Introduction to the 5th Biometals Webinars, Managing Manganese in Bacillus subtilis, SARS‑CoV2 Nsp1 is a metal‑dependent DNA and RNA endonuclease

Abstract

Introduction to the 5th Biometals WebinarsWelcome to the 5th international webinars series For those who don't already know, these webinars have been set up by the international Biometals society and the journal Biometals with the help of Cassyni platform. • The aim is to promote research in the field of metal interactions in biology and to encourage the interdisciplinary exchange of information at international level. Manganese (Mn) in the spotlight today! Mn, just before iron in the periodic table, is importance is quite often under evaluated compare to iron! As iron Mn is essential but may be toxic due to mainly its redox properties. Mn is a key cofactor in many enzymes and vital system such as photosynthesis… where the tetranuclear manganese cofactor in photosystem 2 catalyzes the key reaction of oxygen production….but The Mn Superoxide dismutase is also a key actor…and many other enzymes contain Mn as cofactor prokaryotic, eukaryotes…and viral!!!! Intracellular or excreted welcome two speakers: John Helmann from Cornell University will be talking about Manganese homeostasis in his favorite playground Bacillus subtilis The 2nd speaker, Celia Romão will be talking about Manganese in a metal‑dependent endonuclease, not just any enzyme, but the one encoded by nsp1, one of the 16 sarsCov2 genes!Managing Manganese in Bacillus subtilisBacteria must adapt to both metal ion limitation and excess. In the model Gram-positive bacterium Bacillus subtilis, the MntR (manganese transport regulator) protein binds Mn(II), represses uptake genes, and activates two genes (mneP, mneS) encoding cation diffusion facilitator proteins that export Mn(II). Genetic studies of mutants dysregulated for Mn homeostasis (mntR) or lacking the major Mn efflux pumps (mneP mneS) revealed a class of mutations with increased expression or activity of two TerC family membrane proteins, MeeF(YceF) and MeeY(YkoY). MeeF and MeeY (metalation of exoenzymes) are Mn efflux proteins that function to support metalation of enzymes that function outside the cell membrane. Unlike MneP and MneS, the MeeF and MeeY proteins do not play a major role in resistance to elevated Mn, although mutations can increase their efflux activity. FY mutant strains, with deletions of both meeF and meeY, are slow growing due to defects in protein secretion and a reduced ability to metalate a key enzyme for cell wall synthesis, the Mn-dependent lipoteichoic acid synthase (LtaS). Consistent with a role in the metalation of Mn-requiring exoenzymes, the MeeY protein is regulated by a Mn-responsive riboswitch. We propose that MeeF and MeeY function, in part, in the co-translocational metalation of secreted enzymes. These findings highlight the challenge of properly metalating enzymes that function outside the cell and may prove relevant to studies of TerC homologs in plants and animals.SARS‑CoV2 Nsp1 is a metal‑dependent DNA and RNA endonucleaseOver recent years, we have been living under a pandemic, caused by the rapid spread of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV2). One of the major virulence factors of Coronaviruses is the Non-structural protein 1 (Nsp1), known to suppress the host cells protein translation machinery, allowing the virus to produce its own proteins, propagate and invade new cells. To unveil the molecular mechanisms of SARS-CoV2 Nsp1, we have addressed its biochemical and biophysical properties in the presence of calcium, magnesium and manganese. Our findings indicate that the protein in solution is a monomer and binds to both manganese and calcium, with high affinity. Surprisingly, our results show that SARS-CoV2 Nsp1 alone displays metal-dependent endonucleolytic activity towards both RNA and DNA, regardless of the presence of host ribosome. These results show Nsp1 as new nuclease within the coronavirus family. Furthermore, the Nsp1 double variant R124A/K125A presents no nuclease activity for RNA, although it retains activity for DNA, suggesting distinct binding sites for DNA and RNA. Thus, we present for the first time, evidence that the activities of Nsp1 are modulated by the presence of different metals, which are proposed to play an important role during viral infection. This research contributes significantly to our understanding of the mechanisms of action of Coronaviruses.