A range of membrane models have been developed to study components of cellular systems. Lipid vesicles or liposomes are one such artificial membrane model which mimics many properties of the biological system: they are lipid bilayers composed of one or more lipids to which other molecules can associate. Liposomes are thus ideal to study the roles of cellular lipids and their interactions with other membrane components to understand a wide range of cellular processes including membrane disruption, membrane transport and catalytic activity. Although liposomes are much simpler than cellular membranes, they are still challenging to study and a variety of complementary techniques are needed. In this review article, we consider several currently used analytical methods for spectroscopic measurements of unilamellar liposomes and their interaction with proteins and peptides. Among the variety of spectroscopic techniques seeing increasing application, we have chosen to discuss: fluorescence based techniques such as FRET (fluorescence resonance energy transfer) and FRAP (fluorescence recovery after photobleaching), that are used to identify localisation and dynamics of molecules in the membrane; circular dichroism (CD) and linear dichroism (LD) for conformational and orientation changes of proteins on membrane binding; and SERS (Surface Enhanced Raman Spectroscopy) as a rapidly developing ultrasensitive technique for site-selective molecular characterisation. The review contains brief theoretical basics of the listed techniques and recent examples of their successful applications for membrane studies.

With millions of signalling events occurring simultaneously, cells process a continuous flux of information. The genesis, processing, and regulation of information are dictated by a huge network of protein interactions. This is proven by the fact that alterations in the levels of proteins, single amino acid changes, post-translational modifications, protein products arising out of gene fusions alter the interaction landscape leading to diseases such as congenital disorders, deleterious syndromes like cancer, and crippling diseases like the neurodegenerative disorders which are often fatal. Needless to say, there is an immense effort to understand the biophysical basis of such direct interactions between any two proteins, the structure, domains, and sequence motifs involved in tethering them, their spatio-temporal regulation in cells, the structure of the network, and their eventual manipulation for intervention in diseases. In this chapter, we will deliberate on a few techniques that allow us to dissect the thermodynamic and kinetic aspects of protein interaction, how innovation has rendered some of the traditional techniques applicable for rapid analysis of multiple samples using small amounts of material. These advances coupled with automation are catching up with the genome-wide or proteome-wide studies aimed at identifying new therapeutic targets. The chapter will also summarize how some of these techniques are suited either in the standalone mode or in combination with other biophysical techniques for the drug discovery process.

Neurodegenerative diseases (NDDs), including Alzheimer's disease (AD) and Parkinson's disease (PD), affect the ageing population worldwide and while severely impairing the quality of life of millions, they also cause a massive economic burden to countries with progressively ageing populations. Parallel with the search for biomarkers for early detection and prediction, the pursuit for therapeutic approaches has become growingly intensive in recent years. Various prospective therapeutic approaches have been explored with an emphasis on early prevention and protection, including, but not limited to, gene therapy, stem cell therapy, immunotherapy and radiotherapy. Many pharmacological interventions have proved to be promising novel avenues, but successful applications are often hampered by the poor delivery of the therapeutics across the blood-brain-barrier (BBB). To overcome this challenge, nanoparticle (NP)-mediated drug delivery has been considered as a promising option, as NP-based drug delivery systems can be functionalized to target specific cell surface receptors and to achieve controlled and long-term release of therapeutics to the target tissue. The usefulness of NPs for loading and delivering of drugs has been extensively studied in the context of NDDs, and their biological efficacy has been demonstrated in numerous preclinical animal models. Efforts have also been made towards the development of NPs which can be used for targeting the BBB and various cell types in the brain. The main focus of this review is to briefly discuss the advantages of functionalized NPs as promising theranostic agents for the diagnosis and therapy of NDDs. We also summarize the results of diverse studies that specifically investigated the usage of different NPs for the treatment of NDDs, with a specific emphasis on AD and PD, and the associated pathophysiological changes. Finally, we offer perspectives on the existing challenges of using NPs as theranostic agents and possible futuristic approaches to improve them.

Influenza virus causes an acute febrile respiratory disease in humans that is commonly known as ‘flu’. Influenza virus has been around for centuries and is one of the most successful, and consequently most studied human viruses. This has generated tremendous amount of data and information, thus it is pertinent to summarise these for, particularly interdisciplinary readers. Viruses are acellular organisms and exist at the interface of living and non-living. Due to this unique characteristic, viruses require another organism, i.e. host to survive. Viruses multiply inside the host cell and are obligate intracellular pathogens, because their relationship with the host is almost always harmful to host. In mammalian cells, the life cycle of a virus, including influenza is divided into five main steps: attachment, entry, synthesis, assembly and release. To complete these steps, some viruses, e.g. influenza utilise all three parts — plasma membrane, cytoplasm and nucleus, of the cell; whereas others, e.g. SARS-CoV-2 utilise only plasma membrane and cytoplasm. Hence, viruses interact with numerous host factors to complete their life cycle, and these interactions are either exploitative or antagonistic in nature. The host factors involved in the life cycle of a virus could be divided in two broad categories — proviral and antiviral. This perspective has endeavoured to assimilate the information about the host factors which promote and suppress influenza virus infection. Furthermore, an insight into host factors that play a dual role during infection or contribute to influenza virus-host adaptation and disease severity has also been provided.

The cellular cytoskeleton self-organizes by specific monomer–monomer interactions resulting in the polymerization of filaments. While we have long thought about the role of polymerization in cytoskeleton formation, we have only begun to consider the role of condensation in cytoskeletal organization. In this review, we highlight how the interplay between polymerization and condensation leads to the formation of the cytoskeleton.

Genes under control of super-enhancers are expressed at extremely high levels and are frequently associated with nuclear speckles. Recent data suggest that the high concentration of unphosphorylated RNA polymerase II (Pol II) and Mediator recruited to super-enhancers create phase-separated condensates. Transcription initiates within or at the surface of these phase-separated droplets and the phosphorylation of Pol II, associated with transcription initiation and elongation, dissociates Pol II from these domains leading to engagement with nuclear speckles, which are enriched with RNA processing factors. The transitioning of Pol II from transcription initiation domains to RNA processing domains effectively co-ordinates transcription and processing of highly expressed RNAs which are then rapidly exported into the cytoplasm.

Enabled by its central role in the molecular networks that govern cell function, RNA has been widely used for constructing components used in biological circuits for synthetic biology. Nucleic acid nanotechnology, which exploits predictable nucleic acid interactions to implement programmable molecular systems, has seen remarkable advances in in vitro nanoscale self-assembly and molecular computation, enabling the production of complex nanostructures and DNA-based neural networks. Living cells genetically engineered to execute nucleic acid nanotechnology programs thus have outstanding potential to significantly extend the current limits of synthetic biology. This perspective discusses the recent developments and future challenges in the field of synthetic bionanotechnology. Thus far, researchers in this emerging area have implemented dozens of programmable RNA nanodevices that provide precise control over gene expression at the transcriptional and translational levels and through CRISPR/Cas effectors. Moreover, they have employed synthetic self-assembling RNA networks in engineered bacteria to carry out computations featuring up to a dozen inputs and to substantially enhance the rate of chemical synthesis. Continued advancement of the field will benefit from improved in vivo strategies for streamlining nucleic acid network synthesis and new approaches for enhancing network function. As the field matures and the complexity gap between in vitro and in vivo systems narrows, synthetic bionanotechnology promises to have diverse potential applications ranging from intracellular circuits that detect and treat disease to synthetic enzymatic pathways that efficiently produce novel drug molecules.

Mechanotransduction via the extracellular matrix (ECM)–myosin pathway is involved in determining cell morphology during development and in coupling external transient mechanical stimuli to the reorganization of the cytoskeleton. Here, we present a review on the molecular mechanisms involved in this pathway and how they influence cellular development and organization. We investigate key proteins involved in the ECM–myosin pathway and discuss how specific binding events and conformational changes under force are related to mechanical signaling. We connect these molecular mechanisms with observed morphological changes at the cellular and organism level. Finally, we propose a model encompassing the biomechanical signals along the ECM–myosin pathway and how it could be involved in cell adhesion, cell migration, and tissue architecture.

This issue of Emerging Topics in the Life Sciences highlights current areas of research in the field of archaeal biology and the following introductory editorial sets the stage by considering some of the key developments over the last four decades since the initial identification of the archaea as a unique form of life. Emerging topics from this vibrant and rapidly expanding field of research are considered and detailed further in the articles within this issue.

Sm-like (Lsm) proteins are found in all three domains of life. They are crucially involved in the RNA metabolism of prokaryotic organisms. To exert their function, they assemble into hexa- or heptameric rings and bind RNA via a conserved binding pocket for uridine stretches in the inner pore of the ring. Despite the conserved secondary structure of Lsm proteins, there are several features that lead to a structural diversification of this protein family that mediates their participation in a variety of processes related to RNA metabolism. Until recently, the cellular function of archaeal Sm-like proteins was not well understood. In this review, we discuss structural features of Lsm proteins with a strong focus on archaeal variants, reflect on the evolutionary development of archaeal Lsm proteins and present recent insights into their biological function.

An adequate response to a sudden temperature rise is crucial for cellular fitness and survival. While heat shock response (HSR) is well described in bacteria and eukaryotes, much less information is available for archaea, of which many characterized species are extremophiles thriving in habitats typified by large temperature gradients. Here, we describe known molecular aspects of archaeal heat shock proteins (HSPs) as key components of the protein homeostasis machinery and place this in a phylogenetic perspective with respect to bacterial and eukaryotic HSPs. Particular emphasis is placed on structure–function details of the archaeal thermosome, which is a major element of the HSR and of which subunit composition is altered in response to temperature changes. In contrast with the structural response, it is largely unclear how archaeal cells sense temperature fluctuations and which molecular mechanisms underlie the corresponding regulation. We frame this gap in knowledge by discussing emerging questions related to archaeal HSR and by proposing methodologies to address them. Additionally, as has been shown in bacteria and eukaryotes, HSR is expected to be relevant for the control of physiology and growth in various stress conditions beyond temperature stress. A better understanding of this essential cellular process in archaea will not only provide insights into the evolution of HSR and of its sensing and regulation, but also inspire the development of biotechnological applications, by enabling transfer of archaeal heat shock components to other biological systems and for the engineering of archaea as robust cell factories.

Archaea are phylogenetically distinct from bacteria, and some of their proteolytic systems reflect this distinction. Here, the current knowledge of archaeal proteolysis is reviewed as it relates to protein metabolism, protein homeostasis, and cellular regulation including targeted proteolysis by proteasomes associated with AAA-ATPase networks and ubiquitin-like modification. Proteases and peptidases that facilitate the recycling of peptides to amino acids as well as membrane-associated and integral membrane proteases are also reviewed.

RecJ proteins belong to the DHH superfamily of phosphoesterases that has members in all three domains of life. In bacteria, the archetypal RecJ is a 5′ → 3′ ssDNA exonuclease that functions in homologous recombination, base excision repair and mismatch repair, while in eukaryotes, the RecJ-like protein Cdc45 (which has lost its nuclease activity) is a key component of the CMG (Cdc45–MCM–GINS) complex, the replicative DNA helicase that unwinds double-stranded DNA at the replication fork. In archaea, database searching identifies genes encoding one or more RecJ family proteins in almost all sequenced genomes. Biochemical analysis has confirmed that some but not all of these proteins are components of archaeal CMG complexes and has revealed a surprising diversity in mode of action and substrate preference. In addition to this, some archaea encode catalytically inactive RecJ-like proteins, and others a mix of active and inactive proteins, with the inactive proteins being confined to structural roles only. Here, I summarise current knowledge of the structure and function of the archaeal RecJ-like proteins, focusing on similarities and differences between proteins from different archaeal species, between proteins within species and between the archaeal proteins and their bacterial and eukaryotic relatives. Models for RecJ-like function are described and key areas for further study highlighted.

Heparan sulfate (HS) is a glycosaminoglycan produced by all mammalian cells that plays important roles in physiology and various pathologies. Heparin is a highly sulfated form of HS that is used clinically as an anticoagulant. Heparin and HSs may also have therapeutic benefits for a wide variety of other indications. Cultured mammalian cells produce HS and, through genetic modification, have been used to elucidate the biosynthetic pathway. Recently, metabolic engineering has been used to produce HS from cultured mammalian cells for clinical purposes. This review describes the HS biosynthetic pathway and its manipulation through metabolic engineering to produce bioengineered HSs. We also discuss current challenges and opportunities to advance the field of HS metabolic engineering.

Electron paramagnetic resonance (EPR) spectroscopy combined with site-directed spin labelling is applicable to biomolecules and their complexes irrespective of system size and in a broad range of environments. Neither short-range nor long-range order is required to obtain structural restraints on accessibility of sites to water or oxygen, on secondary structure, and on distances between sites. Many of the experiments characterize a static ensemble obtained by shock-freezing. Compared with characterizing the dynamic ensemble at ambient temperature, analysis is simplified and information loss due to overlapping timescales of measurement and system dynamics is avoided. The necessity for labelling leads to sparse restraint sets that require integration with data from other methodologies for building models. The double electron–electron resonance experiment provides distance distributions in the nanometre range that carry information not only on the mean conformation but also on the width of the native ensemble. The distribution widths are often inconsistent with Anfinsen's concept that a sequence encodes a single native conformation defined at atomic resolution under physiological conditions.

In recent years, it has become evident that structural characterization would gain significantly in terms of biological relevance if framed within a cellular context, while still maintaining the atomic resolution. Therefore, major efforts have been devoted to developing Cellular Structural Biology approaches. In this respect, in-cell NMR can provide and has provided relevant contributions to the field, not only to investigate the structural and dynamical properties of macromolecules in solution but, even more relevant, to understand functional processes directly in living cells and the factors that modulate them, such as exogenous molecules, partner proteins, and oxidative stress. In this commentary, we review and discuss some of the main contributions to the understanding of protein structural and functional properties achieved by in-cell NMR.

Microorganisms are in constant competition for growth niches and environmental resources. In Gram-negative bacteria, contact-dependent growth inhibition (CDI) systems link the fate of one cell with its immediate neighbor through touch-dependent, receptor-mediated toxin delivery. Though discovered for their ability to confer a competitive growth advantage, CDI systems also play significant roles in intersibling cooperation, promoting both auto-aggregation and biofilm formation. In this review, we detail the mechanisms of CDI toxin delivery and consider how toxin exchange between isogenic sibling cells could regulate gene expression.

The ability of bacterial cells to synchronize their behaviour through quorum sensing (QS) regulatory networks enables bacterial populations to mount co-operative responses against competing micro-organisms and host immune defences and to adapt to environmental challenges. Since QS controls the ability of many pathogenic bacteria to cause disease, it is an attractive target for novel antibacterial agents that control infection through inhibition of virulence and by rendering biofilms more susceptible to conventional antibiotics and host clearance pathways. QS systems provide multiple druggable molecular targets for inhibitors (QSIs) that include the enzymes involved in QS signal molecule biosynthesis and the receptors involved in signal transduction. Considerable advances in our understanding of the chemical biology of QS systems and their inhibition have been made, some promising QS targets structurally characterized, QSI screens devised and inhibitors identified. However, much more work is required before any QSI ‘hits’ with the appropriate pharmacological and pharmacokinetic properties can enter human clinical trials. Indeed, the relative efficacy of QSIs alone or as prophylactics or therapeutics or as adjuvants in combination with conventional antibiotics still needs to be extensively evaluated in vivo . Particular attention must be given to the measurement of successful QSI therapy outcomes with respect to bacterial clearance, immune response and pathophysiology. Currently, our understanding of the potential of QS as a promising antibacterial target suggests that it is likely to be of value with respect to a limited number of major pathogens.