Faculty Bibliography

The LIM domain-containing proteins Pinch1/2 regulate integrin activation and cell-extracellular matrix interaction and adhesion. Here, we report that deleting Pinch1 in limb mesenchymal stem cells (MSCs) and Pinch2 globally (double knockout; dKO) in mice causes severe chondrodysplasia, while single mutant mice do not display marked defects. Pinch deletion decreases chondrocyte proliferation, accelerates cell differentiation and disrupts column formation. Pinch loss drastically reduces Smad2/3 protein expression in proliferative zone (PZ) chondrocytes and increases Runx2 and Col10a1 expression in both PZ and hypertrophic zone (HZ) chondrocytes. Pinch loss increases sclerostin and Rankl expression in HZ chondrocytes, reduces bone formation, and increases bone resorption, leading to low bone mass. In vitro studies revealed that Pinch1 and Smad2/3 colocalize in the nuclei of chondrocytes. Through its C-terminal region, Pinch1 interacts with Smad2/3 proteins. Pinch loss increases Smad2/3 ubiquitination and degradation in primary bone marrow stromal cells (BMSCs). Pinch loss reduces TGF-β-induced Smad2/3 phosphorylation and nuclear localization in primary BMSCs. Interestingly, compared to those from single mutant mice, BMSCs from dKO mice express dramatically lower protein levels of β-catenin and Yap1/Taz and display reduced osteogenic but increased adipogenic differentiation capacity. Finally, ablating Pinch1 in chondrocytes and Pinch2 globally causes severe osteopenia with subtle limb shortening. Collectively, our findings demonstrate critical roles for Pinch1/2 and a functional redundancy of both factors in the control of chondrogenesis and bone mass through distinct mechanisms.

INTRODUCTION The mechanisms underlying B cell-mediated autoimmune disease and the origin of autoreactive B cells are not well understood. Human monoclonal autoantibodies (mAbs) are valuable tools for investigating both. In this study, we used mAbs derived from patients with the autoimmune disorder, myasthenia gravis (MG). A subset of patients with MG have pathogenic autoantibodies that recognize muscle specific tyrosine kinase (MuSK). The autoantibodies of MuSK MG are predominantly of the IgG4 subclass and functionally monovalent as a result of Fab-arm exchange. OBJECTIVE To gain insight into the origin and development of these unique autoantibodies. METHODS AND RESULTS We examined MG patient-derived mAbs, their corresponding germline-encoded unmutated common ancestors (UCA) and monovalent antigen-binding fragments (Fabs) to investigate how antigen-driven affinity maturation contributes to both binding and immunopathology. Mature mAbs, their UCA counterparts and mature monovalent Fabs bound to the MuSK autoantigen and retained their pathogenic capacity. However, monovalent UCA Fabs, which still bound the autoantigen, lost their pathogenic capacity. The mature Fabs were characterized by very high affinity (sub-nanomolar) driven by a rapid on-rate and slow off-rate. However, the UCA affinity was approximately 100-fold less than the mature Fabs, which was driven by a rapid off-rate. SUMMARY/CONCLUSION These findings indicate that the autoantigen initiates the autoimmune response in MuSK MG and drives autoimmunity through the accumulation of somatic hypermutations such that monovalent IgG4 Fab-arm exchanged MG autoantibodies reach a high affinity threshold required for pathogenic capacity

Recently, the nuclear receptor farnesoid X receptor (FXR) has been considered to be a liver tumor suppressor. However, the role of FXR in liver cancer invasion and metastasis remains unclear. The results of the current study demonstrated that FXR suppressed the migratory and invasive capacities of SK-Hep-1 cells in vitro and that FXR overexpression inhibited local invasion and lung metastasis of SK-Hep-1 ×enografts in vivo. Bioinformatics analysis of the gene expression profile of SK-Hep-1 cells with different FXR levels indicated that FXR may regulate the Wnt/β-catenin pathway. Compared with controls, FXR-overexpressing SK-Hep-1 cells exhibited decreased expression of β-catenin target genes and reduced nuclear translocation of β-catenin proteins in vitro and in vivo. In conclusion, these results indicated that FXR may suppress SK-Hep-1 cell invasion and metastasis by suppressing the Wnt/β-catenin signaling pathway. The current study provided novel insight into the diagnosis and treatment of liver cancer.

Cancer genomes often harbor hundreds of somatic DNA rearrangement junctions, many of which cannot be easily classified into simple (e.g., deletion) or complex (e.g., chromothripsis) structural variant classes. Applying a novel genome graph computational paradigm to analyze the topology of junction copy number (JCN) across 2,778 tumor whole-genome sequences, we uncovered three novel complex rearrangement phenomena: pyrgo, rigma, and tyfonas. Pyrgo are "towers" of low-JCN duplications associated with early-replicating regions, superenhancers, and breast or ovarian cancers. Rigma comprise "chasms" of low-JCN deletions enriched in late-replicating fragile sites and gastrointestinal carcinomas. Tyfonas are "typhoons" of high-JCN junctions and fold-back inversions associated with expressed protein-coding fusions, breakend hypermutation, and acral, but not cutaneous, melanomas. Clustering of tumors according to genome graph-derived features identified subgroups associated with DNA repair defects and poor prognosis.

We write to introduce our novel group formed to confront some of the issues raised by the COVID-19 pandemic. Information about the group, which we named "cure COVid for Ever and for All" (RxCOVEA), its dynamic membership (changing regularly), and some of its activities-described in more technical detail for expert perusal and commentary-are available upon request.

A primary reason for the intense interest in structural biology is the fact that knowledge of structure can elucidate macromolecular functions in living organisms. Sustained effort has resulted in an impressive arsenal of tools for determining the static structures. But under physiological conditions, macromolecules undergo continuous conformational changes, a subset of which are functionally important. Techniques for capturing the continuous conformational changes underlying function are essential for further progress. Here, we present chemically-detailed conformational movies of biological function, extracted data-analytically from experimental single-particle cryo-electron microscopy (cryo-EM) snapshots of ryanodine receptor type 1 (RyR1), a calcium-activated calcium channel engaged in the binding of ligands. The functional motions differ substantially from those inferred from static structures in the nature of conformationally active structural domains, the sequence and extent of conformational motions, and the way allosteric signals are transduced within and between domains. Our approach highlights the importance of combining experiment, advanced data analysis, and molecular simulations.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Members of the ATP-binding cassette (ABC) transporter superfamily translocate a broad spectrum of chemically diverse substrates. While their eponymous ATP-binding cassette in the nucleotide-binding domains (NBDs) is highly conserved, their transmembrane domains (TMDs) forming the translocation pathway exhibit distinct folds and topologies, suggesting that during evolution, the ancient motor domains were combined with different transmembrane mechanical systems to orchestrate a variety of cellular processes. In recent years, it has become increasingly evident that the distinct TMD folds are best suited to categorize the multitude of ABC transporters. We therefore propose a new ABC transporter classification that currently comprises seven different types based on structural homology in the TMDs.