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52 Publications visible to you, out of a total of 52
ALS-linked loss of Cyclin-F function affects HSP90.

Abstract (Expand)

The founding member of the F-box protein family, Cyclin-F, serves as a substrate adaptor for the E3 ligase Skp1-Cul1-F-box (SCF)<sup>Cyclin-F</sup> which is responsible for … ubiquitination of proteins involved in cell cycle progression, DNA damage and mitotic fidelity. Missense mutations in <i>CCNF</i> encoding for Cyclin-F are associated with amyotrophic lateral sclerosis (ALS). However, it remains elusive whether <i>CCNF</i> mutations affect the substrate adaptor function of Cyclin-F and whether altered SCF<sup>Cyclin-F</sup>-mediated ubiquitination contributes to pathogenesis in <i>CCNF</i> mutation carriers. To address these questions, we set out to identify new SCF<sup>Cyclin-F</sup> targets in neuronal and ALS patient-derived cells. Mass spectrometry-based ubiquitinome profiling of <i>CCNF</i> knockout and mutant cell lines as well as Cyclin-F proximity and interaction proteomics converged on the HSP90 chaperone machinery as new substrate candidate. Biochemical analyses provided evidence for a Cyclin-F-dependent association and ubiquitination of HSP90AB1 and implied a regulatory role that could affect the binding of a number of HSP90 clients and co-factors. Together, our results point to a possible Cyclin-F loss-of-function-mediated chaperone dysregulation that might be relevant for ALS.

Authors: A. Siebert, V. Gattringer, J. H. Weishaupt, C. Behrends

Date Published: 16th Sep 2022

Publication Type: Journal

Mapping autophagosome contents identifies interleukin-7 receptor-alpha as a key cargo modulating CD4+ T cell proliferation.

Abstract (Expand)

CD4+ T cells are pivotal cells playing roles in the orchestration of humoral and cytotoxic immune responses. It is known that CD4+ T cell proliferation relies on autophagy, but identification of the … autophagosomal cargo involved is missing. Here we create a transgenic mouse model, to enable direct mapping of the proteinaceous content of autophagosomes in primary cells by LC3 proximity labelling. Interleukin-7 receptor-alpha, a cytokine receptor mostly found in naive and memory T cells, is reproducibly detected in autophagosomes of activated CD4+ T cells. Consistently, CD4+ T cells lacking autophagy show increased interleukin-7 receptor-alpha surface expression, while no defect in internalisation is observed. Mechanistically, excessive surface interleukin-7 receptor-alpha sequestrates the common gamma chain, impairing the interleukin-2 receptor assembly and downstream signalling crucial for T cell proliferation. This study shows that key autophagy substrates can be reliably identified in this mouse model and help mechanistically unravel autophagy's contribution to healthy physiology and disease.

Authors: D. Zhou, M. Borsa, D. J. Puleston, S. Zellner, J. Capera, S. Sanderson, M. Schifferer, S. S. Hester, X. Ge, R. Fischer, L. Jostins, C. Behrends, G. Alsaleh, A. K. Simon

Date Published: 2nd Sep 2022

Publication Type: Journal

Ubiquitin profiling of lysophagy identifies actin stabilizer CNN2 as a target of VCP/p97 and uncovers a link to HSPB1.

Abstract (Expand)

Lysosomal membrane permeabilization (LMP) is an underlying feature of diverse conditions including neurodegeneration. Cells respond by extensive ubiquitylation of membrane-associated proteins for … clearance of the organelle through lysophagy that is facilitated by the ubiquitin-directed AAA-ATPase VCP/p97. Here, we assessed the ubiquitylated proteome upon acute LMP and uncovered a large diversity of targets and lysophagy regulators. They include calponin-2 (CNN2) that, along with the Arp2/3 complex, translocates to damaged lysosomes and regulates actin filaments to drive phagophore formation. Importantly, CNN2 needs to be ubiquitylated during the process and removed by VCP/p97 for efficient lysophagy. Moreover, we identified the small heat shock protein HSPB1 that assists VCP/p97 in the extraction of CNN2 and show that other membrane regulators including SNAREs, PICALM, AGFG1, and ARL8B are ubiquitylated during lysophagy. Our data reveal a framework of how ubiquitylation and two effectors, VCP/p97 and HSPB1, cooperate to protect cells from the deleterious effects of LMP.

Authors: B. Kravic, T. Bionda, A. Siebert, P. Gahlot, S. Levantovsky, C. Behrends, H. Meyer

Date Published: 21st Jul 2022

Publication Type: Journal

Multi-omics profiling identifies a deregulated FUS-MAP1B axis in ALS/FTD-associated UBQLN2 mutants.

Abstract (Expand)

Ubiquilin-2 (UBQLN2) is a ubiquitin-binding protein that shuttles ubiquitinated proteins to proteasomal and autophagic degradation. UBQLN2 mutations are genetically linked to the neurodegenerative … disorders amyotrophic lateral sclerosis and frontotemporal dementia (ALS/FTD). However, it remains elusive how UBQLN2 mutations cause ALS/FTD. Here, we systematically examined proteomic and transcriptomic changes in patient-derived lymphoblasts and CRISPR/Cas9-engineered HeLa cells carrying ALS/FTD UBQLN2 mutations. This analysis revealed a strong up-regulation of the microtubule-associated protein 1B (MAP1B) which was also observed in UBQLN2 knockout cells and primary rodent neurons depleted of UBQLN2, suggesting that a UBQLN2 loss-of-function mechanism is responsible for the elevated MAP1B levels. Consistent with MAP1B's role in microtubule binding, we detected an increase in total and acetylated tubulin. Furthermore, we uncovered that UBQLN2 mutations result in decreased phosphorylation of MAP1B and of the ALS/FTD-linked fused in sarcoma (FUS) protein at S439 which is critical for regulating FUS-RNA binding and MAP1B protein abundance. Together, our findings point to a deregulated UBQLN2-FUS-MAP1B axis that may link protein homeostasis, RNA metabolism, and cytoskeleton dynamics, three molecular pathomechanisms of ALS/FTD.

Authors: L. Strohm, Z. Hu, Y. Suk, A. Ruhmkorf, E. Sternburg, V. Gattringer, H. Riemenschneider, R. Berutti, E. Graf, J. H. Weishaupt, M. S. Brill, A. B. Harbauer, D. Dormann, J. Dengjel, D. Edbauer, C. Behrends

Date Published: 1st Jul 2022

Publication Type: Journal

Signatures of glial activity can be detected in the CSF proteome.

Abstract (Expand)

Single-cell transcriptomics has revealed specific glial activation states associated with the pathogenesis of neurodegenerative diseases, such as Alzheimer's and Parkinson's disease. While these … findings may eventually lead to new therapeutic opportunities, little is known about how these glial responses are reflected by biomarker changes in bodily fluids. Such knowledge, however, appears crucial for patient stratification, as well as monitoring disease progression and treatment responses in clinical trials. Here, we took advantage of well-described mouse models of beta-amyloidosis and alpha-synucleinopathy to explore cerebrospinal fluid (CSF) proteome changes related to their respective proteopathic lesions. Nontargeted liquid chromatography-mass spectrometry revealed that the majority of proteins that undergo age-related changes in CSF of either mouse model were linked to microglia and astrocytes. Specifically, we identified a panel of more than 20 glial-derived proteins that were increased in CSF of aged beta-amyloid precursor protein- and alpha-synuclein-transgenic mice and largely overlap with previously described disease-associated glial genes identified by single-cell transcriptomics. Our results also show that enhanced shedding is responsible for the increase of several of the identified glial CSF proteins as exemplified for TREM2. Notably, the vast majority of these proteins can also be quantified in human CSF and reveal changes in Alzheimer's disease cohorts. The finding that cellular transcriptome changes translate into corresponding changes of CSF proteins is of clinical relevance, supporting efforts to identify fluid biomarkers that reflect the various functional states of glial responses in cerebral proteopathies, such as Alzheimer's and Parkinson's disease.

Authors: T. Eninger, S. A. Muller, M. Bacioglu, M. Schweighauser, M. Lambert, L. F. Maia, J. J. Neher, S. M. Hornfeck, U. Obermuller, G. Kleinberger, C. Haass, P. J. Kahle, M. Staufenbiel, L. Ping, D. M. Duong, A. I. Levey, N. T. Seyfried, S. F. Lichtenthaler, M. Jucker, S. A. Kaeser

Date Published: 14th Jun 2022

Publication Type: Journal

Proteomic profiling in cerebral amyloid angiopathy reveals an overlap with CADASIL highlighting accumulation of HTRA1 and its substrates.

Abstract (Expand)

Cerebral amyloid angiopathy (CAA) is an age-related condition and a major cause of intracerebral hemorrhage and cognitive decline that shows close links with Alzheimer's disease (AD). CAA is … characterized by the aggregation of amyloid-beta (Abeta) peptides and formation of Abeta deposits in the brain vasculature resulting in a disruption of the angioarchitecture. Capillaries are a critical site of Abeta pathology in CAA type 1 and become dysfunctional during disease progression. Here, applying an advanced protocol for the isolation of parenchymal microvessels from post-mortem brain tissue combined with liquid chromatography tandem mass spectrometry (LC-MS/MS), we determined the proteomes of CAA type 1 cases (n = 12) including a patient with hereditary cerebral hemorrhage with amyloidosis-Dutch type (HCHWA-D), and of AD cases without microvascular amyloid pathology (n = 13) in comparison to neurologically healthy controls (n = 12). ELISA measurements revealed microvascular Abeta(1-40) levels to be exclusively enriched in CAA samples (mean: > 3000-fold compared to controls). The proteomic profile of CAA type 1 was characterized by massive enrichment of multiple predominantly secreted proteins and showed significant overlap with the recently reported brain microvascular proteome of patients with cerebral autosomal-dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), a hereditary cerebral small vessel disease (SVD) characterized by the aggregation of the Notch3 extracellular domain. We found this overlap to be largely attributable to the accumulation of high-temperature requirement protein A1 (HTRA1), a serine protease with an established role in the brain vasculature, and several of its substrates. Notably, this signature was not present in AD cases. We further show that HTRA1 co-localizes with Abeta deposits in brain capillaries from CAA type 1 patients indicating a pathologic recruitment process. Together, these findings suggest a central role of HTRA1-dependent protein homeostasis in the CAA microvasculature and a molecular connection between multiple types of brain microvascular disease.

Authors: A. Zellner, S. A. Muller, B. Lindner, N. Beaufort, A. J. M. Rozemuller, T. Arzberger, N. C. Gassen, S. F. Lichtenthaler, B. Kuster, C. Haffner, M. Dichgans

Date Published: 24th Jan 2022

Publication Type: Journal

Proteomic and lipidomic profiling of demyelinating lesions identifies fatty acids as modulators in lesion recovery.

Abstract (Expand)

After demyelinating injury of the central nervous system, resolution of the mounting acute inflammation is crucial for the initiation of a regenerative response. Here, we aim to identify fatty acids … and lipid mediators that govern the balance of inflammatory reactions within demyelinating lesions. Using lipidomics, we identify bioactive lipids in the resolution phase of inflammation with markedly elevated levels of n-3 polyunsaturated fatty acids. Using fat-1 transgenic mice, which convert n-6 fatty acids to n-3 fatty acids, we find that reduction of the n-6/n-3 ratio decreases the phagocytic infiltrate. In addition, we observe accelerated decline of microglia/macrophages and enhanced generation of oligodendrocytes in aged mice when n-3 fatty acids are shuttled to the brain. Thus, n-3 fatty acids enhance lesion recovery and may, therefore, provide the basis for pro-regenerative medicines of demyelinating diseases in the central nervous system.

Authors: H. Penkert, A. Bertrand, V. Tiwari, S. Breimann, S. A. Muller, P. M. Jordan, M. J. Gerl, C. Klose, L. Cantuti-Castelvetri, M. Bosch-Queralt, I. Levental, S. F. Lichtenthaler, O. Werz, M. Simons

Date Published: 26th Oct 2021

Publication Type: Journal

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