Expression of motor proteins in transgenic HeLa cell lines
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE33271 | GEO | 2013/01/23
SECONDARY ACCESSION(S): PRJNA149323
REPOSITORIES: GEO
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Project description:We employed whole transcriptome microarray profiling to measure the expression of myosin and kinesin genes in multiple HeLa cell lines and confirm the expression of recombinantly-tagged BAC transgenes encoding motor genes. Normal and transfected HeLa cells grown to confluence were harvested for extraction of total RNA and microarray analysis
2013-01-23 | E-GEOD-33271 | biostudies-arrayexpress
Project description:Kinesins are a superfamily of molecular motors that undertake ATP-dependent microtubule-based movement or regulate microtubule dynamics. Plasmodium species, which cause malaria and kill hundreds of thousands annually, encode ~9 kinesins in their genomes. Of these, two – kinesin-8B and kinesin-8X - are canonically classified as kinesin-8s, which in other eukaryotes typically modulate microtubule dynamics. Unexpectedly, Plasmodium kinesin-8B is required for development of the flagellated male gamete in the mosquito host, and its absence completely blocks parasite transmission. To understand the molecular basis of kinesin-8B’s essential role, we characterised the in vitro properties of the kinesin-8B motor domains from P. berghei and P. falciparum. Both motors drive plus-end directed ATP-dependent microtubule gliding, but also catalyse ATP-dependent microtubule depolymerisation. We determined the microtubule-bound structures of these motors using cryo-electron microscopy. P. berghei and P. falciparum kinesin-8B exhibit a very similar mode of microtubule interaction, in which Plasmodium-distinct sequences at the microtubule-kinesin interface influence motor function. Intriguingly, however, P. berghei kinesin-8B exhibits a non-canonical structural response to ATP analogue binding such that neck linker docking is not induced. Nevertheless, the neck linker region is absolutely required for motility and depolymerisation activities of these motors. Taken together, these data suggest that the mechanochemistry of Plasmodium kinesin-8Bs has been functionally tuned to efficiently contribute to flagella formation.
2022-12-05 | PXD037474 | Pride
Project description:We report cell type specific Nova HITS-CLIP using BAC-transgenic lines expressing GFP-Nova under the motor neuron specific choline acetyltransferase (Chat) promoter. By comparing transcriptome wide Nova binding map in motor neurons and that in the whole spinal cord, we identified differential Nova binding sites in motor neurons, which correlate with motor neuron specific RNA processing.
2018-07-04 | GSE71294 | GEO
Project description: Not available
| S-ECPF-GEOD-33271 | biostudies-other
Project description:This is the 2 state model of Myosin V movement described in the article:
A simple kinetic model describes the processivity of myosin-v.
Kolomeisky AB , Fisher ME Biophys. J. 84(3):1642-50 (2003); PubmedID: 12609867
Abstract:
Myosin-V is a motor protein responsible for organelle and vesicle transport in cells. Recent single-molecule experiments have shown that it is an efficient processive motor that walks along actin filaments taking steps of mean size close to 36 nm. A theoretical study of myosin-V motility is presented following an approach used successfully to analyze the dynamics of conventional kinesin but also taking some account of step-size variations. Much of the present experimental data for myosin-V can be well described by a two-state chemical kinetic model with three load-dependent rates. In addition, the analysis predicts the variation of the mean velocity and of the randomness-a quantitative measure of the stochastic deviations from uniform, constant-speed motion-with ATP concentration under both resisting and assisting loads, and indicates a substep of size d(0) approximately 13-14 nm (from the ATP-binding state) that appears to accord with independent observations.
The model differs slightly from the published version. The ATP and ADP bound forms of myosin are called S0 and S1. The state transition and binding constants are called k_1, k_2, k_3 and k_4 instead of k0
0, u0
1, k'
0 and w0
1. Similarly the state loading factors are named th_1, th_2, th_3 and th_4 instead of θ+
0, θ+
1, θ-
0 and θ-
1. The species fwd_step1, fwd_step2, back_step1 and back_step2 count the number of state changes of each kind the myosine molecules have taken over time.
The model can be evaluated in a deterministic continuous or stochastic discreet fashion. The parameter V holds the (forward) speed at each time point, the V_avg the overall way divided by the simulation time and the amount of myosine molecules.
Originally created by libAntimony v1.4 (using libSBML 3.4.1)
This model originates from BioModels Database: A Database of Annotated Published Models (http://www.ebi.ac.uk/biomodels/). It is copyright (c) 2005-2011 The BioModels.net Team.
For more information see the terms of use.
To cite BioModels Database, please use: Li C, Donizelli M, Rodriguez N, Dharuri H, Endler L, Chelliah V, Li L, He E, Henry A, Stefan MI, Snoep JL, Hucka M, Le Novère N, Laibe C (2010) BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models. BMC Syst Biol., 4:92.
2006-09-29 | BIOMD0000000305 | BioModels
Project description:The accompanying dataset is the result of a systematic study to identify the RNA cargoes associated with the cytoskeletal motor proteins of Saccharomyces cerevisiae. We immunopurified, via the use of integrated, C-terminal GFP and 9Myc tags, the five actomyosin motors, Myo1, Myo2, Myo3, Myo4, and Myo5; the kinesin-like proteins Kar3, Kip1, Kip2, Kip3, and Smy1; and the dynein, Dyn1, from S. cerevisiae. Cells were either treated with formaldehyde or with the small molecule latrunculin B. We used formaldehyde crosslinking to stabilize associations between motors proteins and interacting RNAs before IP. Yeast cells growing exponentially in rich medium were treated with formaldehyde, lysed and sonicated, then incubated with magnetic beads coupled to monoclonal antibodies against either 9Myc or GFP to isolate the tagged motor proteins along with any associated RNAs. After IP, the formaldehyde crosslinks were reversed and the enriched RNAs and RNAs purified from the corresponding total lysate were amplified and labeled respectively with Cy5 and Cy3, then jointly hybridized to custom-made, S. cerevisiae oligonucleotide microarrays (Hogan et al., PLoS Biology, 2008). Alternatively, the addition of a low concentration of latrunculin B (2 ug/ml) to live cells to partially solubilize the actin cytoskeleton allowed for successful IP of the motor proteins and associated mRNAs without the need for a chemical crosslinker. In both the case of latrunculin B and formaldehyde treatment, we also performed IPs in which the untagged parent yeast strains were processed for IP and microarray analysis in a manner identical to that of the tagged strains (labeled as mock). transcription profiling
2012-01-27 | E-GEOD-35343 | biostudies-arrayexpress
Project description:Background Amyotrophic lateral sclerosis (ALS) is an age-dependent and incurable neurodegenerative disease. It is pathologically characterized by selective degeneration of motor neurons resulting in a catastrophic loss of motor function. Previous studies showed that excess copper (Cu) might have a role in ALS etiology. However, the underlying mechanism remains incompletely elucidated. Therefore, we aimed to investigate the neurotoxicity and underlying mechanism of Cu exposure in female ALS mice with SOD1G93A gene mutation. Methods After three months of treatment with low-dose Cu exposure (0.13 PPM copper chloride drinking water), the motor ability was assessed by various behavioral methods, including climbing-pole test, rotarod test, hanging endurance test, grip strength test and gait analysis. Muscle atrophy and fibrosis were evaluated by histopathology. The proliferation of astrocytes and microglia was assessed by immunohistochemical analysis. The number of motor neurons in the spinal cord was determined by Nissl staining and ChAT immunohistochemical staining. Western blotting and proteomics revealed the effect of Cu exposure on mitochondrial protein expression in mice. Results Behavioral tests showed that Cu exposure worsened motor function in SOD1G93A mice. Compared with SOD1G93A transgenic mice, there was no significant difference in the pole-climbing test after Cu exposure (P =0.6156). There were significant changes in rotarod test, hanging endurance test and grip strength test (P =0.0121; P = 0.0085; P =0.0105). At the same time, Cu exposure aggravated muscle fibrosis and the loss of spinal motor neurons (P <0.05) and was associated with neuroinflammation (P <0.05). The proteomic analysis further showed that differentially expressed proteins among wild-type mice (WT), ALS mice, and Cu-treated ALS mice were mainly involved in muscle contraction, ATP metabolic processes, immune responses, peroxisomes, mitochondrial electrons respiratory transport chain, GMP biological processes, and were co-related with disturbances in energy metabolism, which was reflected in mitochondrial dysfunction in ALS mice. In addition, Western Blot showed that Cu exposure decreased the expression of mitochondria-related proteins (P <0.05), which was consistent with the results of proteomic analysis. Conclusions In conclusion, our findings showed that Cu exposure aggravates motor dysfunction and pathological changes in ALS mice via mitochondrial dysfunction.
2023-01-29 | PXD039728 | iProX
Project description:TDP-43, a DNA/RNA binding protein involved in RNA transcription and splicing has been associated with the pathophysiology of neurodegenerative diseases, including ALS. However, the function of TDP-43 in motor neurons remains undefined. Here, we employ both gain- and loss-of-function approaches to determine roles of TDP-43 in motor neurons. Mice expressing human TDP-43 in neurons exhibited growth retardation and premature death that are characterized by abnormal intranuclear inclusions comprised of TDP-43 and Fused in Sarcoma (FUS), and massive accumulation of mitochondria in TDP-43-negative cytoplasmic inclusions in motor neurons, lack of mitochondria in motor axon terminals and immature neuromuscular junctions. Whereas elevated level of TDP-43 disrupts the normal nuclear distribution of Survival Motor Neuron (SMN)-associated Gemini of coiled bodies (GEMs) in motor neurons, its absence prevents the formation of GEMs in the nuclei of these cells. Moreover, transcriptome-wide deep sequencing analysis revealed that decrease in abundance of neurofilament transcripts contributed to the reduction of caliber of motor axons in TDP-43 mice. In concert, our findings indicate that TDP-43 participates in pathways critical for motor neuron physiology, including those that regulate the normal distributions of SMN-associated GEMs in the nucleus and mitochondria in the cytoplasm. Human TDP-43 coding region were inserted into pThy1.2 expression cassette and subsequently injected into C57BL/6;SJL hybrid mouse embryos to make human TDP-43 transgenic mice
2010-07-30 | E-GEOD-22351 | biostudies-arrayexpress
Project description:TDP-43, a DNA/RNA binding protein involved in RNA transcription and splicing has been associated with the pathophysiology of neurodegenerative diseases, including ALS. However, the function of TDP-43 in motor neurons remains undefined. Here, we employ both gain- and loss-of-function approaches to determine roles of TDP-43 in motor neurons. Mice expressing human TDP-43 in neurons exhibited growth retardation and premature death that are characterized by abnormal intranuclear inclusions comprised of TDP-43 and Fused in Sarcoma (FUS), and massive accumulation of mitochondria in TDP-43-negative cytoplasmic inclusions in motor neurons, lack of mitochondria in motor axon terminals and immature neuromuscular junctions. Whereas elevated level of TDP-43 disrupts the normal nuclear distribution of Survival Motor Neuron (SMN)-associated Gemini of coiled bodies (GEMs) in motor neurons, its absence prevents the formation of GEMs in the nuclei of these cells. Moreover, transcriptome-wide deep sequencing analysis revealed that decrease in abundance of neurofilament transcripts contributed to the reduction of caliber of motor axons in TDP-43 mice. In concert, our findings indicate that TDP-43 participates in pathways critical for motor neuron physiology, including those that regulate the normal distributions of SMN-associated GEMs in the nucleus and mitochondria in the cytoplasm.
2010-07-31 | GSE22351 | GEO
Project description:<p>This study contains all authorized whole genome sequence data of the HeLa cell line from datasets currently in dbGaP. These data have been approved for health, medical, and/or biomedical research purposes. Access to these data can be granted for one year. Accessible data will include the studies listed on this page and any additional authorized datasets that become available during this one-year period.</p> <p><a href="http://acd.od.nih.gov/hlgda.htm">The HeLa Genome Data Access Working Group of the Advisory Committee to the Director (ACD)</a> will review requests from the research community for access to these datasets and assess whether the requests align with the terms of use defined in the HeLa Genome Data Use Agreement. The Working Group's findings will be reported to the ACD, and the ACD will make recommendations to the NIH Director about whether a request should be approved or disapproved. The NIH Director will decide whether access to the data will be granted.</p>
| phs000640.v1.p1 | EGA