Libicher, K., Hornberger, R., Heymann, M., and Mutschler, H.
Nat Commun, 2020, 11, 904.
In vitro self-replication and multicistronic expression of large synthetic genomes
The generation of a chemical system capable of replication and evolution is a key objective of synthetic biology. This could be achieved by in vitro reconstitution of a minimal self-sustaining central dogma consisting of DNA replication, transcription and translation. Here, we present an in vitro translation system, which enables self-encoded replication and expression of large DNA genomes under well-defined, cell-free conditions. In particular, we demonstrate self-replication of a multipartite genome of more than 116 kb encompassing the full set of Escherichia coli translation factors, all three ribosomal RNAs, an energy regeneration system, as well as RNA and DNA polymerases. Parallel to DNA replication, our system enables synthesis of at least 30 encoded translation factors, half of which are expressed in amounts equal to or greater than their respective input levels. Our optimized cell-free expression platform could provide a chassis for the generation of a partially self-replicating in vitro translation system.
Baek, K., Krist, D.T., Prabu, J.R., Hill, S., Klügel, M., Neumaier, L.-M., von Gronau, S., Kleiger, G., and Schulman, B.A.
Nature, 2020 578, 461-466.
NEDD8 nucleates a multivalent cullin-RING-UBE2D ubiquitin ligation assembly
Eukaryotic cell biology depends on cullin-RING E3 ligase (CRL)-catalysed protein ubiquitylation, which is tightly controlled by the modification of cullin with the ubiquitin-like protein NEDD8. However, how CRLs catalyse ubiquitylation, and the basis of NEDD8 activation, remain unknown. Here we report the cryo-electron microscopy structure of a chemically trapped complex that represents the ubiquitylation intermediate, in which the neddylated CRL1β-TRCP promotes the transfer of ubiquitin from the E2 ubiquitin-conjugating enzyme UBE2D to its recruited substrate, phosphorylated IκBα. NEDD8 acts as a nexus that binds disparate cullin elements and the RING-activated ubiquitin-linked UBE2D. Local structural remodelling of NEDD8 and large-scale movements of CRL domains converge to juxtapose the substrate and the ubiquitylation active site. These findings explain how a distinctive ubiquitin-like protein alters the functions of its targets, and show how numerous NEDD8-dependent interprotein interactions and conformational changes synergistically configure a catalytic CRL architecture that is both robust, to enable rapid ubiquitylation of the substrate, and fragile, to enable the subsequent functions of cullin-RING proteins.
Lee, C.-W., Wilfling, F., Ronchi, P., Allegretti, M., Mosalaganti, S., Jentsch, S., Beck, M., and Pfander, B.
Nat Cell Biol, 2020 22, 159-166.
Selective autophagy degrades nuclear pore complexes
Nuclear pore complexes (NPCs) are very large proteinaceous assemblies that consist of more than 500 individual proteins1,2. NPCs are essential for nucleocytoplasmic transport of different cellular components, and disruption of the integrity of NPCs has been linked to aging, cancer and neurodegenerative diseases3-7. However, the mechanism by which membrane-embedded NPCs are turned over is currently unknown. Here we show that, after nitrogen starvation or genetic interference with the architecture of NPCs, nucleoporins are rapidly degraded in the budding yeast Saccharomyces cerevisiae. We demonstrate that NPC turnover involves vacuolar proteases and the core autophagy machinery. Autophagic degradation is mediated by the cytoplasmically exposed Nup159, which serves as intrinsic cargo receptor and directly binds to the autophagy marker protein Atg8. Autophagic degradation of NPCs is therefore inducible, enabling the removal of individual NPCs from the nuclear envelope.
Identification of a protein complex that attracts or repels nerve cells during development
The three proteins Teneurin, Latrophilin and FLRT hold together and bring neighboring neurons into close contact, enabling the formation of synapses and the exchange of information between the cells. In the early phase of brain development, however, the interaction of the same proteins leads to the repulsion of migrating nerve cells, as researchers from the Max Planck Institute of Neurobiology and the University of Oxford have now shown. The detailed insight into the molecular guidance mechanisms of brain cells was possible due to the structural analyses of the protein complex.
Well anchored, the proteins Teneurin and FLRT are located on the surface of nerve cells. They are on the lookout for their partner protein, Latrophilin, on other neurons. When the three proteins come into contact, they interconnect and hold the membranes together. They then trigger still largely unknown signaling cascades and thus promote the formation of a synapse at this site. Teneurin and its partner proteins are known to establish these important cell contacts in the brain. Teneurin is also an evolutionary very old protein, with related proteins found in diverse organisms ranging from bacteria to worms, fruit flies and vertebrates. However, the role of these proteins during brain development, when neurons are not yet forming synapses, remained unknown.
Hergenhan S, Holtkamp S, and Scheiermann C.
J Mol Biol. 2020;S0022-2836(20)30028-0.
(*IMPRS-LS students are in bold)
Molecular interactions between components of the circadian clock and the immune system
The immune system is under control of the circadian clock. Many of the circadian rhythms observed in the immune system originate in direct interactions between components of the circadian clock and components of the immune system. The main means of circadian control over the immune system is by direct control of circadian clock proteins acting as transcription factors driving the expression or repression of immune genes. A second circadian control of immunity lies in the acetylation or methylation of histones to regulate gene transcription or inflammatory proteins. Furthermore, circadian clock proteins can engage in direct physical interactions with components of key inflammatory pathways such as members of the NFκB protein family. This regulation is transcription independent and allows the immune system to also reciprocally exert control over circadian clock function. Thus, the molecular interactions between the circadian clock and the immune system are manifold. We highlight and discuss here the recent findings with respect to the molecular mechanisms that control time-of-day dependent immunity. This review provides a structured overview focusing on the key circadian clock proteins and discusses their reciprocal interactions with the immune system.
Kuo TT, and Ladurner AG.
Front Genet. 2019;10:1210.
Exploiting the Circadian Clock for Improved Cancer Therapy: Perspective From a Cell Biologist
Since the discovery of the biological clock, the concept of treating cancer according to biological rhythms, here termed cancer chronotherapy, has rapidly evolved. Its fundamental aim is to improve the efficacy of drugs and to minimize adverse effects by administering chemotherapeutic drugs at the appropriate time-of-day. In the last two decades, several experimental and clinical studies have reported positive associations between the circadian clock and drug response in cancer patients. However, the lack of mechanistic insights into critical, deterministic clock-controlled genetic, and metabolic variations between and within individual cancer patients continue to cast a shadow on the potential benefits cancer chronotherapy may provide. Here, we provide first a simplified overview on our biological clocks and how our life-style induces complex biochemical reactions and genetic interactions. Next, we summarize how these reactions directly and indirectly modulate the effectiveness and toxicity of oncological drug treatments. Since cytotoxic chemotherapy represents the most common and affordable of cancer treatments, a case should be made that we need to ensure these treatments are used in the best possible manner. Thus, we list current challenges and future directions toward that goal.
Swietlik JJ, Sinha A, and Meissner F.
Curr Opin Cell Biol. 2020; 63:20–30.
Dissecting intercellular signaling with mass spectrometry-based proteomics
Physiological functions depend on a coordinated interplay of numerous different cell types. Proteins serve as major signaling molecules between cells; however, their comprehensive investigation in physiologically relevant settings has remained challenging. Mass spectrometry (MS)-based shotgun proteomics is emerging as a powerful technology for the systematic analysis of protein-mediated intercellular signaling and regulated post-translational modifications. Here, we discuss recent advancements in cell biological, chemical, and biochemical MS-based approaches for the profiling of cellular messengers released by sending cells, receptors expressed on the cell surface, and their interactions. We highlight methods tailored toward the mapping of dynamic signal transduction mechanisms at cellular interfaces and approaches to dissect communication cell specifically in heterocellular systems. Thereby, MS-based proteomics contributes a unique systems biology perspective for the identification of intercellular signaling pathways deregulated in disease.