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Life Science Alliance Jul 2024Rab6 is a key modulator of protein secretion. The dynein adapter Bicaudal D2 (BicD2) recruits the motors cytoplasmic dynein and kinesin-1 to Rab6-positive vesicles for...
Rab6 is a key modulator of protein secretion. The dynein adapter Bicaudal D2 (BicD2) recruits the motors cytoplasmic dynein and kinesin-1 to Rab6-positive vesicles for transport; however, it is unknown how BicD2 recognizes Rab6. Here, we establish a structural model for recognition of Rab6 by BicD2, using structure prediction and mutagenesis. The binding site of BicD2 spans two regions of Rab6 that undergo structural changes upon the transition from the GDP- to GTP-bound state, and several hydrophobic interface residues are rearranged, explaining the increased affinity of the active GTP-bound state. Mutations of Rab6 that abolish binding to BicD2 also result in reduced co-migration of Rab6/BicD2 in cells, validating our model. These mutations also severely diminished the motility of Rab6-positive vesicles in cells, highlighting the importance of the Rab6/BicD2 interaction for overall motility of the multi-motor complex that contains both kinesin-1 and dynein. Our results provide insights into trafficking of secretory and Golgi-derived vesicles and will help devise therapies for diseases caused by BicD2 mutations, which selectively affect the affinity to Rab6 and other cargoes.
Topics: rab GTP-Binding Proteins; Humans; Dyneins; Protein Binding; Binding Sites; Kinesins; Mutation; Microtubule-Associated Proteins; Protein Transport; Models, Molecular; Guanosine Triphosphate
PubMed: 38719748
DOI: 10.26508/lsa.202302430 -
The Journal of Biological Chemistry Apr 2024Organelles and vesicular cargoes are transported by teams of kinesin and dynein motors along microtubules. We isolated endocytic organelles from cells at different...
Organelles and vesicular cargoes are transported by teams of kinesin and dynein motors along microtubules. We isolated endocytic organelles from cells at different stages of maturation and reconstituted their motility along microtubules in vitro. We asked how the sets of motors transporting a cargo determine its motility and response to the microtubule-associated protein tau. Here, we find that phagosomes move in both directions along microtubules, but the directional bias changes during maturation. Early phagosomes exhibit retrograde-biased transport while late phagosomes are directionally unbiased. Correspondingly, early and late phagosomes are bound by different numbers and combinations of kinesins-1, -2, -3, and dynein. Tau stabilizes microtubules and directs transport within neurons. While single-molecule studies show that tau differentially regulates the motility of kinesins and dynein in vitro, less is known about its role in modulating the trafficking of endogenous cargoes transported by their native teams of motors. Previous studies showed that tau preferentially inhibits kinesin motors, which biases late phagosome transport towards the microtubule minus-end. Here, we show that tau strongly inhibits long-range, dynein-mediated motility of early phagosomes. Tau reduces forces generated by teams of dynein motors on early phagosomes and accelerates dynein unbinding under load. Thus, cargoes differentially respond to tau, where dynein complexes on early phagosomes are more sensitive to tau inhibition than those on late phagosomes. Mathematical modeling further explains how small changes in the number of kinesins and dynein on cargoes impact the net directionality but also that cargoes with different sets of motors respond differently to tau.
PubMed: 38677516
DOI: 10.1016/j.jbc.2024.107323 -
Nature Communications Apr 2024Intraflagellar transport (IFT) orchestrates entry of proteins into primary cilia. At the ciliary base, assembled IFT trains, driven by kinesin-2 motors, can transport...
Intraflagellar transport (IFT) orchestrates entry of proteins into primary cilia. At the ciliary base, assembled IFT trains, driven by kinesin-2 motors, can transport cargo proteins into the cilium, across the crowded transition zone. How trains assemble at the base and how proteins associate with them is far from understood. Here, we use single-molecule imaging in the cilia of C. elegans chemosensory neurons to directly visualize the entry of kinesin-2 motors, kinesin-II and OSM-3, as well as anterograde cargo proteins, IFT dynein and tubulin. Single-particle tracking shows that IFT components associate with trains sequentially, both in time and space. Super-resolution maps of IFT components in wild-type and mutant worms reveal ciliary ultrastructure and show that kinesin-II is essential for axonemal organization. Finally, imaging cilia lacking kinesin-II and/or transition zone function uncovers the interplay of kinesin-II and OSM-3 in driving efficient transport of IFT trains across the transition zone.
Topics: Caenorhabditis elegans; Animals; Cilia; Caenorhabditis elegans Proteins; Kinesins; Flagella; Tubulin; Axoneme; Dyneins; Biological Transport; Single Molecule Imaging; Protein Transport
PubMed: 38658528
DOI: 10.1038/s41467-024-47807-2 -
BMC Medical Genomics Apr 2024Liver cancer ranks sixth in incidence and third in mortality globally and hepatocellular carcinoma (HCC) accounts for 90% of it. Hypoxia, glycolysis, and lactate...
BACKGROUND
Liver cancer ranks sixth in incidence and third in mortality globally and hepatocellular carcinoma (HCC) accounts for 90% of it. Hypoxia, glycolysis, and lactate metabolism have been found to regulate the progression of HCC separately. However, there is a lack of studies linking the above three to predict the prognosis of HCC. The present study aimed to identify a hypoxia-glycolysis-lactate-related gene signature for assessing the prognosis of HCC.
METHODS
This study collected 510 hypoxia-glycolysis-lactate genes from Molecular Signatures Database (MSigDB) and then classified HCC patients from TCGA-LIHC by analyzing their hypoxia-glycolysis-lactate genes expression. Differentially expressed genes (DEGs) were screened out to construct a gene signature by LASSO-Cox analysis. Univariate and multivariate regression analyses were used to evaluate the independent prognostic value of the gene signature. Analyses of immune infiltration, somatic cell mutations, and correlation heatmap were conducted by "GSVA" R package. Single-cell analysis conducted by "SingleR", "celldex", "Seurat", and "CellCha" R packages revealed how signature genes participated in hypoxia/glycolysis/lactate metabolism and PPI network identified hub genes.
RESULTS
We classified HCC patients from TCGA-LIHC into two clusters and screened out DEGs. An 18-genes prognostic signature including CDCA8, CBX2, PDE6A, MED8, DYNC1LI1, PSMD1, EIF5B, GNL2, SEPHS1, CCNJL, SOCS2, LDHA, G6PD, YBX1, RTN3, ADAMTS5, CLEC3B, and UCK2 was built to stratify the risk of HCC. The risk score of the hypoxia-glycolysis-lactate gene signature was further identified as a valuable independent factor for estimating the prognosis of HCC. Then we found that the features of clinical characteristics, immune infiltration, somatic cell mutations, and correlation analysis differed between the high-risk and low-risk groups. Furthermore, single-cell analysis indicated that the signature genes could interact with the ligand-receptors of hepatocytes/fibroblasts/plasma cells to participate in hypoxia/glycolysis/lactate metabolism and PPI network identified potential hub genes in this process: CDCA8, LDHA, YBX1.
CONCLUSION
The hypoxia-glycolysis-lactate-related gene signature we built could provide prognostic value for HCC and suggest several hub genes for future HCC studies.
Topics: Humans; Lactic Acid; Carcinoma, Hepatocellular; Liver Neoplasms; Prognosis; Hypoxia; Eye Proteins; Cyclic Nucleotide Phosphodiesterases, Type 6; Cytoplasmic Dyneins
PubMed: 38627714
DOI: 10.1186/s12920-024-01867-x -
Experimental & Molecular Medicine Apr 2024Intracellular retrograde transport in eukaryotic cells relies exclusively on the molecular motor cytoplasmic dynein 1. Unlike its counterpart, kinesin, dynein has a... (Review)
Review
Intracellular retrograde transport in eukaryotic cells relies exclusively on the molecular motor cytoplasmic dynein 1. Unlike its counterpart, kinesin, dynein has a single isoform, which raises questions about its cargo specificity and regulatory mechanisms. The precision of dynein-mediated cargo transport is governed by a multitude of factors, including temperature, phosphorylation, the microtubule track, and interactions with a family of activating adaptor proteins. Activating adaptors are of particular importance because they not only activate the unidirectional motility of the motor but also connect a diverse array of cargoes with the dynein motor. Therefore, it is unsurprising that dysregulation of the dynein-activating adaptor transport machinery can lead to diseases such as spinal muscular atrophy, lower extremity, and dominant. Here, we discuss dynein motor motility within cells and in in vitro, and we present several methodologies employed to track the motion of the motor. We highlight several newly identified activating adaptors and their roles in regulating dynein. Finally, we explore the potential therapeutic applications of manipulating dynein transport to address diseases linked to dynein malfunction.
Topics: Humans; Cytoplasmic Dyneins; Animals; Biological Transport; Microtubules; Dyneins
PubMed: 38556551
DOI: 10.1038/s12276-024-01200-7 -
Science (New York, N.Y.) Mar 2024Cytoplasmic dynein is a microtubule motor vital for cellular organization and division. It functions as a ~4-megadalton complex containing its cofactor dynactin and a...
Cytoplasmic dynein is a microtubule motor vital for cellular organization and division. It functions as a ~4-megadalton complex containing its cofactor dynactin and a cargo-specific coiled-coil adaptor. However, how dynein and dynactin recognize diverse adaptors, how they interact with each other during complex formation, and the role of critical regulators such as lissencephaly-1 (LIS1) protein (LIS1) remain unclear. In this study, we determined the cryo-electron microscopy structure of dynein-dynactin on microtubules with LIS1 and the lysosomal adaptor JIP3. This structure reveals the molecular basis of interactions occurring during dynein activation. We show how JIP3 activates dynein despite its atypical architecture. Unexpectedly, LIS1 binds dynactin's p150 subunit, tethering it along the length of dynein. Our data suggest that LIS1 and p150 constrain dynein-dynactin to ensure efficient complex formation.
Topics: Cryoelectron Microscopy; Dynactin Complex; Dyneins; Microtubule-Associated Proteins; Microtubules; Protein Binding; Humans; HeLa Cells; 1-Alkyl-2-acetylglycerophosphocholine Esterase; Nerve Tissue Proteins; Adaptor Proteins, Signal Transducing; WD40 Repeats; Protein Interaction Mapping
PubMed: 38547289
DOI: 10.1126/science.adk8544 -
MicroPublication Biology 2024Ciliary-dynein preassembly in the cytoplasm is critical for the assembly and movement of motile cilia, organelles that function under viscous conditions. Defects in...
Ciliary-dynein preassembly in the cytoplasm is critical for the assembly and movement of motile cilia, organelles that function under viscous conditions. Defects in preassembly often lead to a reduction in specific types of ciliary dyneins. Here, we investigated how environmental viscosity affects the motility of preassembly-deficient cilia in the alga We found that, depending on the type of ciliary dynein deficiency, each mutant displays a characteristic phenotype in cell propulsion. Our results highlight not only the unique function(s) of each dynein species, but also the importance of functional coordination between dyneins for ciliary motility under viscous conditions.
PubMed: 38545438
DOI: 10.17912/micropub.biology.001149 -
Auris, Nasus, Larynx Jun 2024Primary ciliary dyskinesia (PCD) is a relatively rare genetic disorder that affects approximately 1 in 20,000 people. Approximately 50 genes are currently known to cause... (Review)
Review
OBJECTIVE
Primary ciliary dyskinesia (PCD) is a relatively rare genetic disorder that affects approximately 1 in 20,000 people. Approximately 50 genes are currently known to cause PCD. In light of differences in causative genes and the medical system in Japan compared with other countries, a practical guide was needed for the diagnosis and management of Japanese PCD patients.
METHODS
An ad hoc academic committee was organized under the Japanese Rhinologic Society to produce a practical guide, with participation by committee members from several academic societies in Japan. The practical guide including diagnostic criteria for PCD was approved by the Japanese Rhinologic Society, Japanese Society of Otolaryngology-Head and Neck Surgery, Japanese Respiratory Society, and Japanese Society of Pediatric Pulmonology.
RESULTS
The diagnostic criteria for PCD consist of six clinical features, six laboratory findings, differential diagnosis, and genetic testing. The diagnosis of PCD is categorized as definite, probable, or possible PCD based on a combination of the four items above. Diagnosis of definite PCD requires exclusion of cystic fibrosis and primary immunodeficiency, at least one of the six clinical features, and a positive result for at least one of the following: (1) Class 1 defect on electron microscopy of cilia, (2) pathogenic or likely pathogenic variants in a PCD-related gene, or (3) impairment of ciliary motility that can be repaired by correcting the causative gene variants in iPS cells established from the patient's peripheral blood cells.
CONCLUSION
This practical guide provides clinicians with useful information for the diagnosis and management of PCD in Japan.
Topics: Humans; Kartagener Syndrome; Genetic Testing; Diagnosis, Differential; Cilia; Japan; Axonemal Dyneins; Proteins
PubMed: 38537559
DOI: 10.1016/j.anl.2024.02.001 -
Journal of Cell Science Apr 2024Primary cilia are essential eukaryotic organelles required for signalling and secretion. Dynein-2 is a microtubule-motor protein complex and is required for ciliogenesis...
Primary cilia are essential eukaryotic organelles required for signalling and secretion. Dynein-2 is a microtubule-motor protein complex and is required for ciliogenesis via its role in facilitating retrograde intraflagellar transport (IFT) from the cilia tip to the cell body. Dynein-2 must be assembled and loaded onto IFT trains for entry into cilia for this process to occur, but how dynein-2 is assembled and how it is recycled back into a cilium remain poorly understood. Here, we identify centrosomal protein of 170 kDa (CEP170) as a dynein-2-interacting protein in mammalian cells. We show that loss of CEP170 perturbs intraflagellar transport and hedgehog signalling, and alters the stability of dynein-2 holoenzyme complex. Together, our data indicate a role for CEP170 in supporting cilia function and dynein-2 assembly.
Topics: Cilia; Humans; Microtubule-Associated Proteins; Animals; Dyneins; Hedgehog Proteins; Signal Transduction; Mice; Flagella
PubMed: 38533689
DOI: 10.1242/jcs.261816 -
Trends in Parasitology May 2024Microtubules (MTs) play a vital role as key components of the eukaryotic cytoskeleton. The phylum Apicomplexa comprises eukaryotic unicellular parasitic organisms... (Review)
Review
Microtubules (MTs) play a vital role as key components of the eukaryotic cytoskeleton. The phylum Apicomplexa comprises eukaryotic unicellular parasitic organisms defined by the presence of an apical complex which consists of specialized secretory organelles and tubulin-based cytoskeletal elements. One apicomplexan parasite, Toxoplasma gondii, is an omnipresent opportunistic pathogen with significant medical and veterinary implications. To ensure successful infection and widespread dissemination, T. gondii heavily relies on the tubulin structures present in the apical complex. Recent advances in high-resolution imaging, coupled with reverse genetics, have offered deeper insights into the composition, functionality, and dynamics of these tubulin-based structures. The apicomplexan tubulins differ from those of their mammalian hosts, endowing them with unique attributes and susceptibility to specific classes of inhibitory compounds.
Topics: Toxoplasma; Tubulin; Cytoskeleton; Animals; Microtubules; Humans; Protozoan Proteins
PubMed: 38531711
DOI: 10.1016/j.pt.2024.02.010