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The Journal of Cell Biology May 2024Axonal transport is essential for neuronal survival. This is driven by microtubule motors including dynein, which transports cargo from the axon tip back to the cell...
Axonal transport is essential for neuronal survival. This is driven by microtubule motors including dynein, which transports cargo from the axon tip back to the cell body. This function requires its cofactor dynactin and regulators LIS1 and NDEL1. Due to difficulties imaging dynein at a single-molecule level, it is unclear how this motor and its regulators coordinate transport along the length of the axon. Here, we use a neuron-inducible human stem cell line (NGN2-OPTi-OX) to endogenously tag dynein components and visualize them at a near-single molecule regime. In the retrograde direction, we find that dynein and dynactin can move the entire length of the axon (>500 µm). Furthermore, LIS1 and NDEL1 also undergo long-distance movement, despite being mainly implicated with the initiation of dynein transport. Intriguingly, in the anterograde direction, dynein/LIS1 moves faster than dynactin/NDEL1, consistent with transport on different cargos. Therefore, neurons ensure efficient transport by holding dynein/dynactin on cargos over long distances but keeping them separate until required.
Topics: Humans; Axonal Transport; Axons; Dynactin Complex; Dyneins; Neurons; Neural Stem Cells
PubMed: 38407313
DOI: 10.1083/jcb.202309084 -
Biology Open Jul 2023Within cilia, the dynein-2 complex needs to be transported as an anterograde cargo to achieve its role as a motor to drive retrograde trafficking of the intraflagellar...
Within cilia, the dynein-2 complex needs to be transported as an anterograde cargo to achieve its role as a motor to drive retrograde trafficking of the intraflagellar transport (IFT) machinery containing IFT-A and IFT-B complexes. We previously showed that interactions of WDR60 and the DYNC2H1-DYNC2LI1 dimer of dynein-2 with multiple IFT-B subunits, including IFT54, are required for the trafficking of dynein-2 as an IFT cargo. However, specific deletion of the IFT54-binding site from WDR60 demonstrated only a minor effect on dynein-2 trafficking and function. We here show that the C-terminal coiled-coil region of IFT54, which participates in its interaction with the DYNC2H1-DYNC2LI1 dimer of dynein-2 and with IFT20 of the IFT-B complex, is essential for IFT-B function, and suggest that the IFT54 middle linker region between the N-terminal WDR60-binding region and the C-terminal coiled-coil is required for ciliary retrograde trafficking, probably by mediating the effective binding of IFT-B to the dynein-2 complex, and thereby ensuring dynein-2 loading onto the anterograde IFT trains. The results presented here agree with the notion predicted from the previous structural models that the dynein-2 loading onto the anterograde IFT train relies on intricate, multivalent interactions between the dynein-2 and IFT-B complexes.
Topics: Dyneins; Cilia; Cytoskeleton; Biological Transport; Protein Binding
PubMed: 37309605
DOI: 10.1242/bio.059976 -
Neurobiology of Disease Jul 2023Cytoplasmic dynein is an important intracellular motor protein that plays an important role in neuronal growth, axonal polarity formation, dendritic differentiation, and...
Cytoplasmic dynein is an important intracellular motor protein that plays an important role in neuronal growth, axonal polarity formation, dendritic differentiation, and dendritic spine development among others. The intermediate chain of dynein, encoded by Dync1i1, plays a vital role in the dynein complex. Therefore, we assessed the behavioral and related neuronal activities in mice with dync1i1 gene knockout. Neuronal activities in primary somatosensory cortex were recorded by in vivo electrophysiology and manipulated by optogenetic and chemogenetics. Nociception of mechanical, thermal, and cold pain in Dync1i1 mice were impaired. The activities of parvalbumin (PV) interneurons and gamma oscillation in primary somatosensory were also impaired when exposed to mechanical nociceptive stimulation. This neuronal dysfunction was rescued by optogenetic activation of PV neurons in Dync1i1 mice, and mimicked by suppressing PV neurons using chemogenetics in WT mice. Impaired pain sensations in Dync1i1 mice were correlated with impaired gamma oscillations due to a loss of interneurons, especially the PV type. This genotype-driven approach revealed an association between impaired pain sensation and cytoplasmic dynein complex.
Topics: Mice; Animals; Parvalbumins; Somatosensory Cortex; Cytoplasmic Dyneins; Dyneins; Interneurons; Pain Threshold
PubMed: 37257662
DOI: 10.1016/j.nbd.2023.106170 -
The Journal of Biological Chemistry Sep 2023The Vps10p domain receptor SorCS2 is crucial for the development and function of the nervous system and essential for brain-derived neurotrophic factor (BDNF)-induced...
The Vps10p domain receptor SorCS2 is crucial for the development and function of the nervous system and essential for brain-derived neurotrophic factor (BDNF)-induced changes in neuronal morphology and plasticity. SorCS2 regulates the subcellular trafficking of the BDNF signaling receptor TrkB as well as selected neurotransmitter receptors in a manner that is dependent on the SorCS2 intracellular domain (ICD). However, the cellular machinery and adaptor protein (AP) interactions that regulate receptor trafficking via the SorCS2 ICD are unknown. We here identify four splice variants of human SorCS2 differing in the insertion of an acidic cluster motif and/or a serine residue within the ICD. We show that each variant undergoes posttranslational proteolytic processing into a one- or two-chain receptor, giving rise to eight protein isoforms, the expression of which differs between neuronal and nonneuronal tissues and is affected by cellular stressors. We found that the only variants without the serine were able to rescue BDNF-induced branching of SorCS2 knockout hippocampal neurons, while variants without the acidic cluster showed increased interactions with clathrin-associated APs AP-1, AP-2, and AP-3. Using yeast two-hybrid screens, we further discovered that all variants bound dynein light chain Tctex-type 3; however, only variants with an acidic cluster motif bound kinesin light chain 1. Accordingly, splice variants showed markedly different trafficking properties and localized to different subcellular compartments. Taken together, our findings demonstrate the existence of eight functional SorCS2 isoforms with differential capacity for interactions with cytosolic ligands dynein light chain Tctex-type 3 and kinesin light chain 1, which potentially allows cell-type specific SorCS2 trafficking and BDNF signaling.
Topics: Humans; Adaptor Proteins, Signal Transducing; Alternative Splicing; Brain-Derived Neurotrophic Factor; Dyneins; Kinesins; Protein Binding; Protein Isoforms; Receptor, trkB; Receptors, Cell Surface; Central Nervous System; Protein Processing, Post-Translational; Protein Transport
PubMed: 37507021
DOI: 10.1016/j.jbc.2023.105102 -
Nature Communications Jan 2024Lysosomes help maintain cellular proteostasis, and defects in lysosomal positioning and function can cause disease, including neurodegenerative disorders. The...
Lysosomes help maintain cellular proteostasis, and defects in lysosomal positioning and function can cause disease, including neurodegenerative disorders. The spatiotemporal distribution of lysosomes is regulated by small GTPases including Rabs, which are activated by guanine nucleotide exchange factors (GEFs). DENN domain proteins are the largest family of Rab GEFs. Using a cell-based assay, we screened DENND6A, a member of the DENN domain protein family against all known Rabs and identified it as a potential GEF for 20 Rabs, including Rab34. Here, we demonstrate that DENND6A activates Rab34, which recruits a RILP/dynein complex to lysosomes, promoting lysosome retrograde transport. Further, we identify DENND6A as an effector of Arl8b, a major regulatory GTPase on lysosomes. We demonstrate that Arl8b recruits DENND6A to peripheral lysosomes to activate Rab34 and initiate retrograde transport, regulating nutrient-dependent lysosomal juxtanuclear repositioning. Loss of DENND6A impairs autophagic flux. Our findings support a model whereby Arl8b/DENND6A/Rab34-dependent lysosomal retrograde trafficking controls autophagy.
Topics: Adaptor Proteins, Signal Transducing; Protein Binding; rab GTP-Binding Proteins; Lysosomes; Autophagy; Dyneins
PubMed: 38296963
DOI: 10.1038/s41467-024-44957-1 -
Pediatric Pulmonology Jul 2023Primary ciliary dyskinesia (PCD) is typically an autosomal recessive disease characterized by recurrent infections of the lower respiratory tract, frequent and severe...
BACKGROUND
Primary ciliary dyskinesia (PCD) is typically an autosomal recessive disease characterized by recurrent infections of the lower respiratory tract, frequent and severe otitis media, chronic rhinosinusitis, neonatal respiratory distress, and organ laterality defects. While severe lower respiratory tract infections and bronchiectasis are common in Inuit, PCD has not been recognized in this population.
METHODS
We report a case series of seven Inuit patients with PCD identified by genetic testing in three Canadian PCD centers.
RESULTS
Patients ranged from 4 to 59 years of age (at time of last evaluation) and originated in the Qikiqtaaluk region (Baffin Island, n = 5), Nunavut, or Nunavik (northern Quebec, n = 2), Canada. They had typical features of PCD, including neonatal respiratory distress (five patients), situs inversus totalis (four patients), bronchiectasis (four patients), chronic atelectasis (six patients), and chronic otitis media (six patients). Most had chronic rhinitis. Genetic evaluation demonstrated that all had homozygous pathogenic variants in DNAH11 at NM_001277115.1:c.4095+2C>A.
CONCLUSIONS
The discovery of this homozygous DNAH11 variant in widely disparate parts of the Nunangat (Inuit homelands) suggests this is a founder mutation that may be widespread in Inuit. Thus, PCD may be an important cause of chronic lung, sinus, and middle ear disease in this population. Inuit with chronic lung disease, including bronchiectasis or laterality defects, should undergo genetic testing for PCD. Consideration of including PCD genetic analysis in routine newborn screening should be considered in Inuit regions.
Topics: Humans; Alleles; Axonemal Dyneins; Canada; Cilia; Ciliary Motility Disorders; Inuit; Kartagener Syndrome; Otitis Media; Respiratory Distress Syndrome, Newborn; Child, Preschool; Child; Adolescent; Young Adult; Adult; Middle Aged
PubMed: 37088965
DOI: 10.1002/ppul.26414 -
Seizure Mar 2024The DYNC1H1 variants are associated with abnormal brain morphology and neuromuscular disorders that are accompanied by epilepsy. This study aimed to explore the...
OBJECTIVES
The DYNC1H1 variants are associated with abnormal brain morphology and neuromuscular disorders that are accompanied by epilepsy. This study aimed to explore the relationship between DYNC1H1 variants and epilepsy.
MATERIALS AND METHODS
Trios-based whole-exome sequencing was performed on patients with epilepsy. Previously reported epilepsy-related DYNC1H1 variants were systematically reviewed to analyse genotype-phenotype correlation.
RESULTS
The DYNC1H1 variants were identified in four unrelated cases of infant-onset epilepsy, including two de novo and two biallelic variants. Two patients harbouring de novo missense variants located in the stem and stalk domains presented with refractory epilepsies, whereas two patients harbouring biallelic variants located in the regions between functional domains had mild epilepsy with infrequent focal seizures and favourable outcomes. One patient presented with pachygyria and neurodevelopmental abnormalities, and the other three patients presented with normal development. These variants have no or low frequencies in the Genome Aggregation Database. All the missense variants were predicted to be damaging using silico tools. Previously reported epilepsy-related variants were monoallelic variants, mainly de novo missense variants, and all the patients presented with severe epileptic phenotypes or developmental delay and malformations of cortical development. Epilepsy-related variants were clustered in the dimerization and stalk domains, and generalized epilepsy-associated variants were distributed in the stem domain.
CONCLUSION
This study suggested that DYNC1H1 variants are potentially associated with infant-onset epilepsy without neurodevelopmental disorders, expanding the phenotypic spectrum of DYNC1H1. The genotype-phenotype correlation helps to understand the underlying mechanisms of phenotypic variation.
Topics: Infant; Humans; Mutation; Epilepsy; Neurodevelopmental Disorders; Epilepsy, Generalized; Mutation, Missense; Phenotype; Cytoplasmic Dyneins
PubMed: 37903666
DOI: 10.1016/j.seizure.2023.10.010 -
Frontiers in Immunology 2023Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths worldwide and has a poor prognosis. Thus, there is a need for an effective biomarker...
BACKGROUND
Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths worldwide and has a poor prognosis. Thus, there is a need for an effective biomarker to improve and predict the prognosis of HCC.
METHODS
RNA sequencing data, simple nucleotide variation data, and clinical data of HCC patients from The Cancer Genome Atlas (TCGA) to identify mutant genes, simple nucleotide variation data, and clinical data of HCC patients from the International Cancer Genome Consortium (ICGC) to validate the prognostic value of mutant genes were the data sources of the present study. To identify the overall survival (OS)-related mutant genes, a Kaplan-Meier (KM) analysis was conducted. We carried out univariate Cox and multivariate Cox regression analyses to identify the independent prognostic factors. We also conducted a correlation analysis of immune cells and mutant genes. To explore the molecular mechanisms of mutant genes, we conducted a gene set enrichment analysis (GSEA). A nomogram was constructed to help predict the prognosis of HCC. In addition, we explored the expression profile of mutant genes in HCC based on a TCGA dataset, an ICGC dataset, and our own HCC tissue samples.
RESULTS
We identified and validated a mutant gene, dynein axonemal heavy chain 5 (), which was negatively related to the OS of HCC patients. Univariate Cox and multivariate Cox regression analyses revealed that the mutant gene could act as an independent prognostic factor for HCC. Most pathways of the mutant gene were involved in cancer development and progression based on GSEA analysis. The mutant gene was negatively correlated with monocytes, naive CD4 T cells, activated dendritic cells, and activated mast cells. In addition, the mRNA and protein levels of had a significantly higher level of expression in the tissue samples of patients with HCC. A nomogram consisting of the pathological stage, , and tumor mutation burden (TMB) performed well.
CONCLUSION
The mutant gene had a significantly higher level of expression in the tissue samples of patients with HCC, could act as an independent prognostic factor for HCC, and is a potential new immunotherapy target for HCC.
Topics: Humans; Carcinoma, Hepatocellular; Prognosis; Liver Neoplasms; Nomograms; Nucleotides; Axonemal Dyneins
PubMed: 38022557
DOI: 10.3389/fimmu.2023.1236995 -
Advanced Science (Weinheim,... Nov 2023The principal cause of death in cancer patients is metastasis, which remains an unresolved problem. Conventionally, metastatic dissemination is linked to...
The principal cause of death in cancer patients is metastasis, which remains an unresolved problem. Conventionally, metastatic dissemination is linked to actomyosin-driven cell locomotion. However, the locomotion of cancer cells often does not strictly line up with the measured actomyosin forces. Here, a complementary mechanism of metastatic locomotion powered by dynein-generated forces is identified. These forces arise within a non-stretchable microtubule network and drive persistent contact guidance of migrating cancer cells along the biomimetic collagen fibers. It is also shown that the dynein-powered locomotion becomes indispensable during invasive 3D migration within a tissue-like luminal network formed by spatially confining granular hydrogel scaffolds (GHS) made up of microscale hydrogel particles (microgels). These results indicate that the complementary motricity mediated by dynein is always necessary and, in certain instances, sufficient for disseminating metastatic breast cancer cells. These findings advance the fundamental understanding of cell locomotion mechanisms and expand the spectrum of clinical targets against metastasis.
Topics: Humans; Female; Dyneins; Breast Neoplasms; Actomyosin; Cell Movement; Hydrogels
PubMed: 37726225
DOI: 10.1002/advs.202302229 -
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