Faculty Bibliography

The human sodium-citrate cotransporter NaCT imports various tri- and di-carboxylates into the cell as TCA cycle intermediates. This substrate uptake process is driven by an inward sodium gradient. The protein is a member of the divalent anion-sodium symporter (DASS) family. Whereas extensive biochemical and structural studies have been carried out for NaCT, how the substrate binding and translocation is coupled to the sodium gradient remains unclear. Here using single particle cryo-electron microscopy, we determined the structures of the human NaCT protein in three states: sodium-free, in the presence of sodium, and bound to sodium and a substrate mimicking inhibitor. These structures suggest a simultaneous binding mechanism for sodium-substrate coupling, distinct from the sequential binding, conformational selection mechanism previously observed for the bacterial DASS protein VcINDY.

Complex human diseases exhibit substantial clinical heterogeneity driven by poorly understood molecular mechanisms, while many also lack sufficient molecular and omics data for mechanistic investigation, hindering therapeutic development. We introduce PiMInfer, a "phenotype-to-mechanism" framework that leveraged largely available real-world clinical data-based deep phenotypic characterizations with a biomedical knowledge graph approach to resolve disease clinical heterogeneity into phenotype-informed molecular modules, thereby accelerating therapeutic target discovery. We applied PiMInfer to investigate Hidradenitis Suppurativa (HS), an autoimmune skin disease with poorly understood pathogenesis and limited treatment options. PiMInfer identified a coherent, phenotype-informed HS gene module (PiHSM) and functional endotypes, which were validated using multimodal evidence. In silico drug repurposing using PiHSM prioritized Carfilzomib, targeting the immunoproteasome subunit PSMB9, essential for MHC Class I antigen presentation. Preclinical testing using human patient lesional skin explants confirmed its anti-inflammatory activity and demonstrated a significant downregulation of IFN-γ, IL-17, and mTOR signaling pathways within HS lesional microenvironment through single-cell RNA sequencing. PiHSM-based network predictions further suggest a potential enhanced efficacy of combining Carfilzomib with approved HS agents. Collectively, PiMInfer provides a scalable framework that bridges real-world phenome-wide comorbid associations to mechanism-anchored therapeutic discovery, enabling a paradigm shift in precision medicine approaches for complex diseases with limited molecular characterization and in need of better therapeutic strategies.

VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic) is a severe adult-onset inflammatory disease caused by somatic mutations that reduce cytoplasmic activity of UBA1, the primary initiating enzyme for ubiquitylation. How this hypomorphic state drives cell-intrinsic immune activation in mature myeloid cells is unknown. Using unbiased multi-omic, biochemical, and cell biological analyses of model systems and patient-derived cells, we show that loss of cytoplasmic UBA1 activity convergently disrupts endoplasmic reticulum-associated degradation (ERAD) and mitochondrial homeostasis. ERAD failure arises from preferential under-charging of ERAD E2 enzymes, explaining hallmark VEXAS features, including ER-derived vacuoles and unfolded protein response activation, and promotes accumulation of the ERAD substrate STING. Simultaneously, mitochondrial dysfunction drives cytosolic leakage of mitochondrial DNA, inducing cGAS-dependent STING signaling and inflammatory cytokine production. STING inhibition or reversal of mitochondrial DNA leakage resolves multi-cytokine inflammation in VEXAS models and patient myeloid cells, establishing the cGAS-STING pathway as a therapeutically actionable vulnerability.

Age is the greatest risk factor for mortality and morbidity. We previously reported premature ageing in PKP2 arrhythmogenic cardiomyopathy (PKP2-ACM). However, depiction of cardiomyocyte molecular anatomy in premature ageing remains incomplete. Furthermore, the relationship between myocyte premature ageing and the molecular components of electrical homeostasis in PKP2-deficient hearts remains understudied. Therefore, we aimed to identify molecular anatomical changes of premature ageing in PKP2-deficient adult cardiomyocytes, the corresponding gene cohort and its imprint in the human electrocardiogram (ECG). We used a murine model of cardiomyocyte-specific PKP2 knockout. Molecular anatomy was resolved by expansion and structured illumination microscopy, allowing 3D-nanometric visualization, and by serial block-face scanning electron microscopy. We used omics databases to distill a subset of genes related to ageing and PKP2 deficiency containing single nucleotide polymorphisms (SNPs) with an ECG-relevant genomic signature. We found increased DNA damage and reduced abundance of transcriptionally repressed heterochromatin at the lamin-associated domain (LAD), and disrupted mitochondria ultrastructure at the intercalated disc. Genes dysregulated in ageing hearts and PKP2-deficient cells were distilled to identify those in the PKP2 human left ventricular gene network (by GTEx) that contain SNPs with an imprint in the human ECG (based on ECG GWAS). These findings indicate that in PKP2 deficiency, transcriptionally repressed heterochromatin in the LAD is vulnerable to damage, and erasing of otherwise transcriptionally inaccessible genomic regions, consistent with premature cellular ageing and transcriptional reprogramming. Multi-omics analysis indicates that reduced PKP2 expression accelerates cardiomyocyte ageing, and the age-related gene network contributes to dysfunction of cell metabolism and arrhythmia risk in PKP2-ACM. KEY POINTS: We used advanced imaging technology to unveil DNA damage and loss of heterochromatin architecture in the lamin-associated domain (LAD), and mitochondrial remodelling at the intercalated disc, as features of premature myocyte ageing in adult PKP2-deficient murine cardiomyocytes. Multi-omics analysis identified a network of genes dysregulated by ageing and by PKP2 deficiency. Cross-correlation with the ECG GWAS database found a subset of single nucleotide polymorphisms with an imprint in the human electrocardiogram. These results provide novel insight into the mechanisms of transcriptional remodelling and take an initial step toward a better understanding of the oligogenic bases of arrhythmia risk in PKP2-deficient hearts.

Autotomy is a unique phenotype whereby an animal sheds a body part to escape predation^1-3. The timing and location of autotomy are tightly regulated by preformed planes of weakness (aka fracture planes) which facilitate tissue loss. While autotomy is often followed by regeneration, these phenotypes are rarely reported in mammals^4-9. A notable exception are spiny mice (Acomys) which exhibit skin autotomy and more remarkably, complete tissue regeneration^10-14. Presently, mechanisms underlying autotomy and complete regeneration in Acomys skin remain elusive. Here, we report the discovery of a honeycomb-like fracture lattice in Acomys skin whose design directs tissue destruction but also facilitates regenerative healing. Unlike the single continuous surface of a fracture plane, this fracture lattice consists of a three-dimensional array of hexagonal units whose boundaries guide tissue breakage. Moreover, we identify collagen VI as the main constituent of the fracture lattice and find that it is distinctly arranged to initiate fracturing and propagation of skin tearing. By preconditioning the tissue for autotomy, the fracture lattice dampens the damage-induced inflammatory response but also upregulates a pro-regenerative gene signature, accelerating skin appendage regeneration. Lastly, we discovered the key role of spiny hairs in fracture lattice formation, as inhibiting their development leads to abnormal pattern formation and changes in skin fracture mechanics. Our results present a novel example of a uniquely evolved structural adaptation in mammalian skin that links tissue patterning, autotomy and regeneration. We expect that the application of a modular compartment structure to artificial skin and other organ engineering may enhance resilience to injury and facilitate efficient regeneration.

Members of the Mammalian Cell Entry (MCE) superfamily interact with other proteins to form diverse architectures for the transport of hydrophobic molecules across the cell envelope in Gram-negative bacteria. Some of these trans-envelope MCE protein complexes include a PqiC-like outer membrane (OM) lipoprotein component. The best-studied member of this group of OM lipoproteins is E. coli PqiC, from the PqiABC system, which can form an octameric ring. How PqiC-like lipoproteins interact with their MCE protein binding partners to facilitate transport is not well understood. Here we report the cryo-electron microscopy structures of Pseudomonas aeruginosa PA3214, a homolog of PqiC, in the context of the full MCE transport PA3211-PA3214 system. Our structure provides insight into the biological assembly of the lipoprotein and interactions with its binding partner, MCE protein PA3213. We utilize deep mutational scanning to identify functionally important sites in E. coli PqiC in an unbiased manner. Through phenotypic and biochemical experiments, we characterize the interactions of the lipoproteins PqiC and PA3214 with their associated MCE proteins PqiB and PA3213, thus providing a model for how some MCE proteins employ a C-terminal peptide to mediate key interactions with their cognate lipoproteins at the OM.

Glucagon-like peptide-1 receptor agonists (GLP-1 RAs) use in children is rapidly expanding, though in contrast to adults, morphologic data on dermatologic adverse events (AEs) in pediatric patients remain sparse, underscoring the need for further focused investigation. We review and synthesize the existing literature on dermatologic AEs of GLP-1 RAs in children in comparison to adults, to summarize current evidence, identify knowledge gaps, and highlight areas for future research. Available reports suggest that rash, urticaria, and alopecia may occur with greater frequency in children on injectable GLP-1 RAs compared with adults, while facial lipodystrophy has been more often described in adults. However, pediatric dermatologic AE data are not as robust as adult data, reducing the ability for direct comparisons. Based on this initial investigation, dermatologic AEs of GLP-1 RAs are seemingly under-described in children and may have important differences compared with adult dermatologic AEs.

Temporal patterning is an evolutionarily conserved mechanism to produce neuronal and glial diversity from common cells of origin during neurodevelopment. This process is controlled by a series of temporal transcription factors that are transiently expressed and drive the sequential production of specific progeny subtypes. Intermediate neural progenitors (INPs) and optic lobe neural stem cells (OL NSCs) share striking similarities in temporal factor expression despite divergent cells of origin. Tailless (Tll) is a terminal temporal factor in the visual system, in OL NSCs. Tll expression coincides with the termination of neurogenesis and onset of gliogenesis. Here, we report that Tll also acts as a terminal factor in INPs, demonstrating functional conservation. Tll expression is activated by the preceding temporal factor, Scarecrow, and represses odd-paired and hamlet. tll also plays a partial role in promoting gliogenesis in gliogenic NSCs. We performed genome-wide binding analysis of Tll in the OL NSCs and INPs by Targeted DamID, revealing both conserved and divergent targets, reflecting differences in regulatory outcomes. We show that temporal patterning mechanisms are conserved between different brain regions, whilst facilitating lineage-specific outputs.

The divalent anion sodium symporter (DASS) family mediates the uptake of Krebs cycle intermediates and sulfate and influences adiposity, insulin resistance, and metabolism in mammals. While Na⁺:substrate stoichiometry is known for several DASS transporters, the location of key Na⁺-binding sites remains elusive; important information for understanding the mechanism. In VcINDY, a bacterial DASS protein, we visualized a nonprotein cryo-electron microscopy (cryo-EM) density in the middle of the transport domain. Its size and coordination suggest that it may represent either a third Na⁺ ion or a structural water molecule. Using a combination of in vitro binding and transport assays, cryo-EM structural determination, and molecular dynamic simulations, we show that the density is not a Na⁺ ion. Instead, the data indicate that the density likely represents a structural water molecule critical for transport domain integrity. Sequence and structural similarities suggest this feature may be conserved across human DASS transporters such as NaCT and NaDC3.

Alopecia areata (AA) is an autoimmune hair disease with variable presentations necessitating individualized, stepwise management given patient age, extent of involvement, comorbidities, and treatment tolerability. Historically, severity was defined by percentage of scalp hair loss using the Severity of Alopecia Tool (SALT), with SALT≥50 defining severe disease and systemic therapy eligibility. Recently, severity assessment has expanded beyond scalp involvement with the AA Scale for Clinical Use (AASc), incorporating eyebrow/eyelash involvement, psychosocial impairment, and treatment response to better capture disease burden. First-line therapy for mild-to-moderate AA (SALT<50) includes topical or intralesional corticosteroids, often with topical or oral minoxidil. Observation may be appropriate in select cases given potential for spontaneous regrowth. FDA approval of oral Janus kinase inhibitors (JAKis) for severe AA (SALT≥50) has changed the therapeutic landscape. Selection among baricitinib, ritlecitinib, and deuruxolitinib is guided by patient age, safety considerations, comorbidities, laboratory monitoring, and insurance coverage. Switching between JAKis may still be beneficial after nonresponse to one agent. Adjunctive and alternative therapies include topical immunotherapy, systemic immunosuppressants, emerging biologic/targeted therapies, and procedural modalities, such as platelet-rich plasma, laser- and light-based therapies, and microneedling. This review synthesizes current evidence on AA treatment and offers a practical, contemporary framework for AA management.