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Innovative Algorithmic Approaches in Revenue Management
Full text linkIn this dissertation, I design and analyze tractable algorithms for sequential decision-making problems under uncertainty, with various applications in revenue management. More specifically, I focus on innovative business strategies that have emerged in recent years, with applications ranging from reusable product sales, opaque selling, and maritime shipping. For each application, I identify the unique operational requirements, highlight the complexity of the underlying decision problems, and develop tractable solution approaches. I also provide both theoretical analysis and numerical evaluation to evaluate the performance of the proposed policies. In the first chapter, I study the joint inventory and online assortment problem, wherein a decision maker (DM) must first select initial inventory levels for a collection of available products or resources, and then offer personalized assortments to customers who arrive over a finite selling horizon, and who make purchasing decisions according to a multinomial logit choice model. The goal across both sets of decisions is to maximize the expected revenue earned by the end of the selling horizon. We are the first to consider this joint optimization framework when the resources are reusable. That is, upon purchase or rental, each unit is consumed for a random duration, after which it returns to the DM for future use. Our cornerstone result when reusability is modeled in its classic form, is a constant factor approximation scheme when the usage duration distributions satisfy the increasing failure rate (IFR) property. In a nutshell, our approach exploits notions of submodularity within a fluid approximation of the original problem. This fluid problem approximates the IFR-based usage durations with appropriately defined geometric random variables. To show that this approximate approach is indeed valid requires establishing a novel link between the CDFs of geometric and IFR-distributed random variables, which may find broader applications beyond those considered in this paper. Next, we consider our joint optimization problem under an augmented version of basic reusability, wherein consumed resources can return to the DM as transformed versions of their original selves. The intent of this novel modeling feature is to capture reusability settings where the identity of a product can possibly change due to its consumption (a product purchased online and returned to the seller may become damaged during the try-on process) or through the very nature of a return (a bike rented at one dock may be returned to a different one). In this so-called network reusability setting, we propose a novel inventory refinement process that iteratively adjusts inventory decisions based on feedback from the online assortment stage. We ultimately establish a strong performance bound for our overall approach, which is network dependent. Through numerical experiments, we show that our approximation strategies perform nearly optimally across a wide range of reusability scenarios, demonstrating the robustness and practicality of our approach. In the second chapter, I study the algorithmic approach to mitigate the overbooking and no-show behavior in the maritime industry. In the airline industry, the practice of overbooking has been a celebrated operational tool that has led to revenue gains exceeding hundreds of millions of dollars when implemented correctly. By contrast, in the container shipping industry, the story of overbooking is filled with tales of chronic mistrust between shippers and carriers. Specifically, loose and unenforceable contracting practices have led to a failed market where shippers constantly renege on their agreement to produce containers as promised, and as a result, carriers overbook too frequently in an effort to hedge against this no-show behavior. The cost of such behaviors has been estimated to be in the range of \$30-40 billion annually, which highlights the glaring need for a remedy to this issue. In this paper, we propose and study a deposit-based booking system that draws inspiration from current practices that have been shown to be successful in mitigating no-show behavior and overbooking in the container shipping industry. Specifically, we consider a reservation system where inquiring shippers book cargo space using a customized deposit. The carrier, upon accepting the shipper’s booking request, matches the shipper’s deposit with a deposit of their own of equal size. If either party reneges on the agreement, the defaulting party loses their deposit to the more trustworthy party. However, if both parties uphold their side of the deal, the deposits are returned in full to both sides. Under this booking mechanism, we study the carrier’s sequential online booking problem, which gives rise to a new class of revenue management problems with overbooking and no-show behavior that share only superficial commonalities with existing frameworks. In the third chapter, I study the operational decisions involved in opaque selling—a practically motivated mechanism employed by platforms such as Priceline.com. Under this mechanism, customers are offered an opaque option, which provides them with one item from a predefined bundle without revealing which specific item they will receive at the time of purchase. In exchange for this uncertainty, customers receive a discounted price, allowing the retailer to gain greater flexibility in managing inventory levels. This work contributes to the growing literature on choice-based online matching by extending the classical model to incorporate this additional decision lever. Customers who choose the opaque product delegate the final allocation decision to the platform, introducing consumer-side uncertainty that must be accounted for in the underlying choice model. This trade-off between allocation flexibility and customer uncertainty presents new challenges in dynamic assortment and pricing. I rigorously formulate the problem and propose tractable algorithms that address these challenges, providing both theoretical performance guarantees and numerical evidence of the effectiveness and robustness of the proposed policies under realistic settings
Improving Particle-Phase Nitrate Measurement in PM2.5 Filter Sampling: Evaluation of a Denuder–Nylon Filter Modification in the SPARTAN Network
Full text linkAccurate measurement of PM2.5 composition is of global importance. Evaporation-induced mass loss introduces potential bias in filter-based PM2.5 measurements, with ammonium nitrate volatilization from Teflon filters being a major contributor to negative mass artifacts. Previous studies have demonstrated that using an acid gas denuder in combination with nylon filters can help recover the nitrate mass. This study evaluates design modifications to the AirPhoton sampling station setup within the globally distributed Surface Particulate Matter Network (SPARTAN) incorporating an additional acid gas denuder upstream and a nylon filter downstream of the existing Teflon filter.
Two co-located AirPhoton Sampling stations were installed on the roof of an academic building at Washington University in St. Louis. One of the stations used the design modifications and one did not. The design modifications were also installed in SPARTAN stations worldwide. The post sampling analysis for the Teflon filters remained unchanged. Ion Chromatography (IC) was used for laboratory measurement of nitrate on nylon filters. Adding the design modification increased nitrate and ammonium collection in St. Louis by an average of 60% and 4% during the spring 2024. The design modifications showed no significant impact on the Teflon filter PM2.5 mass and the major ionic species composition.
Global deployment of the denuder–nylon modification revealed substantial variability in nitrate partitioning, with higher nitrate fractional recovery on nylon filters more common in warmer climates and during summer months. Temperature emerged as the dominant factor influencing nitrate fraction on nylon filters, while correlations with relative humidity, daily fluctuations, and aerosol chemical characteristics—including cation-to-anion balance and dust composition—were weak.
Post-sampling nitrate stability in St. Louis was assessed using paired filters with and without immediate refrigeration. Most measurements fell within detection bounds, indicating limited evaporative loss under the observed conditions. Teflon-associated nitrate showed the most evidence of loss, while PM2.5 mass remained largely stable with only minor deviations. The findings suggest that under moderate climate and nitrate conditions, post-sampling volatilization is a minor contributor to overall measurement uncertainty, though losses may be different in warmer or more polluted environments
Older Adults and Financial Exploitation
Full text linkThe National Adult Protective Services Association defines elder financial exploitation as misuse, mishandling, or exploitation of an older adult’s property, possession, or assets. These actions often occur without consent and are typically carried out through false pretense, undue influence, coercion, or manipulation
Structure and function of type 1 pilus assembly machinery in uropathogenic Escherichia coli
Full text linkGram-negative bacteria produce chaperone-usher pathway (CUP) pili, which are extracellular adhesive protein fibers critical for host-pathogen interactions. An outer-membrane usher protein, together with a periplasmic chaperone, assembles thousands of pilin subunits into a pilus fiber. Each pilus is tipped by an adhesin that mediates host and tissue tropisms by binding to receptors with stereochemical specificity. Type 1 pili expressed by uropathogenic Escherichia coli (UPEC) are essential virulence factors in urinary tract infections (UTIs) and facilitate attachment to mannosylated glycoproteins on bladder epithelial cells. Although type 1 pili have been extensively studied, critical gaps remain in our understanding of the molecular mechanisms governing their assembly and termination at the outer membrane (OM) usher FimD, as well as in the development of antibiotic-sparing therapeutics targeting usher function. Chapter 1 provides molecular details of adhesin translocation across the outer membrane and elucidates a unique conformational state adopted by the adhesin during stepwise secretion through the usher pore. A cryo-electron microscopy (cryoEM) structure of a quaternary tip complex consisting of the usher FimD, chaperone FimC, adhesin FimH, and the tip adapter FimF reveal the usher caught in the act of secreting its cognate adhesin. Comparison with previous structures depicting the adhesin either first entering or having completely exited the usher pore reveals remarkable structural plasticity of the two-domain adhesin during translocation, and identify the “stretched-relaxed” conformation of FimH. A piliation assay demonstrated that the stretched-relaxed conformation of FimH is required for transport through the usher. These findings reveal the molecular details of adhesin translocation across the outer membrane and provide the basis for the design of rational therapeutics. Chapter 2 focuses on the development of antibiotic-sparing therapeutics targeting type 1 pili, in response to the global rise of multidrug-resistant bacteria. Fabs that bind the outer membrane usher FimD were generated with the goal of either inhibiting conformational changes required for usher function or blocking interactions between the usher and pilus subunits. Candidate Fabs were structurally characterized by cryoEM and functionally assessed for their ability to inhibit usher activity. One Fab targeting the N-terminal domain (NTD) of FimD was found to inhibit usher-mediated donor-strand exchange (DSE), a critical step in pilus assembly. This work demonstrates that Fab-mediated inhibition can effectively disrupt usher function and identifies a functionally important epitope on the outer membrane usher, providing a foundation for the development of small-molecule inhibitors. Chapter 3 investigates the molecular basis of FimI-mediated termination and anchoring of type 1 pili at the outer membrane usher FimD. A minimal termination complex was engineered that preserved native interactions between FimI and the preceding rod subunit FimA during incorporation into the FimD usher and was visualized by cryoEM. In the structure, FimH adopted a distorted orientation when directly linked to FimA, but this was corrected by reintroducing a tip adapter, suggesting that tip adapter subunits play a structural role in orienting FimH. FimI was found to bind the FimD C-terminal domain 2 (CTD2) in a manner similar to other subunits, suggesting it does not engage in unique interactions with the usher\u27s periplasmic domains during termination and anchoring. Interestingly, when preceded by FimI, FimA occupied a significantly lower position within the usher pore than the analogous subunit from a similar structure. Deletion of the elongated residues in FimI’s N-terminal extension (Nte), a conserved feature among terminating subunits, did not affect the location or orientation of FimI or FimA within the usher. These findings provide a structural framework for understanding how FimI mediates termination and anchoring of the pilus, and establish a platform for further investigation into termination and anchoring of pili at the FimD usher. Overall, the studies presented here advance our understanding of type 1 pilus assembly by revealing key conformational intermediates during adhesin secretion, identifying functionally critical epitopes on the FimD usher that could serve as targets for antibiotic-sparing therapeutics, and contributing to a comprehensive model of type 1 pilus termination. By integrating highresolution cryoEM analysis with functional assays, this work not only addresses long-standing gaps in the mechanistic understanding of CUP pilus assembly but also highlights promising new avenues for therapeutic intervention. The insights gained provide a strong foundation for the rational design of usher-targeted inhibitors and open the door to broader investigations of conserved mechanisms across diverse CUP pili systems
New Paradigms in E. coli Hemolysin Function and Pathogenesis
Full text linkBacterial pore-forming toxins are potent secreted protein effectors that disrupt cellular ion homeostasis by integrating into the plasma membrane. Among them, the RTX toxin alphahemolysin (HlyA) from uropathogenic Escherichia coli (UPEC) has been extensively studied for its hemolytic activity, broad cell tropism, and role in virulence. HlyA was identified as the first type 1 secreted substrate, with β2 integrin CD18/LFA-1 proposed as its receptor. However, its diverse mechanisms of cellular toxicity and impact on outcomes of urinary tract infection (UTI) have remained incompletely defined. During pyelonephritis, renal epithelial cells are at the front line of infection, directly contacting ascending UPEC and playing an essential role in initiating innate immune responses. Importantly, these cells are highly sensitive to HlyA cytotoxicity during in vitro exposure. Here, leveraging an updated preclinical mouse model of ascending pyelonephritis, we demonstrate that HlyA functions as a renal damage factor, exacerbating renal tubular epithelial injury in vivo. As CD18 is expressed exclusively on hematopoietic cells, we leveraged a CRISPR-Cas9 loss-ofxii function screen to uncover an alternative HlyA targeting mechanism in renal epithelial cells. We found that HlyA enters epithelial cells via clathrin-mediated endocytosis (CME) and induces cytotoxicity through a mechanism distinct from the previously proposed plasma membrane poration model. Our findings show that internalized HlyA triggers permeabilization of the endolysosome, causing rapid cytoplasmic acidification, protease release, mitochondrial dysfunction, and caspase-independent cell death. Additionally, we identify the low-density lipoprotein receptor (LDLR) as an epithelial receptor for HlyA and demonstrate that targeting of LDLRHlyA interactions via peptide biologics neutralizes HlyA toxicity to renal epithelial cells. Further, we demonstrate a sex-biased response to HlyA-induced epithelial cell death, where testosterone exposure enhances HlyA-mediated killing of renal epithelial cells through an androgen receptor-independent mechanism. Given that renal damage during pyelonephritis contributes to scarring and chronic kidney disease, our findings reveal a novel HlyA mechanism of action and suggest therapeutic targeting of LDLR-HlyA interactions to mitigate UPECassociated renal injury. Beyond the context of UTI, E. coli is among the common causes of bacterial sepsis, commonly disseminating into the bloodstream during severe pyelonephritis or gastrointestinal tract infections. Sepsis is a life-threatening condition by which infectious organisms (predominantly bacteria) trigger a dysregulated proinflammatory response followed by subsequent immunosuppression. Despite intensive research on immune modulators of septic shock, bacterial sepsis continues to have a substantial mortality rate, with current host-targeted therapeutics exhibiting limited efficacy. Here, we establish a mouse model of E. coli-induced sepsis, using the urosepsis isolate CFT073, that yields nearly 80% mortality and significant manifestations of illness in the first few days. Leveraging this model, we identify that genetic knockout of HlyA in CFT073 diminishes sepsis-induced weight loss and increases survival rates 4-fold. Together, these findings uncover a previously unrecognized intracellular mechanism of HlyA cytotoxicity, establish LDLR as a functional epithelial receptor for HlyA, and demonstrate that HlyA is a critical virulence determinant in both localized and systemic E. coli infections. Our work provides a foundation for therapeutic strategies aimed at neutralizing HlyA and other bacterial toxins to reduce renal injury and improve sepsis outcomes
Single-cell and spatial immunogenomics to dissect immune mechanisms in inflammatory bowel diseases and the central nervous system
Full text linkSingle-cell genomics, particularly single-cell RNA sequencing (scRNA-seq), enables the comprehensive profiling of gene expression at the resolution of individual cells, uncovering cellular heterogeneity, identifying rare populations, and reconstructing developmental trajectories that are obscured in bulk analyses. My studies focused on the combination of complementary single-cell analyses and experimental approaches to dissect immune cell dynamics in the gut and brain. We first examined Crohn’s like disease of the pouch (CDP), a clinically challenging condition that exclusively affects patients who underwent ileal pouch anal anastomosis (IPAA) for ulcerative colitis (UC), by performing scRNA-seq/TCR/BCR-seq of paired ileoanal pouch and pre-pouch ileum tissues of individuals with CDP and healthy controls. We identified myeloid cell as the pathogenic signaling hub for chronic inflammation of the ileoanal pouch, and that the myeloid cells in CDP represent a more terminally differentiated state compared to pouchitis. We found that Th17 cells are clonally expanded in the pouch, but not the inflamed pre-pouch ileum, suggesting that pre-pouch ileitis in CDP occurs following acute-to-chronic transition of pouch inflammation. We identified shared immune and stromal inflammatory mediators between CDP, pouchitis, Crohn’s disease, and UC. Lastly, we identified endoplasmic reticulum stress as a potential biomarker for CDP diagnosis and treatment. We followed with studying perianal fistulizing Crohn’s disease (PCD) using scRNA-seq and spatial transcriptomics. We found extensive upregulation of type II interferon responses in PCD, and further show that type II interferon and TNF signal on rectal epithelial cells and potentiate their epithelial to mesenchymal transition. We identified pathogenic Th17 cells as the chief source of IFN-g in PCD, and demonstrate their co-localization with elevated interferon signaling in the fistula tract. These findings suggest the use of systemic anti-IFNg therapy in the treatment of this highly morbid complication. We then characterized macrophage populations in the choroid plexus, a critical interface between the central nervous system and systemic circulation facing continuous immunological challenges. Through high-throughput scRNA-seq, flow cytometry, lineage tracing, and imaging techniques, we identified three phenotypically and functionally distinct macrophage populations in the homeostatic choroid plexus, characterized by differential expression of CD163, MHCII, or CD9. These subsets originate from distinct combinations of primitive and definitive hematopoietic waves, occupy discrete spatial niches, and rely on different survival factors. During neuroinflammation, these macrophages orchestrated IFN responses and chemokine production, targeting effector CD8 T cells, while microglia predominantly expressed chemokines for B cells. We also identified both conserved and unique macrophage subsets within the human choroid plexus. Finally, we developed and validated novel anti-amyloid chimeric antigen receptor astrocytes (CAR-A) targeting Alzheimer’s disease, characterized by a pathological cascade initiated by amyloid accumulation and progressing to tau-mediated neurodegeneration. We engineered CAR-A constructs and validated their in vitro functionality. In vivo studies demonstrated the efficacy of two CAR-A designs in reducing amyloid plaque burden and ameliorating related pathologies in a murine amyloidosis model. Single-nucleus analysis indicated that CAR-A treatment induced a distinctive glial response towards amyloid pathology, implicating both astrocytes and microglia in amyloid clearance. These findings provide in vivo evidence supporting CAR-A-based cell or gene therapies as viable strategies for treating Alzheimer’s disease
Mass Spectrometry-Based Probing of Nucleic Acid and Protein Higher-Order Structures: From UV-Induced G-Quadruplex Crosslinking to mRNA Footprinting and ApoE4 Footprinting
No full textThe structural complexity of nucleic acids plays a crucial role in genomic stability, gene regulation, and RNA functionality, yet studying their higher-order structures (HOS) and chemical modifications remains challenging due to their dynamic nature. This thesis employs mass spectrometry-based approaches—particularly ion mobility mass spectrometry (IM-MS) and MS-based footprinting—to probe DNA and RNA structures at the molecular level, with a focus on two systems: DNA photoadducts and messenger RNA (mRNA). Additionally, the study extends to protein systems, emphasizing the role of higher-order protein structure (HOS) in biological functions. In Chapter 2, we investigate the formation of cyclobutane pyrimidine dimers (CPDs) induced by UV irradiation in DNA, which have significant implications for skin cancer. Using IM-MS and MS/MS, we differentiate six stereoisomers of thymidine dimers (cis/syn vs. trans/anti), offering insights into their structural and ion mobility characteristics. The high resolution of IM-MS, complemented by fragmentation data from MS/MS, enables the baseline separation and precise characterization of these DNA photoproducts, advancing the understanding of UV-induced DNA damage. Building on these findings, Chapter 3 explores the application of trapped ion mobility spectrometry time-of-flight mass spectrometry (TIMS-TOF) to resolve closely related isomeric thymidine dimers with greater sensitivity and resolution. We utilize TIMS-TOF to completely resolve all six distinct isomers of thymidine dimers, enhancing our ability to characterize DNA conformational dynamics, particularly in the context of G-quadruplex (G4) structures. This data is compared and validated with existing isotopic dilution mass spectrometry (IDMS) results from Dr. Taylor\u27s lab, providing further corroboration of our findings. In Chapter 4, we transition to the study of RNA higher-order structures, specifically focusing on mRNA. Traditional biophysical techniques provide valuable global structural insights but lack site-specific resolution. In contrast, our new mass spectrometry-based chemical footprinting strategy combines backbone- and base-specific reagents to probe RNA structure. This approach, demonstrated on the PreQ1 riboswitch, reveals structural changes upon ligand binding, providing a powerful and amplification-free method for probing RNA-ligand interactions. The method holds potential for advancing RNA-based research, with applications in RNA therapeutics, including mRNA vaccines. Finally, Chapter 5 shifts focus to the structural characterization of proteins, specifically Apolipoprotein E4 (ApoE4), a protein variant associated with Alzheimer’s disease. Using fast photochemical oxidation of proteins (FPOP) and mass spectrometry-based footprinting, we examine the conformational differences between ApoE4 and its engineered variants. Our findings reveal how mutations affecting oligomerization interfaces influence solvent accessibility and lipid-binding properties, offering new insights into the structural basis of ApoE4’s role in disease. By integrating advanced mass spectrometry techniques for the structural analysis of DNA, RNA, and proteins, this dissertation provides a comprehensive framework for investigating higher-order structures across these biomolecular systems. The methodologies developed here offer transformative opportunities for understanding nucleic acid and protein dynamics and have broad implications for biomedical research, including drug design, disease understanding, and therapeutic development
Robust Embeddings of Genetics, Anatomy, and State Decoded from a Neuron’s Activity
Full text linkNeural spike trains are shaped by both extrinsic inputs and intrinsic structural constraints. While prior work has focused on how spike timing encodes externally driven variables, such as stimuli or behavior, this dissertation explores whether principles that organize neural activity in space and time—such as genetic cell type, anatomical location, and arousal state—are also embedded in the spiking output of individual neurons. I hypothesize that these organizing principles are not quenched as irrelevant variability, but rather multiplexed within the neural code itself. To test this, I attempt to decode these principles from the spike trains of neurons recorded in behaving mice, and show that, like signals of stimulus or behavior, these principles are reliably embedded in the neural code. I demonstrate that the spike timing of individual neurons carries enough information to classify: (1) genetic cell type, using a novel deep-learning architecture (LOLCAT) on datasets from the Allen Institute; (2) anatomical location, across diverse brain regions, structures, and cortical layers in datasets spanning multiple labs and behavioral paradigms; and (3) arousal state, revealing a surprisingly local scale of this presumed global signal in freely-behaving mice. These signatures are consistently observed across neurons, indicating that any individual neuron may carry this embedded information. These findings expand beyond a moment-to-moment, stimulus-and-response-centric view of the neural code, revealing that individual spike trains carry high-dimensional, temporally embedded signatures of the principles that organize the brain. The spike train reveals not just what the brain does, but what it is. In doing so, this work reframes neural coding as the simultaneous representation of both function and form—suggesting that these embedded signatures of structure and state may be foundational, though often overlooked, dimensions of the neural code
The role of the poly-N repeat in the Plasmodium falciparum histone acetyltransferase protein, PfGCN5
Full text linkP. falciparum possesses the most AT-rich genome sequenced to date, with long AAT repeats coding for asparagines (N) in intragenic regions. As a result, nearly a quarter of its proteome is composed of proteins with poly-N repeats. Since the discovery of its genome, the function of these repeats has remained as one of the biggest mysteries in the biology of this parasite. Furthermore, it has long been known that proteins with N-rich domains aggregate easily, particularly during heat stress (HS). Given that the hallmark of this disease is recurrent febrile episodes, it is not well understood how the parasite survives this stress with an aggregation-prone proteome. Only two of the 1,302 proteins containing poly-N domains in P. falciparum have been experimentally studied. In the first case, Rpn6, a non-enzymatic component of the proteasome, was found to remain stable and active when its 28-residue poly-N domain was removed, during standard and heat stress (HS) conditions, resulting in unaltered parasite growth. This study suggests that poly-N repeats may not play a functional role in the parasite. In the second case, PfHsp110, but not mammalian or yeast homologs, was able to fold the parasite’s putative CDK2-regulatory subunit (possessing an 83 N run) upon heat shock (HS), highlighting the robust specialization that parasite chaperones have developed. Recent discoveries in other organisms whose proteomes are N-rich have shed light on the numerous functions that low-complexity regions, such as poly-N domains, carry out. Mounting data indicate that this is a pervasive phenomenon in the cell, with multiple pivotal roles, including cell recovery, signaling, complex assembly, cellular organization, and transcriptional regulation. Inspired by these discoveries in biology, we decided against the odds to study the role of an imperfect repeat of 98 residues containing 81 N, in the P. falciparum histone acetyltransferase protein, GCN5 (PfGCN5). We observed that the extended low-complexity intrinsically disordered regions in the N-terminus of P. falciparum or P. vivax GCN5 are important for parasite growth. We revealed that the poly-N repeat of PfGCN5 interacts directly with the C-terminal catalytic domain of the protein following cleavage. Deletion of the poly-N repeat in PfGCN5 leads to parasite growth impairment by destabilizing the N-terminal polypeptide after cleavage from the catalytic domain. Interestingly, deletion lines are highly susceptible to stress and display a substantial decrease in histone acetylation. We confirmed PfGCN5 as the enzyme responsible for H3K9ac and newly implicated it in the acetylation of histone variants H3.3 and H4. Moreover, the interactions and localization of PfGCN5 in the context of heat stress are further explored, and the essentiality of two other poly-N repeat-containing proteins is determined. To our knowledge, this is the first study to discover a functional role for a poly-N domain in P. falciparum. Future studies should focus on the role of the poly-N repeat in the SAGA complex and its mechanistic contribution to PfGCN5 function
Investigating the Genetics and Genomics of Vein of Galen Malformations
No full textVein of Galen malformations (VOGMs) represent the most common and most severe subtype of congenital brain arteriovenous malformations, yet the underlying genetic and developmental mechanisms remain incompletely understood. This dissertation investigates the etiology of VOGMs through an integrated genomic and transcriptomic approach, combining whole-exome sequencing of 310 proband-parent trios with single-cell transcriptomic profiling of 336,326 human cerebrovascular cells. Genetic analyses identified a genome-wide significant enrichment of de novo loss-of-function variants in RASA1, which encodes the Ras suppressor p120 RasGAP (2042.5-fold enrichment; p = 4.79 × 10⁻⁷). Rare, damaging inherited variants were significantly enriched in EPHB4 (17.5-fold; p = 1.22 × 10⁻⁵), a receptor tyrosine kinase that functions in concert with p120 RasGAP to regulate endothelial signaling and vascular development. Additional candidate genes implicated in VOGM pathogenesis include ACVRL1, NOTCH1 and PTPN11, with ACVRL1 variants also observed in a multigenerational pedigree, supporting a heritable contribution in a subset of cases. Integrative transcriptomic analyses localized VOGM-associated gene expression to endothelial cells during late gestational development, defining this population as a key spatiotemporal locus of disease pathophysiology. Functional modeling in mice demonstrated that endothelial expression of a VOGM-linked EPHB4 variant (p.Phe867Leu) disrupted developmental angiogenesis and impaired arterial-capillary-venous hierarchy, but only in the context of a “second-hit” genetic background, highlighting the interplay between genetic susceptibility and developmental context. Together, these findings elucidate a genetic and mechanistic framework for VOGM pathogenesis centered on disrupted endothelial signaling. By establishing RASA1 and EPHB4 as core contributors and revealing key molecular pathways and cellular contexts, this work provides a foundation for improved genetic diagnosis, functional studies, and potential therapeutic strategies in cerebrovascular malformations