Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Identification of CCNF/Cyclin F targets

ABSTRACT: The founding member of the F-box protein family, Cyclin F, serves as substrate adaptor for the Ubiquitin E3 ligase Skp1-Cul1-F-box (SCF)Cyclin F which is responsible for ubiquitination of proteins involved in cell cycle progression, DNA damage and mitotic fidelity. Missense mutations in CCNF encoding for Cyclin F are associated with amyotrophic lateral sclerosis (ALS). However, it remains elusive whether CCNF mutations affect the substrate adaptor function of Cyclin F and whether altered SCFCyclin F mediated ubiquitination contributes to ALS pathogenesis in CCNF mutation carrier. To start addressing these questions, we set out to identify new SCFCyclin F targets in neuronal and patient-derived cells. Mass spectrometry-based ubiquitinome profiling of CCNF 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 confirmed the Cyclin F dependent binding and ubiquitination of HSP90AB1 and unveiled a regulatory role of this ubiquitination in controlling the binding of a number of HSP90 clients and co-factors, thereby implying chaperone dysregulation and CCNF loss-of-function mechanism in ALS.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human) Mus Musculus (mouse)

TISSUE(S): Brain, Blood Cell, Epithelial Cell, Cell Culture, Lymph Node

DISEASE(S): Amyotrophic Lateral Sclerosis,Neuroblastoma

SUBMITTER: Christian Behrends

LAB HEAD: Christian Behrends

PROVIDER: PXD030729 | Pride | 2022-09-13

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	20191216_AS_128.raw	Raw
	20191216_AS_129.raw	Raw
	20191216_AS_130.raw	Raw
	20191216_AS_131.raw	Raw
	20191216_AS_132.raw	Raw

Items per page:

1 - 5 of 118

Similar Datasets

Mutations to the ALS/FTD gene CCNF reveals elevated Lys48-linked ubiquitination by the SCF(cyclin F) complex

Project description:Approximately 10% of Amyotrophic lateral sclerosis (ALS) cases have a positive family history (familial ALS) and appear clinically indistinguishable from sporadic cases. ALS and frontotemporal dementia (FTD) are fatal neurodegenerative disorders that have common molecular and pathogenic characteristics; however, their biological mechanisms remain poorly understood. We have previously identified CCNF missense mutations in cohorts of familial and sporadic ALS and FTD cases [ref]. CCNF encodes cyclin F, a component of an E3 ubiquitin-protein ligase (SCFCyclin F) complex that is responsible for ubiquitinating proteins for degradation by the Ubiquitin-Proteasome System (UPS). The SCFCyclin F complex is essential for maintaining cellular homeostasis in cells; but mutations appear to lead to aberrant motor neuron development and neuron degeneration. We revealed elevated Lys48-specific ubiquitination of proteins in neuronal cells expressing mutant CCNFS621G compared to the CCNFWT control, which is consistent with increased Lys48-specific E3 ligase activity of cyclin FS621G (>1.3-fold). Different subsets of immunoprecipitated Lys48-ubiquitinated proteins were identified between CCNFWT and CCNFS621G cells indicating that transfected cyclin F contribute to the protein ubiquitination profile. These findings provide mechanistic insights into the effects of CCNF gene mutations on the function of the SCFcyclin F complex on neuronal proteostasis.

2017-12-01 | PXD002011 | Pride

ALS/FTD-causing mutation in cyclin F causes the dysregulation of SFPQ

Project description:Recently, we identified missense mutations in CCNF that are causative of familial and sporadic amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). CCNF encodes for cyclin F, a substrate recognition component of an E3-ubiquitin ligase. Mutations in CCNF directly implicates disruption in the ubiquitin-proteasome system in the pathogenesis of ALS/FTD. In this study we used an unbiased proteomic screening workflow using the proximity-based ligation method, BioID, to identify putative interaction partners of cyclin F. In doing so, we found over 100 putative interaction partners of cyclin F. Notably, we demonstrated that cyclin F closely associates with a number of essential paraspeckle proteins, which are stress-responsive proteins that have recently been implicated in ALS pathogenesis. We further demonstrate that SCFcyclin F mediates the direct ubiquitylation and subsequent proteasomal degradation of RBM14, a core component of the paraspeckle complex. This degradation is defective when cyclin F carries an ALS/FTD-causing mutation, leading to RBM14 accumulation in primary neurons upon proteasome inhibition. Additionally, analysis of ALS patient post-mortem tissue revealed that RBM14 levels were significantly reduced in post-mortem ALS patient motor cortex and significantly reduced in the neurons of spinal cord tissue. Together, our data indicate that defects in RBM14 homeostasis, which can be due to defects in cyclin F-mediated degradation, may be a common factor underlying ALS/FTD disease pathogenesis.

2021-03-18 | PXD014163 | Pride

Expression of ALS-linked mutant CCNF in zebrafish leads to increased cell death in the spinal cord and an aberrant motor phenotype

Project description:Amyotrophic lateral sclerosis (ALS) is a rapidly progressive, fatal neurodegenerative disease characterised by the loss of upper and lower motor neurons. Approximately 10% of ALS cases have a known family history of the disease and mutations in multiple genes have been identified. ALS-linked mutations in CCNF were recently reported, however the pathogenic mechanisms associated with these mutations are yet to be established. To investigate possible mechanisms, an in vitro model of ALS was developed that expressed mutant CCNF in a neuronal cell line (Neuro-2a). Proteomic analysis of this in vitro model identified the disruption of several cellular pathways, including those associated with caspase-3 mediated cell death and axonal outgrowth. To establish whether these findings were replicated in vivo, a zebrafish model was developed. Transient overexpression of human mutant CCNF in zebrafish led to increased caspase-3 activity, increased cell death and a motor neuron axonopathy consisting of shortened primary motor axons and increased frequency of aberrant axonal branching. A significant correlation between the severity of this mutant CCNF-induced axonopathy and reduced motor function was also demonstrated in this model, with zebrafish expressing the mutant protein demonstrating an impaired motor response to a light stimulus. This is the first report of an ALS-linked CCNF mutation in vivo and indicates that zebrafish will be a useful tool to model the pathogenesis of CCNF-linked motor neuron degeneration.

2017-05-22 | PXD005131 | Pride

Phosphorylation of Cyclin F at Serine 621 regulates the Lys48-ubiquitylation E3 ligase activity of the SCF(Cyclin F) complex.

Project description:Amyotrophic Lateral Sclerosis (ALS) is a fatal neurodegenerative disorder that is characterized by progressive weakness, paralysis and cachexia often resulting in patient death within 3-5 years of diagnosis. Recently, we identified mutations to the CCNF gene, which encodes the Cyclin F protein, in cohorts of patients with familial and sporadic ALS (1). Cyclin F is a part of a Skp1-Cul-F-box (SCF) E3 ubiquitin-protein ligase complex and is responsible for ubiquitinating proteins for the degradation by the proteasome. In this study, we investigated the phosphorylation status of Cyclin F and the effect of the serine mutation at site 621 to glycine on E3 ligase Lys48-specific ubiquitylation activity. We identified seven phosphorylation sites on Cyclin F, of which five were unique including Ser621. These phosphorylation sites were mostly identified within the PEST sequence of Cyclin F at the C-terminus. Additionally, we determined that Casein Kinase II (CK2) was responsible for phosphorylating Ser621 and controls the E3 ligase activity of the SCF(Cyclin F) complex. Thus, the glycine mutation at this site on Cyclin F prevents phosphorylation by CK2 and confers elevated Lys48-ubiquitylation activity, a hallmark of ALS pathogenesis. These findings highlight the convergence of post-translational modifications (ubiquitylation and phosphorylation) to sustain cellular homeostasis, and aberrations such as a single missense mutation can affect downstream cellular processes and lead to aberrant motor neuron development, neuron degeneration and ultimately ALS.

2017-12-08 | PXD004531 | Pride

Promoters Active in Interphase are Bookmarked during Mitosis by Ubiquitination

Project description:We analyzed genome-wide modification of chromatin by ubiquitination in human cells and whether this mark changes through the cell cycle using modified ChIP-seq technique (ChAP-seq). Chromatin affinity purification was based on a standard ChIP method with modification of a two-step affinity purification.

2013-04-04 | E-GEOD-45771 | biostudies-arrayexpress

The IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation (ChIP-seq)

Project description:The IRE1a-XBP1 pathway, a conserved adaptive response to the unfolded protein response, is indispensable for development of the secretory cells. It maintains endoplasmic reticulum homeostasis by enhancing protein folding and the secretory capacity of the cells. Here, we used a modified ChIP-seq protocol (ChIPmentation) to investigate the genome-wide binding events of the transcription factor XBP1 in differentiated mouse Th2 cells.

2018-10-06 | E-MTAB-6327 | biostudies-arrayexpress

T helper cell type 2 ChIP-seq

Project description:CD4+ T cells were extracted from mouse and human. They were activated in vitro with CD3/28 and cultured with Il4. ChIPmentation was then performed

2017-12-06 | E-MTAB-6276 | biostudies-arrayexpress

Mouse ES cells expressing catalytically inactive Ring1B display impaired Ring1B and H3K27me3 deposition.

Project description:ChIP-seq for H3K27me3 and Ring1B was performed in WT mESCs and mESCs containing catalytically inactive Ring1B (I53A mutant). Cells expressing catalytically inactive Ring1B maintain the spatial distribution of Ring1B and H3K27me3 but at reduced levels. These findings support the notion that PRC2 recruitment is, in part, dependent on H2A ubiquitination (H2AK119ub). Two biological replicates were performed for Ring1B and H3K27me3 ChIPs in WT and Ring1B I53A/I53A mouse ESCs. Input chromatin was sequenced for each replicate as a control for ChIP enrichment.

2015-09-20 | E-GEOD-69955 | biostudies-arrayexpress

A simple and robust plate-based single-cell ATAC-seq on various tissues and cell lines

Project description:The assay for transposase-accessible chromatin using sequencing (ATAC-seq) is widely used to identify regulatory regions throughout the genome. However, only a few studies have been done at the single cell level (scATAC-seq) due to technical difficulties. Here we developed a simple and robust plate-based scATAC-seq method, combining upfront bulk tagmentation with single-nuclei sorting, to investigate open chromatin regions. We applied this method on mouse splenocytes and unbiasedly revealed key regulatory regions and transcription factors that define each cell (sub)type.

2018-10-10 | E-MTAB-6714 | biostudies-arrayexpress

Proteomics analysis zebrafish ALS model

Project description:Despite advances in studies of C9orf72-ALS, understanding the function of C9orf72 remains a key research element that is poorly explored. We generated a C9orf72-related ALS stable zebrafish line with a reduced expression of C9orf72.Using this stable transgenic zebrafish model, we analyzed the effects of reduced C9orf72 function on the zebrafish’s neuromuscular system.

2022-02-17 | PXD025850 | Pride

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data