3 and FOXO1 TF functions. Employing sequential functional genomic CRISPR-Cas9 screens, we identified FANCM, a DNA translocase in the Fanconi anemia pathway, as a selective dependency in PAX3::FOXO1+ ARMS. FANCM loss reduces fusion protein levels, induces myogenic differentiation, and disrupts the PAX3::FOXO1 transcriptional program, thereby halting oncogenic proliferation. Mechanistically, FANCM depletion exacerbates replication stress (RS) and DNA damage signaling, with chromatin-associated RS enriched at PAX3::FOXO1 target gene loci, resulting in selective downregulation of the oncogenic program. CRISPR exon-tiling screens prioritized FANCM's helicase and DNA-binding domains as essential for this dependency, linking FANCM-mediated replication fork binding to sustained oncogenesis.

ARMS↗

PubMed2026

Expanding the human proteome with microproteins and peptideins.

A major scientific drive is to characterize the protein-coding genome, which is a primary basis for studying human health. But the fundamental question remains of what has been missed in previous analyses. Over the past decade, the translation of non-canonical open reading frames (ncORFs) has been observed across human cell types and disease states1-3, with major implications for biomedical science. However, a key gap in knowledge has been which ncORFs produce small microproteins or alternative protein molecules that contribute to the human proteome. Here we report the collaborative efforts of the TransCODE Consortium4 to produce a consensus landscape of protein-level evidence for ncORFs. We show that about 25% of a set of 7,264 ncORFs gives rise to detectable peptides in a large-scale analysis of 95,520 proteomics experiments. We develop an annotation framework for ncORF-encoded microproteins as human proteins and codify the new conceptual model of 'peptideins' as microproteins that have indeterminate potential as functional proteins. To probe the biological implications of peptideins, we create an evolutionary analysis approach, termed ORF relative branch length (ORBL), and determine that evolutionary constraint is common and associates with observation of ncORF-derived peptides. We then characterize a pan-essential cellular phenotype for one peptidein from the OLMALINC long non-coding RNA. Overall, we generate public research tools supported by GENCODE and PeptideAtlas and advance biomedical discovery for understudied components of the human proteome.

Humans↗

PubMed2004

Indoprofen upregulates the survival motor neuron protein through a cyclooxygenase-independent mechanism.

Most patients with the pediatric neurodegenerative disease spinal muscular atrophy have a homozygous deletion of the survival motor neuron 1 (SMN1) gene, but retain one or more copies of the closely related SMN2 gene. The SMN2 gene encodes the same protein (SMN) but produces it at a low efficiency compared with the SMN1 gene. We performed a high-throughput screen of approximately 47,000 compounds to identify those that increase production of an SMN2-luciferase reporter protein, but not an SMN1-luciferase reporter protein. Indoprofen, a nonsteroidal anti-inflammatory drug (NSAID) and cyclooxygenase (COX) inhibitor, selectively increased SMN2-luciferase reporter protein and endogenous SMN protein and caused a 5-fold increase in the number of nuclear gems in fibroblasts from SMA patients. No other NSAIDs or COX inhibitors tested exhibited this activity.

Animals↗

PubMed2004

A flexible data analysis tool for chemical genetic screens.

High-throughput assays generate immense quantities of data that require sophisticated data analysis tools. We have created a freely available software tool, SLIMS (Small Laboratory Information Management System), for chemical genetics which facilitates the collection and analysis of large-scale chemical screening data. Compound structures, physical locations, and raw data can be loaded into SLIMS. Raw data from high-throughput assays are normalized using flexible analysis protocols, and systematic spatial errors are automatically identified and corrected. Various computational analyses are performed on tested compounds, and dilution-series data are processed using standard or user-defined algorithms. Finally, published literature associated with active compounds is automatically retrieved from Medline and processed to yield potential mechanisms of actions. SLIMS provides a framework for analyzing high-throughput assay data both as a laboratory information management system and as a platform for experimental analysis.

Cyclic AMP Response Element-Binding Protein↗

PubMed2003

Conserved pathways within bacteria and yeast as revealed by global protein network alignment.

We implement a strategy for aligning two protein-protein interaction networks that combines interaction topology and protein sequence similarity to identify conserved interaction pathways and complexes. Using this approach we show that the protein-protein interaction networks of two distantly related species, Saccharomyces cerevisiae and Helicobacter pylori, harbor a large complement of evolutionarily conserved pathways, and that a large number of pathways appears to have duplicated and specialized within yeast. Analysis of these findings reveals many well characterized interaction pathways as well as many unanticipated pathways, the significance of which is reinforced by their presence in the networks of both species.

Bacteria↗

PubMed2003

Spectroscopic and electronic structure studies of the diamagnetic side-on CuII-superoxo complex Cu(O2)[HB(3-R-5-iPrpz)3]: antiferromagnetic coupling versus covalent delocalization.

Magnetic, vibrational, and optical techniques are combined with density functional calculations to elucidate the electronic structure of the diamagnetic mononuclear side-on CuII-superoxo complex. The electronic nature of its lowest singlet/triplet states and the ground-state diamagnetism are explored. The triplet state is found to involve the interaction between the Cu xy and the superoxide pi v * orbitals, which are orthogonal to each other. The singlet ground state involves the interaction between the Cu xy and the in-plane superoxide pi v * orbitals, which have a large overlap and thus strong bonding. The ground-state singlet/triplet states are therefore fundamentally different in orbital origin and not appropriately described by an exchange model. The ground-state singlet is highly delocalized with no spin polarization.

Copper↗

PubMed2003

Biological mechanism profiling using an annotated compound library.

We present a method for testing many biological mechanisms in cellular assays using an annotated library of 2036 small organic molecules. This annotated compound library represents a large-scale collection of compounds with diverse, experimentally confirmed biological mechanisms and effects. We found that this chemical library is (1) more structurally diverse than conventional, commercially available libraries, (2) enriched in active compounds in a tumor cell viability assay, and (3) capable of generating hypotheses regarding biological mechanisms underlying cellular processes. We elucidated biological mechanisms relevant to the antiproliferative activity of 85 compounds from this library that were selected using a high-throughput cell viability screen. We developed a novel automated scoring system for identifying statistically enriched mechanisms among such a subset of compounds. This scoring system can identify both previously known and potentially novel antiproliferative mechanisms.

Cell Line, Tumor↗

PubMed2003

Detecting spatial patterns in biological array experiments.

Chemical genetic screening and DNA and protein microarrays are among a number of increasingly important and widely used biological research tools that involve large numbers of parallel experiments arranged in a spatial array. It is often difficult to ensure that uniform experimental conditions are present throughout the entire array, and as a result, one often observes systematic spatially correlated errors, especially when array experiments are performed using robots. Here, the authors apply techniques based on the discrete Fourier transform to identify and quantify spatially correlated errors superimposed on a spatially random background. They demonstrate that these techniques are effective in identifying common spatially systematic errors in high-throughput 384-well microplate assay data. In addition, the authors employ a statistical test to allow for automatic detection of such errors. Software tools for using this approach are provided.

Analysis of Variance↗

PubMed2002

Global analysis of large-scale chemical and biological experiments.

Research in the life sciences is increasingly dominated by high-throughput data collection methods that benefit from a global approach to data analysis. Recent innovations that facilitate such comprehensive analyses are highlighted. Several developments enable the study of the relationships between newly derived experimental information, such as biological activity in chemical screens or gene expression studies, and prior information, such as physical descriptors for small molecules or functional annotation for genes. The way in which global analyses can be applied to both chemical screens and transcription profiling experiments using a set of common machine learning tools is discussed.

Animals↗