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Guo, Q., Lehmer, C., Martinez-Sanchez, A., Rudack, T., Beck, F., Hartmann, H., Perez-Berlanga, M., Frottin, F., Hipp, M.S., Hartl, F.U., Edbauer, D., Baumeister, W., and Fernandez-Busnadiego, R
Cell 2018, [Epub ahead of print].
doi: 10.1016/j.cell.2017.12.030

In Situ Structure of Neuronal C9orf72 Poly-GA Aggregates Reveals Proteasome Recruitment

Protein aggregation and dysfunction of the ubiquitin-proteasome system are hallmarks of many neurodegenerative diseases. Here, we address the elusive link between these phenomena by employing cryo-electron tomography to dissect the molecular architecture of protein aggregates within intact neurons at high resolution. We focus on the poly-Gly-Ala (poly-GA) aggregates resulting from aberrant translation of an expanded GGGGCC repeat in C9orf72, the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia. We find that poly-GA aggregates consist of densely packed twisted ribbons that recruit numerous 26S proteasome complexes, while other macromolecules are largely excluded. Proximity to poly-GA ribbons stabilizes a transient substrate-processing conformation of the 26S proteasome, suggesting stalled degradation. Thus, poly-GA aggregates may compromise neuronal proteostasis by driving the accumulation and functional impairment of a large fraction of cellular proteasomes.


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Prytuliak, R., Pfeiffer, F., and Habermann, B.H.
BMC Bioinformatics 2018, 19, 24.
doi: 10.1186/s12859-018-2020-x

SLALOM, a flexible method for the identification and statistical analysis of overlapping continuous sequence elements in sequence- and time-series data.

Protein or nucleic acid sequences contain a multitude of associated annotations representing continuous sequence elements (CSEs). Comparing these CSEs is needed, whenever we want to match identical annotations or integrate distinctive ones. Currently, there is no ready-to-use software available that provides comprehensive statistical readout for comparing two annotations of the same type with each other, which can be adapted to the application logic of the scientific question.
We have developed a method, SLALOM (for StatisticaL Analysis of Locus Overlap Method), to perform comparative analysis of sequence annotations in a highly flexible way. SLALOM implements six major operation modes and a number of additional options that can answer a variety of statistical questions about a pair of input annotations of a given sequence collection. We demonstrate the results of SLALOM on three different examples from biology and economics and compare our method to already existing software. We discuss the importance of carefully choosing the application logic to address specific scientific questions.
SLALOM is a highly versatile, command-line based method for comparing annotations in a collection of sequences, with a statistical read-out for performance evaluation and benchmarking of predictors and gene annotation pipelines. Abstraction from sequence content even allows SLALOM to compare other kinds of positional data including, for example, data coming from time series.


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Yan X, Shi Q, Bracher A, Miličić G, Singh AK, Hartl FU, Hayer-Hartl M. (IMPRS-LS students are in bold)
Cell 2017, [Epub ahead of print].
doi: 10.1016/j.cell.2017.12.010

GroEL Ring Separation and Exchange in the Chaperonin Reaction

The bacterial chaperonin GroEL and its cofactor, GroES, form a nano-cage for a single molecule of substrate protein (SP) to fold in isolation. GroEL and GroES undergo an ATP-regulated interaction cycle to close and open the folding cage. GroEL consists of two heptameric rings stacked back to back. Here, we show that GroEL undergoes transient ring separation, resulting in ring exchange between complexes. Ring separation occurs upon ATP-binding to the trans ring of the asymmetric GroEL:7ADP:GroES complex in the presence or absence of SP and is a consequence of inter-ring negative allostery. We find that a GroEL mutant unable to perform ring separation is folding active but populates symmetric GroEL:GroES2 complexes, where both GroEL rings function simultaneously rather than sequentially. As a consequence, SP binding and release from the folding chamber is inefficient, and E. coli growth is impaired. We suggest that transient ring separation is an integral part of the chaperonin mechanism.


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Rosam, M., Krader, D., Nickels, C., Hochmair, J., Back, K.C., Agam, G., Barth, A., Zeymer, C., Hendrix, J., Schneider, M., Antes, I., Reinstein, J., Lamb, D.C., and Buchner, J.
Nat Struct Mol Biol 2018, 25, 90-100.
doi: 10.1038/s41594-017-0012-6

Bap (Sil1) regulates the molecular chaperone BiP by coupling release of nucleotide and substrate

BiP is the endoplasmic member of the Hsp70 family. BiP is regulated by several co-chaperones including the nucleotide-exchange factor (NEF) Bap (Sil1 in yeast). Bap is a two-domain protein. The interaction of the Bap C-terminal domain with the BiP ATPase domain is sufficient for its weak NEF activity. However, stimulation of the BiP ATPase activity requires full-length Bap, suggesting a complex interplay of these two factors. Here, single-molecule FRET experiments with mammalian proteins reveal that Bap affects the conformation of both BiP domains, including the lid subdomain, which is important for substrate binding. The largely unstructured Bap N-terminal domain promotes the substrate release from BiP. Thus, Bap is a conformational regulator affecting both nucleotide and substrate interactions. The preferential interaction with BiP in its ADP state places Bap at a late stage of the chaperone cycle, in which it coordinates release of substrate and ADP, thereby resetting BiP for ATP and substrate binding.


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Park, D.I., Stambuk, J., Razdorov, G., Pucic-Bakovic, M., Martins-de-Souza, D., Lauc, G., and Turck, C.W.
Sci Rep 2018, 8, 179.
doi: 10.1038/s41598-017-17500-0

Blood plasma/IgG N-glycome biosignatures associated with major depressive disorder symptom severity and the antidepressant response

While N-linked glycosylation has been extensively studied in the context of inflammatory and metabolic disorders, its relationship with major depressive disorder (MDD) and antidepressant treatment response has not been investigated. In our exploratory study, we analysed N-glycan profiles in blood plasma samples collected from MDD patients (n = 18) and found gender-dependent correlations with severity of depressive symptoms prior to initiating antidepressant treatment. In addition, several N-glycosylation traits showed gender-dependent associations with clinical antidepressant response. Follow up proteomics analysis in peripheral blood mononuclear cells (PBMCs) collected from MDD patients (n = 20) identified baseline and post-antidepressant treatment pathway differences between responder and non-responder patients. Reactome data analysis further delineated potential biological reaction differences between responder and non-responder patients. Our preliminary results suggest that specific glycosylation traits are associated with depressive symptom severity and antidepressant response and may be of use as biomarkers.


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Mangal, S., Sacher, J., Kim, T., Osorio, D.S., Motegi, F., Carvalho, A.X., Oegema, K., and Zanin, E.
J Cell Biol, 2018, [Epub ahead of print].
doi: 10.1083/jcb.201706021

TPXL-1 activates Aurora A to clear contractile ring components from the polar cortex during cytokinesis

During cytokinesis, a signal from the central spindle that forms between the separating anaphase chromosomes promotes the accumulation of contractile ring components at the cell equator, while a signal from the centrosomal microtubule asters inhibits accumulation of contractile ring components at the cell poles. However, the molecular identity of the inhibitory signal has remained unknown. To identify molecular components of the aster-based inhibitory signal, we developed a means to monitor the removal of contractile ring proteins from the polar cortex after anaphase onset. Using this assay, we show that polar clearing is an active process that requires activation of Aurora A kinase by TPXL-1. TPXL-1 concentrates on astral microtubules coincident with polar clearing in anaphase, and its ability to recruit Aurora A and activate its kinase activity are essential for clearing. In summary, our data identify Aurora A kinase as an aster-based inhibitory signal that restricts contractile ring components to the cell equator during cytokinesis.


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Scherr, M.J., Safaric, B., and Duderstadt, K.E.
Bioessays, 2017, [Epub ahead of print].
doi: 10.1002/bies.201700159

Noise in the Machine: Alternative Pathway Sampling is the Rule During DNA Replication

The astonishing efficiency and accuracy of DNA replication has long suggested that refined rules enforce a single highly reproducible sequence of molecular events during the process. This view was solidified by early demonstrations that DNA unwinding and synthesis are coupled within a stable molecular factory, known as the replisome, which consists of conserved components that each play unique and complementary roles. However, recent single-molecule observations of replisome dynamics have begun to challenge this view, revealing that replication may not be defined by a uniform sequence of events. Instead, multiple exchange pathways, pauses, and DNA loop types appear to dominate replisome function. These observations suggest we must rethink our fundamental assumptions and acknowledge that each replication cycle may involve sampling of alternative, sometimes parallel, pathways. Here, we review our current mechanistic understanding of DNA replication while highlighting findings that exemplify multi-pathway aspects of replisome function and considering the broader implications.


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Glock P., Broichhagen J., Kretschmer S., Blumhardt P., Mücksch J., Trauner D., Schwille P.
(IMPRS-LS students are in bold)
Angew Chem Int Ed Engl, 2017, [Epub ahead of print].
doi: 10.1002/anie.201712002

Optical control of protein pattern formation

Patterns formed by protein reactions and diffusion are the foundation for many phenomena in biology. Yet, the experimental study of reaction-diffusion (R-D) systems has so far been dominated by chemical oscillators, for which many manipulation tools are available. Here, we developed a photoswitch for the Min system of Escherichia coli, a versatile biological in vitro R-D system consisting of the antagonistic proteins MinD and MinE. A MinE-derived peptide of 19 amino acids is covalently modified with a photoisomerizable crosslinker based on azobenzene to externally control peptide-mediated depletion of MinD from the membrane. In addition to providing an on-off switch for pattern formation, we achieve frequency-locked entrainment with a precise 2D spatial memory, allowing new insights into Min protein action on the membrane. Taken together, we provide a tool to externally control protein patterns formed by self-organization.


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Vincenz-Donnelly, L., Holthusen, H., Korner, R., Hansen, E.C., Presto, J., Johansson, J., Sawarkar, R., Hartl, F.U., and Hipp, M.S.
EMBO J, 2017, [Epub ahead of print].
doi 10.15252/embj.201695841

High capacity of the endoplasmic reticulum to prevent secretion and aggregation of amyloidogenic proteins

Protein aggregation is associated with neurodegeneration and various other pathologies. How specific cellular environments modulate the aggregation of disease proteins is not well understood. Here, we investigated how the endoplasmic reticulum (ER) quality control system handles β-sheet proteins that were designed de novo to form amyloid-like fibrils. While these proteins undergo toxic aggregation in the cytosol, we find that targeting them to the ER (ER-β) strongly reduces their toxicity. ER-β is retained within the ER in a soluble, polymeric state, despite reaching very high concentrations exceeding those of ER-resident molecular chaperones. ER-β is not removed by ER-associated degradation (ERAD) but interferes with ERAD of other proteins. These findings demonstrate a remarkable capacity of the ER to prevent the formation of insoluble β-aggregates and the secretion of potentially toxic protein species. Our results also suggest a generic mechanism by which proteins with exposed β-sheet structure in the ER interfere with proteostasis.


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Albert, S., Schaffer, M., Beck, F., Mosalaganti, S., Asano, S., Thomas, H.F., Plitzko, J.M., Beck, M., Baumeister, W., and Engel, B.D.
Proc Natl Acad Sci USA, 2017, [Epub ahead of print].
doi: 10.1073/pnas.1716305114

Proteasomes tether to two distinct sites at the nuclear pore complex.

The partitioning of cellular components between the nucleus and cytoplasm is the defining feature of eukaryotic life. The nuclear pore complex (NPC) selectively gates the transport of macromolecules between these compartments, but it is unknown whether surveillance mechanisms exist to reinforce this function. By leveraging in situ cryo-electron tomography to image the native cellular environment of Chlamydomonas reinhardtii, we observed that nuclear 26S proteasomes crowd around NPCs. Through a combination of subtomogram averaging and nanometer-precision localization, we identified two classes of proteasomes tethered via their Rpn9 subunits to two specific NPC locations: binding sites on the NPC basket that reflect its eightfold symmetry and more abundant binding sites at the inner nuclear membrane that encircle the NPC. These basket-tethered and membrane-tethered proteasomes, which have similar substrate-processing state frequencies as proteasomes elsewhere in the cell, are ideally positioned to regulate transcription and perform quality control of both soluble and membrane proteins transiting the NPC.