Intracellular microbial rhodopsin-based optogenetics to control metabolism and cell signaling.

Microbial rhodopsin (MRs) ion channels and pumps have become invaluable optogenetic tools for neuroscience as well as biomedical applications. Recently, MR-optogenetics expanded towards subcellular organelles opening principally new opportunities in optogenetic control of intracellular metabolism and signaling via precise manipulations of organelle ion gradients using light. This new optogenetic field expands the opportunities for basic and medical studies of cancer, cardiovascular, and metabolic disorders, providing more detailed and accurate control of cell physiology. This review summarizes recent advances in studies of the cellular metabolic processes and signaling mediated by optogenetic tools targeting mitochondria, endoplasmic reticulum (ER), lysosomes, and synaptic vesicles. Finally, we discuss perspectives of such an optogenetic approach in both fundamental and applied research.

Mirror proteorhodopsins.

Proteorhodopsins (PRs), bacterial light-driven outward proton pumps comprise the first discovered and largest family of rhodopsins, they play a significant role in life on the Earth. A big remaining mystery was that up-to-date there was no described bacterial rhodopsins pumping protons at acidic pH despite the fact that bacteria live in different pH environment. Here we describe conceptually new bacterial rhodopsins which are operating as outward proton pumps at acidic pH. A comprehensive function-structure study of a representative of a new clade of proton pumping rhodopsins which we name "mirror proteorhodopsins", from Sphingomonas paucimobilis (SpaR) shows cavity/gate architecture of the proton translocation pathway rather resembling channelrhodopsins than the known rhodopsin proton pumps. Another unique property of mirror proteorhodopsins is that proton pumping is inhibited by a millimolar concentration of zinc. We also show that mirror proteorhodopsins are extensively represented in opportunistic multidrug resistant human pathogens, plant growth-promoting and zinc solubilizing bacteria. They may be of optogenetic interest.

ATP synthase FOF1 structure, function, and structure-based drug design.

ATP synthases are unique rotatory molecular machines that supply biochemical reactions with adenosine triphosphate (ATP)-the universal "currency", which cells use for synthesis of vital molecules and sustaining life. ATP synthases of F-type (FOF1) are found embedded in bacterial cellular membrane, in thylakoid membranes of chloroplasts, and in mitochondrial inner membranes in eukaryotes. The main functions of ATP synthases are control of the ATP synthesis and transmembrane potential. Although the key subunits of the enzyme remain highly conserved, subunit composition and structural organization of ATP synthases and their assemblies are significantly different. In addition, there are hypotheses that the enzyme might be involved in the formation of the mitochondrial permeability transition pore and play a role in regulation of the cell death processes. Dysfunctions of this enzyme lead to numerous severe disorders with high fatality levels. In our review, we focus on FOF1-structure-based approach towards development of new therapies by using FOF1 structural features inherited by the representatives of this enzyme family from different taxonomy groups. We analyzed and systematized the most relevant information about the structural organization of FOF1 to discuss how this approach might help in the development of new therapies targeting ATP synthases and design tools for cellular bioenergetics control.

Low-resolution structures of modular nanotransporters shed light on their functional activity.

Modular nanotransporters (MNTs) are multifunctional chimeric polypeptides for the multistep transport of locally acting cytotoxic agents into the nuclei of cancer target cells. MNTs consist of several polypeptide domains (functional modules) for the recognition of a cell-surface internalizable receptor, pH-dependent endosomal escape and subsequent transport into the nucleus through the nuclear pores. MNTs are a promising means for cancer treatment. As has been shown previously, all of the modules of MNTs retain their functionalities. Despite their importance, there is no structural information available about these chimeric polypeptides, which hampers the creation of new MNT variants. Here, a low-resolution 3D structure of an MNT is presented which was obtained by atomic force microscopy, transmission electron microscopy and small-angle X-ray scattering coupled to size-exclusion chromatography. The data suggest that the MNT can adopt two main conformations, but in both conformations the protein N- and C-termini are distanced and do not influence each other. The change in the MNT conformation during acidification of the medium was also studied. It was shown that the fraction of the elongated conformation increases upon acidification. The results of this work will be useful for the development of MNTs that are suitable for clinical trials and possible therapeutic applications.