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Integrin Regulation of Ferroptosis in the Context of Tumor Heterogeneity
Get PDFBreast cancer diagnosis and treatment are complicated by the fact that most solid tumors are heterogeneous and composed of diverse populations of tumor cells, as well as stromal cells. Tumor heterogeneity challenges therapeutic approaches because different populations of tumor cells within a given tumor respond differently to a given therapy. For this reason, it is essential to understand the mechanisms by which specific populations of tumor cells respond to therapy so that effective combinatorial strategies can be developed that overcome the challenge of heterogeneity. Within this context, a major goal of this thesis is to understand how diverse populations of tumor cells in breast cancer respond to ferroptosis. Using single cell RNA sequencing, I found that a subpopulation of luminal tumor cells with high expression of GATA3 in a patient derived organoid, show selective resistance to ferroptosis. Investigation of mechanism involved revealed that GATA3 promotes ferroptosis resistance through the downregulation of integrin β1 and the consequent inactivation of a FAK-ROCK-YAP pathway that culminates in the repression of ACSL4, a lipid-modifying enzyme that is essential for ferroptosis. These findings imply a novel role for integrin β1 signaling in promoting ferroptosis sensitivity and highlight the role of intratumor heterogeneity in determining ferroptosis resistance.
The second part of my thesis focuses on the role of integrins in manipulating transcriptional patterns in epithelial and mesenchymal cell types. In this work, I highlight opposing roles for YAP/TAZ in promoting transcription of the LAMC2 subunit of laminin 332 in epithelial cell types while repressing transcription in mesenchymal cell types. Further investigation of this mechanism revealed that ZEB1 represses transcription of LAMC2 in mesenchymal cell types. In epithelial cell types, however, integrin β4 promotes downregulation of ZEB1 through mir200 and this repression allows for YAP/TAZ mediated transcription of LAMC2. Finally, I demonstrate that this positive feedback loop, in which integrin β4 promotes expression of its ligand laminin332, promotes an epithelial differentiated ferroptosis resistant cell state.
In summary, this thesis highlights the effects of tumor heterogeneity on the response to ferroptosis inhibitors and demonstrates the role that integrins have in influencing cell state and ferroptosis sensitivity.Cancer Biology6 months2025-11-2
UMCCTS Newsletter, November 2024
Get PDFThis is the Novembr 2024 issue of the UMass Center for Clinical and Translational Science Newsletter containing news and events of interest.Supported by the National Center for Advancing Translational Sciences, National Institutes of Health, through Grants UL1 TR001453, TL1 TR001454 and KL2 TR001455.No embarg
Microglia promote inflammatory cell death upon neuronal mitochondrial impairment during neurodegeneration
No full textThe failure to clear dysfunctional mitochondria, cell death and inflammation have been linked in neurodegenerative disease, but their relationship and role in these conditions is not fully understood. Loss of Vps13d prevents clearance of mitochondria, and mutations in human VPS13D have been associated with neurological movement disorders. To investigate the relationship between mitochondrial health, inflammation and neurodegeneration, we created a conditional Vps13d-knockout mouse. Loss of Vps13d in excitatory neurons resulted in behavioral changes and neurodegeneration. Vacuolar protein sorting 13D (VPS13D) deficiency also caused mitochondrial ultrastructural defects and dysfunction in neurons followed by gasdermin E processing, cyclic GMP-AMP synthase (cGAS)-stimulator of interferon response cGAMP interactor (STING) signaling, microglial activation and cell death. Gasdermin E localization with mitochondria in Vps13d-mutant neurons was required for elevated extracellular mitochondrial DNA that promoted activation of microglia. Depletion of microglia suppressed cell death and behavioral phenotypes but not mitochondrial changes in the neuron-specific Vps13d-knockout model, indicating that microglia promote cell death in this model of neurodegenerative disease.No embarg
Deep sequencing of yeast and mouse tRNAs and tRNA fragments using OTTR
Get PDFAmong the major classes of RNAs in the cell, tRNAs remain the most difficult to characterize via deep sequencing approaches, as tRNA structure and nucleotide modifications can each interfere with cDNA synthesis by commonly-used reverse transcriptases (RTs). Here, we benchmark a recently-developed RNA cloning protocol, termed Ordered Two-Template Relay (OTTR), to characterize intact tRNAs and tRNA fragments in budding yeast and in mouse tissues. We show that OTTR successfully captures both full-length tRNAs and tRNA fragments in budding yeast and in mouse reproductive tissues without any prior enzymatic treatment, and that tRNA cloning efficiency can be further enhanced via AlkB-mediated demethylation of modified nucleotides. As with other recent tRNA cloning protocols, we find that a subset of nucleotide modifications leave misincorporation signatures in OTTR datasets, enabling their detection without any additional protocol steps. Focusing on tRNA cleavage products, we compare OTTR with several standard small RNA-Seq protocols, finding that OTTR provides the most accurate picture of tRNA fragment levels by comparison to "ground truth" Northern blots. Applying this protocol to mature mouse spermatozoa, our data dramatically alter our understanding of the small RNA cargo of mature mammalian sperm, revealing a far more complex population of tRNA fragments - including both 5' and 3' tRNA halves derived from the majority of tRNAs - than previously appreciated. Taken together, our data confirm the superior performance of OTTR to commercial protocols in analysis of tRNA fragments, and force a reappraisal of potential epigenetic functions of the sperm small RNA payload.No embarg
High Throughput Genetic Screens in Escherichia coli – From Metabolic Interactions to Antibacterial Mechanism of Action
No full textGenetic screens are powerful tools for functional genomics. I developed a method to substantially increase the throughput of genetic screens in Escherichia coli and applied it to study bacteria–bacteria and bacteria–drug interactions.
To investigate metabolic cross-feeding in microbial communities, I systematically screened single-gene knockouts and found that strains benefiting from shared goods were typically defective in amino acid, nucleotide, or vitamin biosynthesis, or in central carbon metabolism. Pairwise experiments identified vitamin auxotrophs as optimal cross-feeding partners. In larger assemblies of auxotrophs, consortia rapidly coalesced around vitamin-deficient strains, stabilized by multiple cross-feeding interactions. These results suggest vitamins are ideal shared goods by supporting community growth while maintaining member interdependence.
To explore the antibacterial activity of non-antibiotic drugs, I screened 200 compounds using a barcoded E. coli knockout library. Network analysis of drug–drug similarities revealed that antibiotics clustered into modules corresponding to known mechanisms of action, whereas non-antibiotics largely remained unconnected. Nonetheless, about half of non-antibiotics formed distinct clusters, indicating shared and potentially unexploited antibacterial targets. Additionally, analysis of efflux systems revealed they impact antibiotics and non-antibiotics alike, raising concerns that non-antibiotic exposure may promote cross-resistance in vivo. Based on these findings I designed a custom library and machine learning pipeline for mechanism-of-action discovery.
My studies demonstrate the versatility of high-throughput genetic screening for investigating microbial ecology and pharmacology. By advancing both methodological and biological understanding, this work highlights how genetic screens can uncover principles of cooperation in microbial communities and illuminate the under-characterized antibacterial effects of non-antibiotic drugs.Systems, Computational, and Quantitative Biology1 year2026-11-1
Childhood adversity is associated with longitudinal white matter changes after adulthood trauma
No full textBackground: Childhood adversity is associated with susceptibility to posttraumatic stress disorder (PTSD) in adulthood. PTSD and childhood adversity are linked to white matter microstructure, yet the role of white matter as a potential neural mechanism connecting childhood adversity to PTSD remains unclear. The present study investigated the potential moderating role of previous childhood adversity on longitudinal changes in white matter microstructure and posttraumatic stress symptoms following a recent traumatic event in adulthood.
Methods: As part of the AURORA Study, 114 recent trauma survivors completed diffusion weighted imaging at 2-weeks and 6-months after exposure. Participants reported on prior childhood adversity and PTSD symptoms at 2-weeks, 6-months, and 12-months post-trauma. We performed both region-of-interest (ROI) using fractional anisotropy (FA) and whole-brain correlational tractography using quantitative anisotropy (QA) analyses to index associations between white matter microstructure changes and prior adversity.
Results: ROI-based analyses did not identify significant associations between childhood adversity and changes in FA. Whole-brain correlational tractography revealed that greater childhood adversity moderated the QA changes within threat and visual processing tracts including the cingulum bundle and inferior fronto-occipital fasciculus (IFOF). QA changes within cingulum bundle and IFOF were associated with changes in PTSD symptoms between 2-weeks and 6-months.
Conclusions: Our findings suggest temporal variability in threat and visual white matter tracts may be a potential neural pathway through which childhood adversity confers risk to PTSD symptoms after adulthood trauma. Future studies should take the temporal properties of white matter into consideration to better understand the neurobiology of childhood adversity and PTSD.No embarg
Animal Models of Orthopedic Implant-Associated Infections and Revisions
No full textOrthopedic implant-associated infections such as prosthetic joint infections (PJIs) lead to devastating complications for patients and impose significant financial burdens on the healthcare systems. Although the primary orthopedic implant associated infection rate is relatively low (0.3-9%), the reinfection rate after implant revisions can be as high as 20% to 40%. To evaluate novel therapeutic strategies for preventing and treating infections associated with primary and revision implants, it is essential to develop appropriate animal models that closely emulate clinical realities. Here we discuss existing animal models developed for orthopedic implant revision surgeries including small animal models in rats and mice, and larger animal models in rabbits, sheep, and mini-pigs. While larger animal models offer the advantage of more closely mimicking human surgical procedures, implant dimensions, and infection treatment protocols, rodent models are more cost-effective and better suited for screening experimental prophylaxes and therapeutics. Existing animal revision models have focused on primary infections established by () and revisions involving both one-stage and two-stage procedures. Further development of smaller animal implant revision models that implement more clinically relevant surgical procedures and recapitulate polymicrobial infections could facilitate the discovery and more rigorous evaluation of novel implant coating prophylaxes and therapeutics for reducing reinfection rates following implant revisions.No embarg
Paramagnetic rim lesions are highly specific for multiple sclerosis in real-world data
Get PDFParamagnetic rim lesions (PRLs) are an emerging biomarker for multiple sclerosis representing chronic, low-grade intraparenchymal brain inflammation. In addition to associating with greater disease severity, PRLs may be diagnostically supportive. Our aim in this study was to determine PRL specificity and sensitivity for discriminating multiple sclerosis from its diagnostic mimics using real-world clinical diagnostic and imaging data. This is a retrospective, cross-sectional analysis of a longitudinal cohort of patients with prospectively collected observational data. Patients were included if they underwent clinical evaluation in our academic neuroimmunology centre and had an available MRI scan from the same clinical 3-T magnet that included a T2*-weighted sequence with susceptibility post-processing (Susceptibility Weighted ANgiography protocol, General Electric). Susceptibility imaging-derived filtered phase maps and corresponding T2-fluid attenuated inversion recovery images were manually reviewed to determine PRLs. PRLs were categorized as 'definite', 'probable' or 'possible' based on modified, recent consensus criteria. We hypothesized that PRLs would convey a high specificity to discriminate multiple sclerosis from its MRI mimics. Five hundred seventy-four patients were evaluated in total: 473 with multiple sclerosis, 53 with non-inflammatory neurological disease and 48 with other inflammatory neurological disease. Identification of 'definite' or 'probable' PRL provided a specificity of 98% to discriminate multiple sclerosis from non-inflammatory neurological disease and other inflammatory neurological disease; sensitivity was 36%. Interrater agreement was almost perfect for definite/probable identification at a subject level. PRLs convey high specificity for multiple sclerosis and can aid in the diagnostic evaluation. Modest sensitivity limits their use as single diagnostic indicators. Including lesions with lower confidence ('possible') rapidly erodes specificity and should be interpreted with caution given the potential harms associated with misdiagnosis.No embarg
A National Evaluation of Risk Factors for Colorectal Cancer Using NHANES Data (2013-2023)
No full textBackground: Colorectal cancer (CRC) is a common malignancy with nearly 2 million new cases documented in 2020. There were approximately 152,810 new cases of CRC in 2024 emphasizing a need for ongoing preventive research. This study evaluated the relationships between CRC diagnosis and disease risk factors. Current literature provides information about risk factors, however there are gaps in knowledge about factor interactions. This study aims to address these gaps to guide by exploring potential interactions that could increase CRC risk.
Methods: Using National Health and Nutrition Examination Survey (NHANES) data from 2013 to 2023, a secondary data analysis explore the associations between previously identified CRC risk factors and a CRC diagnosis.
Results: Major findings showed that individuals who identified as ‘Other’ race were most likely to have CRC. Study findings supported prior research associations between education level, race/ethnicity, diet, body composition, alcohol consumption, and CRC diagnosis. Additionally, education level mediated the relationship between race/ethnicity and CRC diagnosis. There was a significant interaction between race/ethnicity and waist to height ratio (WHR) with non-Hispanic Asians (NHAs) with an increased WHR the highest likelihood of CRC diagnosis.
Conclusions: Enhanced longitudinal data collection among diverse populations is warranted to guide intervention work on CRC incidence reduction, particularly among high-risk groups.6 months2/19/202
Investigating the Basic Biology of Cryptic Splicing and Associated Therapeutic Opportunities
No full textThe odyssey our genetic information undergoes as described in the Central Dogma of Molecular Biology is rife with potential pitfalls and yet we remain relatively - remarkably - unscathed. To shuttle our genetic information, messenger RNA (mRNA), must undergo modifications such as splicing. Splicing, which involves a plethora of small nuclear RNAs (snRNAs) and proteins, serves to remove introns and ligate flanking exons. The complexity of mRNA sequence recognition and spliceosome formation leaves the process vulnerable to mistakes such as the recognition of an erroneous site, hereafter referred to as cryptic splicing. In this thesis, I sought to elucidate the prevalence and peculiarities of cryptic splice sites to better understand the molecular characteristics governing their usage. To this end, I isolated, sequenced, categorized, and characterized these cryptic splice sites and discovered a number of sequence motifs and gene features that influence the erroneous usage of these sites. Subdividing the identified cryptic sites into 3 classes (recursive, high-fidelity, and low-fidelity), I assessed the predicted consequences of splicing at those sites with an explicit interest in sites that induce inclusion of a novel, cryptic exon. Additionally, I explored the therapeutic potential of redirecting cryptic splicing through steric inhibition of erroneous sites and subsequent skipping of cryptic exons. Ultimately, this thesis significantly expands our previously limited knowledge of cryptic splicing phenomena while better defining which classes of cryptic splice sites possess the greatest therapeutic potential.Neuroscience2 years2027-03-3