Syntaxin of plants31 (SYP31) and SYP32 is essential for Golgi morphology maintenance and pollen development

Abstract Pollen development is a key process for the sexual reproduction of angiosperms. The Golgi plays a critical role in pollen development via the synthesis and transport of cell wall materials. However, little is known about the molecular mechanisms underlying the maintenance of Golgi integrity in plants. In Arabidopsis thaliana, syntaxin of plants (SYP) 3 family proteins SYP31 and SYP32 are the only two Golgi-localized Qa-soluble N-ethylmaleimide sensitive factor attachment protein receptors (SNAREs) with unknown endogenous functions. Here, we demonstrate the roles of SYP31 and SYP32 in modulating Golgi morphology and pollen development. Two independent lines of syp31/+ syp32/+ double mutants were male gametophytic lethal; the zero transmission rate of syp31 syp32 mutations was restored to largely normal levels by pSYP32:SYP32 but not pSYP32:SYP31 transgenes, indicating their functional differences in pollen development. The initial arrest of syp31 syp32 pollen occurred during the transition from the microspore to the bicellular stage, where cell plate formation in pollen mitosis I (PMI) and deposition of intine were abnormal. In syp31 syp32 pollen, the number and length of Golgi cisterna were significantly reduced, accompanied by many surrounding vesicles, which could be largely attributed to defects in anterograde and retrograde trafficking routes. SYP31 and SYP32 directly interacted with COG3, a subunit of the conserved oligomeric Golgi (COG) complex and were responsible for its Golgi localization, providing an underlying mechanism for SYP31/32 function in intra-Golgi trafficking. We propose that SYP31 and SYP32 play partially redundant roles in pollen development by modulating protein trafficking and Golgi structure.

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4-phenylbutyrate restored GABA uptake and reduced seizures in SLC6A1 variants-mediated disorders

10.1101/2021.12.08.471799 ◽

2021 ◽

Author(s):

Gerald I Nwosu ◽

Felicia Mermer ◽

Carson Flamm ◽

Sarah Poliquin ◽

Wangzhen Shen ◽

...

Keyword(s):

Cystic Fibrosis ◽

Membrane Trafficking ◽

Protein Trafficking ◽

Molecular Mechanisms ◽

Critical Role ◽

Gaba Uptake ◽

Functional Protein ◽

Great Opportunity ◽

Treatment Development ◽

The Impact

We have previously studied the molecular mechanisms of solute carrier family 6 member 1 (SLC6A1) associated with a continuum of neurodevelopmental disorders, including various epilepsy syndromes, autism, and intellectual disability. Based on functional assays of variants in a large cohort with heterogenous clinical phenotypes, we conclude that partial or complete loss of GABA uptake function in the mutant GAT-1 is the primary etiology as identified in GABAA receptor mutation-mediated epilepsy and in cystic fibrosis. Importantly, we identified that there are common patterns of the mutant protein trafficking from biogenesis, oligomerization, glycosylation, and translocation to the cell membrane across variants with the conservation of this process across cell types. Conversely any approach to facilitate membrane trafficking would increase presence of the functional protein in the targeted destination in all involved cells. PBA is an FDA-approved drug for pediatric use and is orally bioavailable so it can be quickly translated to patient use. It has been demonstrated that PBA can correct protein misfolding, reduce ER stress, and attenuate unfolded protein response in neurodegenerative diseases, it has also showed promise in treatment of cystic fibrosis. The common cellular mechanisms shared by the mutant GAT-1 and the mutant cystic fibrosis transmembrane conductance regulator led us to test if PBA and other pharmaco-chaperones could be a potential treatment option for SLC6A1 mutations. We examined the impact of PBA and other small molecules in a library of variants and in cell and knockin mouse models. Because of the critical role of astrocytic GAT-1 deficit in seizures, we focused on astrocytes, and demonstrated that the existence of the mutant GAT-1 retained the wildtype GAT-1, suggesting aberrant protein oligomerization and trafficking caused by the mutant GAT-1. PBA increased GABA uptake in both mouse and human astrocytes bearing the mutations. Importantly, PBA increased GAT-1 expression and suppressed spike wave discharges (SWDS) in the heterozygous knockin mice. Although the detailed mechanisms of action for PBA are ambiguous, it is likely that PBA can facilitate the forward trafficking of the wildtype GAT-1 favoring over the mutant GAT-1, thus increasing GABA uptake. Since all patients with SLC6A1 mutations are heterozygous and carry one wildtype functional allele, this suggests a great opportunity for treatment development by leveraging the endogenous protein trafficking pathway to promote forward trafficking of the wildtype in combination with enhancing the disposal of the mutant allele as treatment mode. The study opens a novel avenue of treatment development for genetic epilepsy via drug repurposing.

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VPS35 Protects Against TMEM230-Mutation-Induced Progressive Locomotor Deficits in Drosophila

10.21203/rs.3.rs-952504/v1 ◽

2021 ◽

Author(s):

Chao Ma ◽

Xiaobo Wang ◽

Wanli W Smith ◽

Zhaohui Liu

Keyword(s):

Protein Trafficking ◽

Molecular Mechanisms ◽

Caspase 3 ◽

Transmembrane Protein ◽

Critical Role ◽

In Vivo Studies ◽

Wild Type ◽

Dopaminergic Neurodegeneration ◽

Locomotor Deficits

Abstract BackgroundRecently, four Parkinson’s disease (PD)-linked mutations (Y92C, R141L, 184PGext*5 and 184Wext*5) in transmembrane protein 230 (TMEM230) were identified in PD patients, and these mutations have implications in protein trafficking and neurodegeneration. However, there is a lack of in vivo studies on the roles of PD-related variants of TMEM230 in PD pathogenesis.MethodsIn this study, we generated human wild-type (WT) and mutant TMEM230 (Y92C, R141L, 184PGext*5 and 184Wext*5) transgenic Drosophila using isoform Ⅱ cDNA. ResultsWe found that the expression of TMEM230 184PGext*5 in pan-neurons or dopaminergic neurons in Drosophila induced PD-like phenotypes, which included impaired locomotor ability, a shortened lifespan, reduced TH levels, and increased phosphorylated JNK and cleaved caspase-3 levels. Moreover, rotenone, a common pesticide, enhanced TMEM230-184PGext*5-induced PD-like phenotypes. In contrast, the overexpression of wild-type (WT) VPS35 rescued TMEM230-184PGext*5-induced PD-like phenotypes, while the knockdown of VPS35 by RNA interference (RNAi) or the expression of mutant VPS35 D620N worsened PD-like phenotypes. ConclusionThese results indicate that VPS35, as a downstream effector of TMEM230, plays a critical role in TMEM230-linked JNK/caspase-3 signalling pathways and that mutations in TMEM230 and VPS35 disrupt these pathways, resulting in dopaminergic neurodegeneration and PD-like phenotypes. These findings provide novel insight into the molecular mechanisms of mutant TME230- and VPS35-induced abnormalities underlying the pathogenesis of PD.

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Differential Roles of Syntaxin 7 and Syntaxin 8 in Endosomal Trafficking

Molecular Biology of the Cell ◽

10.1091/mbc.10.11.3891 ◽

1999 ◽

Vol 10 (11) ◽

pp. 3891-3908 ◽

Cited By ~ 91

Author(s):

Rytis Prekeris ◽

Bin Yang ◽

Viola Oorschot ◽

Judith Klumperman ◽

Richard H. Scheller

Keyword(s):

Membrane Fusion ◽

Protein Trafficking ◽

Molecular Mechanisms ◽

Protein Transport ◽

Growth Factor Receptor ◽

Endosomal Trafficking ◽

Attachment Protein ◽

Early Endosomes ◽

Protein Receptors ◽

And Function

To understand molecular mechanisms that regulate the intricate and dynamic organization of the endosomal compartment, it is important to establish the morphology, molecular composition, and functions of the different organelles involved in endosomal trafficking. Syntaxins and vesicle-associated membrane protein (VAMP) families, also known as soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein receptors (SNAREs), have been implicated in mediating membrane fusion and may play a role in determining the specificity of vesicular trafficking. Although several SNAREs, including VAMP3/cellubrevin, VAMP8/endobrevin, syntaxin 13, and syntaxin 7, have been localized to the endosomal membranes, their precise localization, biochemical interactions, and function remain unclear. Furthermore, little is known about SNAREs involved in lysosomal trafficking. So far, only one SNARE, VAMP7, has been localized to late endosomes (LEs), where it is proposed to mediate trafficking of epidermal growth factor receptor to LEs and lysosomes. Here we characterize the localization and function of two additional endosomal syntaxins, syntaxins 7 and 8, and propose that they mediate distinct steps of endosomal protein trafficking. Both syntaxins are found in SNARE complexes that are dissociated by α-soluble NSF attachment protein and NSF. Syntaxin 7 is mainly localized to vacuolar early endosomes (EEs) and may be involved in protein trafficking from the plasma membrane to the EE as well as in homotypic fusion of endocytic organelles. In contrast, syntaxin 8 is likely to function in clathrin-independent vesicular transport and membrane fusion events necessary for protein transport from EEs to LEs.

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Conserved Molecular Mechanisms Underlying Homeostasis of the Golgi Complex

International Journal of Cell Biology ◽

10.1155/2010/758230 ◽

2010 ◽

Vol 2010 ◽

pp. 1-9 ◽

Cited By ~ 8

Author(s):

Cathal Wilson ◽

Antonella Ragnini-Wilson

Keyword(s):

Golgi Complex ◽

Protein Trafficking ◽

Secretory Pathway ◽

Molecular Mechanisms ◽

Underlying Mechanism ◽

Central Function ◽

Sequential Processing ◽

The Face ◽

Regulate Protein ◽

Dynamic Structures

The Golgi complex performs a central function in the secretory pathway in the sorting and sequential processing of a large number of proteins destined for other endomembrane organelles, the plasma membrane, or secretion from the cell, in addition to lipid metabolism and signaling. The Golgi apparatus can be regarded as a self-organizing system that maintains a relatively stable morphofunctional organization in the face of an enormous flux of lipids and proteins. A large number of the molecular players that operate in these processes have been identified, their functions and interactions defined, but there is still debate about many aspects that regulate protein trafficking and, in particular, the maintenance of these highly dynamic structures and processes. Here, we consider how an evolutionarily conserved underlying mechanism based on retrograde trafficking that uses lipids, COPI, SNAREs, and tethers could maintain such a homeodynamic system.

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Regulatory Roles of Osteopontin in Production of Monocyte-Origin MCP-1

Cell Transplantation ◽

10.1177/0963689718756070 ◽

2018 ◽

Vol 27 (8) ◽

pp. 1185-1194

Author(s):

Liying Shi ◽

Lin Shi ◽

Xiangdong Wang ◽

Jiantai He

Keyword(s):

Molecular Mechanisms ◽

Kinase Inhibitors ◽

Critical Role ◽

Inflammatory Cells ◽

Underlying Mechanism ◽

Leukocyte Migration ◽

Enzyme Linked Immunosorbent Assay ◽

Monocyte Chemoattractant Protein 1 ◽

Monocyte Migration ◽

Molecular Events

Osteopontin (OPN), expressed by various immune cells, plays a critical role in leukocyte migration. Although OPN was found to selectively induce the expression of proinflammatory chemokines, the molecular mechanisms that control OPN gene expression and its underlying mechanism for migration and recruitment of inflammatory cells remain largely unknown. In this study, real-time polymerase chain reaction and enzyme-linked immunosorbent assay were used to determine OPN and monocyte chemoattractant protein 1 (MCP-1) expression. Signaling and molecular events between OPN and MCP-1 were analyzed by Western blot. Leukocyte migration in the presence of OPN was measured by chemotaxis assay. Our data indicated that phosphoinositide 3-kinase (PI3K), c-Jun NH2-terminal kinase (JNK), and extracellular signal-regulated kinase (ERK) that are activated upon stimulation with lipopolysaccharide were shown to upregulate OPN expression. Endogenous production of OPN was attributable to increased production of MCP-1, and this effect could be blocked by an anti-β1 integrin antibody and JNK and p38 kinase inhibitors. Furthermore, we found that the effect of OPN on inflammatory cell migration was mediated through inducing the expression of MCP-1 in monocytes. These results support a role of OPN in monocyte migration via MCP-1, which may represent an additional mechanism for innate and adaptive immune responses.

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The Role of the Endothelium in Premature Atherosclerosis: Molecular Mechanisms

Current Medicinal Chemistry ◽

10.2174/0929867326666190911141951 ◽

2020 ◽

Vol 27 (7) ◽

pp. 1041-1051 ◽

Cited By ~ 1

Author(s):

Michael Spartalis ◽

Eleftherios Spartalis ◽

Antonios Athanasiou ◽

Stavroula A. Paschou ◽

Christos Kontogiannis ◽

...

Keyword(s):

Molecular Mechanisms ◽

Low Density Lipoprotein ◽

Critical Role ◽

Density Lipoprotein ◽

Number Of Genes ◽

Causes Of Mortality ◽

Artery Disease ◽

Genetic And Environmental Factors ◽

Made In

Atherosclerotic disease is still one of the leading causes of mortality. Atherosclerosis is a complex progressive and systematic artery disease that involves the intima of the large and middle artery vessels. The inflammation has a key role in the pathophysiological process of the disease and the infiltration of the intima from monocytes, macrophages and T-lymphocytes combined with endothelial dysfunction and accumulated oxidized low-density lipoprotein (LDL) are the main findings of atherogenesis. The development of atherosclerosis involves multiple genetic and environmental factors. Although a large number of genes, genetic polymorphisms, and susceptible loci have been identified in chromosomal regions associated with atherosclerosis, it is the epigenetic process that regulates the chromosomal organization and genetic expression that plays a critical role in the pathogenesis of atherosclerosis. Despite the positive progress made in understanding the pathogenesis of atherosclerosis, the knowledge about the disease remains scarce.

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Molecular Mechanisms of Complement System Proteins and Matrix Metalloproteinases in the Pathogenesis of Age-Related Macular Degeneration

Current Molecular Medicine ◽

10.2174/1566524019666190828150625 ◽

2019 ◽

Vol 19 (10) ◽

pp. 705-718 ◽

Cited By ~ 4

Author(s):

Naima Mansoor ◽

Fazli Wahid ◽

Maleeha Azam ◽

Khadim Shah ◽

Anneke I. den Hollander ◽

...

Keyword(s):

Matrix Metalloproteinases ◽

Macular Degeneration ◽

Complement System ◽

Molecular Mechanisms ◽

Critical Role ◽

Age Related Macular Degeneration ◽

Genetic Changes ◽

Complement System Proteins ◽

Age Related ◽

Common Genetic Variants

: Age-related macular degeneration (AMD) is an eye disorder affecting predominantly the older people above the age of 50 years in which the macular region of the retina deteriorates, resulting in the loss of central vision. The key factors associated with the pathogenesis of AMD are age, smoking, dietary, and genetic risk factors. There are few associated and plausible genes involved in AMD pathogenesis. Common genetic variants (with a minor allele frequency of >5% in the population) near the complement genes explain 40–60% of the heritability of AMD. The complement system is a group of proteins that work together to destroy foreign invaders, trigger inflammation, and remove debris from cells and tissues. Genetic changes in and around several complement system genes, including the CFH, contribute to the formation of drusen and progression of AMD. Similarly, Matrix metalloproteinases (MMPs) that are normally involved in tissue remodeling also play a critical role in the pathogenesis of AMD. MMPs are involved in the degradation of cell debris and lipid deposits beneath retina but with age their functions get affected and result in the drusen formation, succeeding to macular degeneration. In this review, AMD pathology, existing knowledge about the normal and pathological role of complement system proteins and MMPs in the eye is reviewed. The scattered data of complement system proteins, MMPs, drusenogenesis, and lipofusogenesis have been gathered and discussed in detail. This might add new dimensions to the understanding of molecular mechanisms of AMD pathophysiology and might help in finding new therapeutic options for AMD.

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LncRNA SNHG12 Improves Cerebral Ischemic-reperfusion Injury by Activating SIRT1/FOXO3a Pathway through I nhibition of Autophagy and Oxidative Stress

Current Neurovascular Research ◽

10.2174/1567202617666200727142019 ◽

2020 ◽

Vol 17 (4) ◽

pp. 394-401

Author(s):

Yuanhua Wu ◽

Yuan Huang ◽

Jing Cai ◽

Donglan Zhang ◽

Shixi Liu ◽

...

Keyword(s):

Oxidative Stress ◽

Molecular Mechanisms ◽

Ischemia Reperfusion ◽

Critical Role ◽

Cell Activity ◽

Ht22 Cells ◽

Cerebral Ischemic ◽

Cerebral Ischemic Reperfusion ◽

And Oxidative Stress

Background: Ischemia/reperfusion (I/R) injury involves complex biological processes and molecular mechanisms such as autophagy. Oxidative stress plays a critical role in the pathogenesis of I/R injury. LncRNAs are the regulatory factor of cerebral I/R injury. Methods: This study constructs cerebral I/R model to investigate role of autophagy and oxidative stress in cerebral I/R injury and the underline regulatory mechanism of SIRT1/ FOXO3a pathway. In this study, lncRNA SNHG12 and FOXO3a expression was up-regulated and SIRT1 expression was down-regulated in HT22 cells of I/R model. Results: Overexpression of lncRNA SNHG12 significantly increased the cell viability and inhibited cerebral ischemicreperfusion injury induced by I/Rthrough inhibition of autophagy. In addition, the transfected p-SIRT1 significantly suppressed the release of LDH and SOD compared with cells co-transfected with SIRT1 and FOXO3a group and cells induced by I/R and transfected with p-SNHG12 group and overexpression of cells co-transfected with SIRT1 and FOXO3 further decreased the I/R induced release of ROS and MDA. Conclusion: In conclusion, lncRNA SNHG12 increased cell activity and inhibited oxidative stress through inhibition of SIRT1/FOXO3a signaling-mediated autophagy in HT22 cells of I/R model. This study might provide new potential therapeutic targets for further investigating the mechanisms in cerebral I/R injury and provide.

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Nano-Curcumin Regulates p53 Phosphorylation and PAI-1 Expression during Bleomycin Induced Injury in Alveolar Basal Epithelial Cells

Current Bioactive Compounds ◽

10.2174/1573407214666180727093612 ◽

2020 ◽

Vol 16 (1) ◽

pp. 85-89

Author(s):

Mahesh M. Gouda ◽

Ashwini Prabhu ◽

Varsha Reddy S.V. ◽

Rafa Jahan ◽

Yashodhar P. Bhandary

Keyword(s):

Epithelial Cells ◽

Lung Injury ◽

Molecular Mechanisms ◽

A549 Cells ◽

Plasminogen Activator Inhibitor ◽

Underlying Mechanism ◽

Protective Role ◽

Alveolar Epithelial Cells ◽

Cellular Damage

Background: Bleomycin (BLM) is known to cause DNA damage in the Alveolar Epithelial Cells (AECs). It is reported that BLM is involved in the up-regulation of inflammatory molecules such as neutrophils, macrophages, chemokines and cytokines. The complex underlying mechanism for inflammation mediated progression of lung injury is still unclear. This investigation was designed to understand the molecular mechanisms associated with p53 mediated modulation of Plasminogen Activator Inhibitor-I (PAI-I) expression and its regulation by nano-curcumin formulation. Methods: A549 cells were treated with BLM to cause the cellular damage in vitro and commercially available nano-curcumin formulation was used as an intervention. Cytotoxic effect of nano-curcumin was analyzed using Methyl Thiazolyl Tetrazolium (MTT) assay. Protein expressions were analyzed using western blot to evaluate the p53 mediated changes in PAI-I expression. Results: Nano-curcumin showed cytotoxicity up to 88.5 % at a concentration of 20 μg/ml after 48 h of treatment. BLM exposure to the cells activated the phosphorylation of p53, which in turn increased PAII expression. Nano-curcumin treatment showed a protective role against phosphorylation of p53 and PAI-I expression, which in turn regulated the fibro-proliferative phase of injury induced by bleomycin. Conclusion: Nano-curcumin could be used as an effective intervention to regulate the severity of lung injury, apoptosis of AECs and fibro-proliferation during pulmonary injury.

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Mechanosensation and Mechanotransduction by Lymphatic Endothelial Cells Act as Important Regulators of Lymphatic Development and Function

International Journal of Molecular Sciences ◽

10.3390/ijms22083955 ◽

2021 ◽

Vol 22 (8) ◽

pp. 3955

Author(s):

László Bálint ◽

Zoltán Jakus

Keyword(s):

Endothelial Cells ◽

Endothelial Cell ◽

Cell Fate ◽

Molecular Mechanisms ◽

Critical Role ◽

Mechanical Forces ◽

Lymphatic Endothelial Cells ◽

Lymphatic Development ◽

And Function ◽

The Impact

Our understanding of the function and development of the lymphatic system is expanding rapidly due to the identification of specific molecular markers and the availability of novel genetic approaches. In connection, it has been demonstrated that mechanical forces contribute to the endothelial cell fate commitment and play a critical role in influencing lymphatic endothelial cell shape and alignment by promoting sprouting, development, maturation of the lymphatic network, and coordinating lymphatic valve morphogenesis and the stabilization of lymphatic valves. However, the mechanosignaling and mechanotransduction pathways involved in these processes are poorly understood. Here, we provide an overview of the impact of mechanical forces on lymphatics and summarize the current understanding of the molecular mechanisms involved in the mechanosensation and mechanotransduction by lymphatic endothelial cells. We also discuss how these mechanosensitive pathways affect endothelial cell fate and regulate lymphatic development and function. A better understanding of these mechanisms may provide a deeper insight into the pathophysiology of various diseases associated with impaired lymphatic function, such as lymphedema and may eventually lead to the discovery of novel therapeutic targets for these conditions.

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