Faculty Bibliography

E-cadherin plays a central role in cell-cell adhesion. The ectodomains of wild-type cadherins form a crystalline-like two-dimensional lattice in cell-cell interfaces mediated by both trans (apposed cell) and cis (same cell) interactions. In addition to these extracellular forces, adhesive strength is further regulated by cytosolic phenomena involving α and β catenin-mediated interactions between cadherin and the actin cytoskeleton. Cell-cell adhesion can be further strengthened under tension through mechanisms that have not been definitively characterized in molecular detail. Here we quantitatively determine the role of the cadherin ectodomain in mechanosensing. To this end, we devise an E-cadherin-coated emulsion system, in which droplet surface tension is balanced by protein binding strength to give rise to stable areas of adhesion. To reach the honeycomb/cohesive limit, an initial emulsion compression by centrifugation facilitates E-cadherin trans binding, whereas a high protein surface concentration enables the cis-enhanced stabilization of the interface. We observe an abrupt concentration dependence on recruitment into adhesions of constant crystalline density, reminiscent of a first-order phase transition. Removing the lateral cis interaction with a "cis mutant" shifts this transition to higher surface densities leading to denser, yet weaker adhesions. In both proteins, the stabilization of progressively larger areas of deformation is consistent with single-molecule experiments that show a force-dependent lifetime enhancement in the cadherin ectodomain, which may be attributed to the "X-dimer" bond.

The low-density lipoprotein (LDL) receptor-related protein 2 (LRP2 or megalin) is representative of the phylogenetically conserved subfamily of giant LDL receptor-related proteins, which function in endocytosis and are implicated in diseases of the kidney and brain. Here, we report high-resolution cryoelectron microscopy structures of LRP2 isolated from mouse kidney, at extracellular and endosomal pH. The structures reveal LRP2 to be a molecular machine that adopts a conformation for ligand binding at the cell surface and for ligand shedding in the endosome. LRP2 forms a homodimer, the conformational transformation of which is governed by pH-sensitive sites at both homodimer and intra-protomer interfaces. A subset of LRP2 deleterious missense variants in humans appears to impair homodimer assembly. These observations lay the foundation for further understanding the function and mechanism of LDL receptors and implicate homodimerization as a conserved feature of the LRP receptor subfamily.

Accumulation of somatic hypermutation (SHM) is the primary mechanism to enhance the binding affinity of antibodies to antigens in vivo. However, the structural basis of the effects of many SHMs remains elusive. Here, we integrated atomistic molecular dynamics (MD) simulation and data mining to build a high-throughput structural bioinformatics pipeline to study the effects of individual and combination SHMs on antibody conformation, flexibility, stability, and affinity. By applying this pipeline, we characterized a common mechanism of modulation of heavy-light pairing orientation by frequent SHMs at framework positions 39_H, 91_H, 38_L, and 87_L through disruption of a conserved hydrogen-bond network. Q39L_H alone and in combination with light chain framework 4 (FWR4_L) insertions further modulated the elbow angle between variable and constant domains of many antibodies, resulting in improved binding affinity for a subset of anti-HIV-1 antibodies. Q39L_H also alleviated aggregation induced by FWR4_L insertion, suggesting remote epistasis between these SHMs. Altogether, this study provides tools and insights for understanding antibody affinity maturation and for engineering functionally improved antibodies.

In the mouse, the osteoblast-derived hormone Lipocalin-2 (LCN2) suppresses food intake and acts as a satiety signal. We show here that meal challenges increase serum LCN2 levels in persons with normal or overweight, but not in individuals with obesity. Postprandial LCN2 serum levels correlate inversely with hunger sensation in challenged subjects. We further show through brain PET scans of monkeys injected with radiolabeled recombinant human LCN2 (rh-LCN2) and autoradiography in baboon, macaque, and human brain sections, that LCN2 crosses the blood-brain barrier and localizes to the hypothalamus in primates. In addition, daily treatment of lean monkeys with rh-LCN2 decreases food intake by 21%, without overt side effects. These studies demonstrate the biology of LCN2 as a satiety factor and indicator and anorexigenic signal in primates. Failure to stimulate postprandial LCN2 in individuals with obesity may contribute to metabolic dysregulation, suggesting that LCN2 may be a novel target for obesity treatment.

OBJECTIVE/UNASSIGNED:) levels. This condition, known as nonthyroidal illness syndrome (NTIS), is associated with poor outcomes. The association of NTIS and outcomes in patients in the intensive care unit (ICU) requiring mechanical ventilation has not been well studied. This study aimed to determine the impact of NTIS on the outcomes of these patients. METHODS/UNASSIGNED:levels. Patients who died while on mechanical ventilation were assigned a VFD of 0. RESULTS/UNASSIGNED:< .001 for both mean and median VFDs). CONCLUSIONS/UNASSIGNED:due to NTIS in patients in the ICU requiring mechanical ventilation is associated with poor outcomes.

Thyroid hormone plays an important role in cardiac function. Low levels of serum triiodothyronine (T₃) due to nonthyroidal illness syndrome may have adverse effects in heart failure (HF). This study was designed to assess the ability of T₃ to predict in-hospital outcomes in patients with acute HF. In total, 137 patients without thyroid disease or treatment with drugs which affect TH levels, who were hospitalized with acute HF were prospectively enrolled and studied. TH levels were tested upon hospital admission, and outcomes were compared between patients with low (<2.3 pg/ml) and normal (≥2.3 pg/ml) free T₃ levels as well as between those with low (<0.6 ng/ml) and normal (≥0.6 ng/ml) total T₃ levels. Low free T₃ correlated with an increased length of stay in the hospital (median 11 vs 7 days, p <0.001) and higher rates of intensive care unit admission (31.8% vs 16.9%, p = 0.047), with a trend toward increased need for invasive mechanical ventilation (9.0% vs 1.4%, p = 0.056). Low total T3 correlated with an increased length of stay in the hospital (median 11 vs 7 days, p <0.001) and increased need for invasive mechanical ventilation (9.8% vs 1.3%, p = 0.045). In conclusion, low T₃ predicts worse hospital outcomes in patients with acute HF and can be useful in the risk stratification of these patients.

A patient with pyelonephritis developed multiorgan failure resulting in death. Clinical findings were consistent with multiple endocrine neoplasia type II, with bilateral pheochromocytomas identified by computed tomography scan. We hypothesize that either the infection or the administration of radiocontrast media led to a massive release of catecholamines from the pheochromocytomas. As a result, tissue perfusion was severely compromised, and multiorgan failure developed. This exceedingly rare complication of pheochromocytoma has been termed pheochromocytoma multisystem crisis.