Molecular Dynamics Simulations of Influenza A Virus NS1 Reveal a Remarkably Stable RNA-Binding Domain Harboring Promising Druggable Pockets.

The non-structural protein NS1 of influenza A viruses is considered to be the major antagonist of the interferon system and antiviral defenses of the cell. It could therefore represent a suitable target for novel antiviral strategies. As a first step towards the identification of small compounds targeting NS1, we here investigated the druggable potential of its RNA-binding domain since this domain is essential to the biological activities of NS1. We explored the flexibility of the full-length protein by running molecular dynamics simulations on one of its published crystal structures. While the RNA-binding domain structure was remarkably stable along the simulations, we identified a flexible site at the two extremities of the "groove" that is delimited by the antiparallel α-helices that make up its RNA-binding interface. This groove region is able to form potential binding pockets, which, in 60% of the conformations, meet the druggability criteria. We characterized these pockets and identified the residues that contribute to their druggability. All the residues involved in the druggable pockets are essential at the same time to the stability of the RNA-binding domain and to the biological activities of NS1. They are also strictly conserved across the large sequence diversity of NS1, emphasizing the robustness of this search towards the identification of broadly active NS1-targeting compounds.

Murine Bone Marrow Mesenchymal Stromal Cells Respond Efficiently to Oxidative Stress Despite the Low Level of Heme Oxygenases 1 and 2.

AIMS: Mesenchymal stromal cells (MSCs) are heterogeneous cells from adult tissues that are able to differentiate in vitro into adipocytes, osteoblasts, or chondrocytes. Such cells are widely studied in regenerative medicine. However, the success of cellular therapy depends on the cell survival. Heme oxygenase-1 (HO-1, encoded by the Hmox1 gene), an enzyme converting heme to biliverdin, carbon monoxide, and Fe2+, is cytoprotective and can affect stem cell performance. Therefore, our study aimed at assessing whether Hmox1 is critical for survival and functions of murine bone marrow MSCs. RESULTS: Both MSC Hmox1+/+ and Hmox1-/- showed similar phenotype, differentiation capacities, and production of cytokines or growth factors. Hmox1+/+ and Hmox1-/- cells showed similar survival in response to 50 µmol/L hemin even in increased glucose concentration, conditions that were unfavorable for Hmox1-/- bone marrow-derived proangiogenic cells (BDMC). Hmox1+/+ MSCs but not fibroblasts retained low ROS levels even after prolonged incubation with 50 µmol/L hemin, although both cell types have a comparable Hmox1 expression and similarly increase its levels in response to hemin. MSCs Hmox1-/- treated with hemin efficiently induced expression of a vast panel of antioxidant genes, especially enzymes of the glutathione pathway. Innovation and Conclusion: Hmox1 overexpression is a popular strategy to enhance viability and performance of MSCs after the transplantation. However, murine MSCs Hmox1-/- do not differ from wild-type MSCs in phenotype and functions. MSC Hmox1-/- show better resistance to hemin than fibroblasts and BDMCs and rapidly react to the stress by upregulation of quintessential genes in antioxidant response. Antioxid. Redox Signal. 00, 000-000.