scholarly journals Golgi outposts locally regulate microtubule orientation in neurons but are not required for the overall polarity of the dendritic cytoskeleton

2019 ◽  
Author(s):  
Sihui Z. Yang ◽  
Jill Wildonger
Keyword(s):  
Matrix Protein ◽  
Cell Function ◽  
Microtubule Cytoskeleton ◽  
Microtubule Polarity ◽  
Drosophila Model ◽  
Ring Complex ◽  
Microtubule Growth ◽  
Microtubule Networks ◽  
Golgi Matrix ◽  
Do So

ABSTRACTMicrotubule-organizing centers (MTOCs) often play a central role in organizing the cellular microtubule networks that underlie cell function. In neurons, microtubules in axons and dendrites have distinct polarities. Dendrite-specific Golgi outposts, in particular multi-compartment outposts, have emerged as regulators of acentrosomal microtubule growth, raising the question of whether outposts contribute to establishing the overall polarity of the dendritic microtubule cytoskeleton. The cis-Golgi matrix protein GM130 has roles in both the MTOC activity of Golgi and in connecting Golgi compartments to form multi-compartment units. Using a combination of genetic approaches and live imaging in a Drosophila model, we found that GM130 is not essential for the overall polarity of the dendritic microtubule cytoskeleton. However, the mislocalization of multi-compartment Golgi outposts to axons disrupts the uniform orientation of axonal microtubules. This suggests that outposts have the capacity to influence microtubule polarity and, as our data indicate, likely do so independently of microtubule nucleation mediated by the γ-tubulin ring complex (γ-TuRC). Altogether, our results are consistent with the model that multi-compartment Golgi outposts may locally influence microtubule polarity, but that outposts are not necessary for the overall polarity of the dendritic microtubule cytoskeleton.

scholarly journals Golgi Outposts Locally Regulate Microtubule Orientation in Neurons but Are Not Required for the Overall Polarity of the Dendritic Cytoskeleton

Genetics ◽  
2020 ◽  
Vol 215 (2) ◽  
pp. 435-447 ◽  
Author(s):  
Sihui Z. Yang ◽  
Jill Wildonger
Keyword(s):  
Matrix Protein ◽  
Cell Function ◽  
Microtubule Nucleation ◽  
Microtubule Organization ◽  
Microtubule Orientation ◽  
Microtubule Polarity ◽  
Ring Complex ◽  
Microtubule Growth ◽  
Microtubule Networks ◽  
Golgi Matrix

Microtubule-organizing centers often play a central role in organizing the cellular microtubule networks that underlie cell function. In neurons, microtubules in axons and dendrites have distinct polarities. Dendrite-specific Golgi “outposts,” in particular multicompartment outposts, have emerged as regulators of acentrosomal microtubule growth, raising the question of whether outposts contribute to establishing or maintaining the overall polarity of the dendritic microtubule cytoskeleton. Using a combination of genetic approaches and live imaging in a Drosophila model, we found that dendritic microtubule polarity is unaffected by eliminating known regulators of Golgi-dependent microtubule organization including the cis-Golgi matrix protein GM130, the fly AKAP450 ortholog pericentrin-like protein, and centrosomin. This indicates that Golgi outposts are not essential for the formation or maintenance of a dendrite-specific cytoskeleton. However, the overexpression of GM130, which promotes the formation of ectopic multicompartment units, is sufficient to alter dendritic microtubule polarity. Axonal microtubule polarity is similarly disrupted by the presence of ectopic multicompartment Golgi outposts. Notably, multicompartment outposts alter microtubule polarity independently of microtubule nucleation mediated by the γ-tubulin ring complex. Thus, although Golgi outposts are not essential to dendritic microtubule polarity, altering their organization correlates with changes to microtubule polarity. Based on these data, we propose that the organization of Golgi outposts is carefully regulated to ensure proper dendritic microtubule polarity.

scholarly journals The Golgi matrix protein GM130: a specific interacting partner of the small GTPase rab1b

EMBO Reports ◽  
2001 ◽  
Vol 2 (4) ◽  
pp. 336-341 ◽  
Author(s):  
Thomas Weide ◽  
Michael Bayer ◽  
Miriam Köster ◽  
Jan‐Peter Siebrasse ◽  
Reiner Peters ◽  
...  
Keyword(s):  
Matrix Protein ◽  
Small Gtpase ◽  
Golgi Matrix

scholarly journals Biochemical reconstitutions reveal principles of human γ-TuRC assembly and function

2021 ◽  
Vol 220 (3) ◽  
Author(s):  
Michal Wieczorek ◽  
Shih-Chieh Ti ◽  
Linas Urnavicius ◽  
Kelly R. Molloy ◽  
Amol Aher ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Self Assembly ◽  
The Self ◽  
Dependent Manner ◽  
Guanine Nucleotide ◽  
Partial Cone ◽  
Ring Complex ◽  
Microtubule Networks ◽  
And Function

The formation of cellular microtubule networks is regulated by the γ-tubulin ring complex (γ-TuRC). This ∼2.3 MD assembly of >31 proteins includes γ-tubulin and GCP2-6, as well as MZT1 and an actin-like protein in a “lumenal bridge” (LB). The challenge of reconstituting the γ-TuRC has limited dissections of its assembly and function. Here, we report a biochemical reconstitution of the human γ-TuRC (γ-TuRC-GFP) as a ∼35 S complex that nucleates microtubules in vitro. In addition, we generate a subcomplex, γ-TuRCΔLB-GFP, which lacks MZT1 and actin. We show that γ-TuRCΔLB-GFP nucleates microtubules in a guanine nucleotide–dependent manner and with similar efficiency as the holocomplex. Electron microscopy reveals that γ-TuRC-GFP resembles the native γ-TuRC architecture, while γ-TuRCΔLB-GFP adopts a partial cone shape presenting only 8–10 γ-tubulin subunits and lacks a well-ordered lumenal bridge. Our results show that the γ-TuRC can be reconstituted using a limited set of proteins and suggest that the LB facilitates the self-assembly of regulatory interfaces around a microtubule-nucleating “core” in the holocomplex.

Hexosamines and TGF-β1 use similar signaling pathways to mediate matrix protein synthesis in mesangial cells

2004 ◽  
Vol 286 (2) ◽  
pp. F409-F416 ◽  
Author(s):  
Lalit P. Singh ◽  
Kenneith Green ◽  
Michelle Alexander ◽  
Shira Bassly ◽  
Errol D. Crook
Keyword(s):  
High Glucose ◽  
Matrix Protein ◽  
Transforming Growth Factor ◽  
Cell Function ◽  
Mesangial Cells ◽  
Rat Kidney ◽  
Protein Accumulation ◽  
Western Blots ◽  
Matrix Production ◽  
Mes Cells

Hyperglycemia-induced alterations in mesangial (MES) cell function and extracellular matrix (ECM) protein accumulation are seen in diabetic glomerulopathy. Transforming growth factor-β1 (TGF-β1) mediates high-glucose-induced matrix production in the kidney. Recent studies demonstrated that some of the effects of high glucose on cellular metabolism are mediated by the hexosamine biosynthesis pathway (HBP) in which fructose-6-phosphate is converted to glucosamine (GlcN) 6-phosphate. We previously showed that the high-glucose-mediated fibronectin and laminin synthesis in MES cells is mediated by the HBP and that GlcN is more potent than glucose in inducing TGF-β1 promoter luciferase activity. In this study, we investigated the hypothesis that the effects of glucose on MES matrix production occur via hexosamine regulation of TGF-β1. Culturing simian virus (SV)-40-transformed rat kidney MES cells in 25 mM glucose (HG) for 48 h increases cellular fibronectin and laminin levels about twofold on Western blots compared with low glucose (5 mM). GlcN (1.5 mM) or TGF-β1 (2.5-5 ng/ml) for 24-48 h also increases ECM synthesis. However, the effects of HG or GlcN with TGF-β1 are not additive. The presence of anti-TGF-β1 antibodies (20 μg/ml) blocks both TGF-β1- and GlcN-induced fibronectin synthesis. TGF-β1 increased ECM levels via PKA (laminin and fibronectin) and PKC (fibronectin) pathways. Similarly, TGF-β1 and hexosamines led to nonadditive increases in phosphorylation of the cAMP responsive element binding transcription factor. These results suggest that the effects of excess glucose on MES ECM synthesis occur via HBP-mediated regulation of TGF-β1.

Loss of the Golgi Matrix Protein 130 Cause Aberrant IgA1 Glycosylation in IgA Nephropathy

2019 ◽  
Vol 49 (4) ◽  
pp. 307-316 ◽  
Author(s):  
Chang Wang ◽  
Muyao Ye ◽  
Qiulan Zhao ◽  
Ming Xia ◽  
Di Liu ◽  
...  
Keyword(s):  
Iga Nephropathy ◽  
Matrix Protein ◽  
Mononuclear Cells ◽  
Lectin Binding ◽  
Underlying Mechanism ◽  
Sirna Transfection ◽  
Peripheral Blood Mononuclear ◽  
Vicia Villosa Lectin ◽  
Iga1 Glycosylation ◽  
Golgi Matrix

Background: Aberrant O-glycosylation IgA1 production is a major factor in the pathogenesis of IgA nephropathy, but the underlying mechanism is still unclear. IgA1 glycosylation modification is in Golgi, and downregulation of the Golgi peripheral membrane protein Golgi matrix protein 130 (GM130) could lead to glycosylation deficiency. In this study, we aimed to explore the role of GM130 in glycosylate deficiency IgA1 (Gd-IgA1) production. Methods: We enrolled 27 IgA nephropathy patients, 12 patients with chronic tonsillitis, 15 non-IgAN chronic kidney disease patients, and 15 healthy volunteers as healthy control. We explored GM130 expression in Tonsillar tissue by immunofluorescence staining and Western blotting and expression in peripheral blood mononuclear cells (PBMCs) by flow cytometry. The concentration of IgA1 and level of O-glycosylation were determined by ELISA and Vicia Villosa lectin-binding assay. Real-time PCR and Western blot were used to analyze the levels of β1,3-Gal transferase (C1GALT1) and ST6GalNAC2, respectively. To explore the contribution of GM130 in IgA1 O-glycosylation modification, cells were subjected to experiments for evaluation of GM130 silencing by GM130-siRNA transfection. Results: GM130 expression was significantly decreased in tonsil tissues and PBMC of IgAN patients; the expression of C1GALT1 decreased and Gd-IgA1 level increased significantly in patients with IgAN patients. The expression of GM130 was negatively related to Gd-IgA1 production. By siRNA transfection, our results clearly indicated that the downregulation of GM130 can increase IgA1 O-glycosylation deficiency, which is thought to reduce C1GALT1 expression but not affect the expression of ST6GalNAC2. Conclusion: We identified and demonstrated that GM130 plays an important role in IgA1 O-glycans deficiency in IgAN patients, by negatively regulating C1GALT1 expression. We believe that this finding will provide theoretical foundations for a new mechanism of Gd-IgA1 production in IgAN patients.

Identification and characterization of AtCASP, a plant transmembrane Golgi matrix protein

2005 ◽  
Vol 58 (1) ◽  
pp. 109-122 ◽  
Author(s):  
Luciana Renna ◽  
Sally L. Hanton ◽  
Giovanni Stefano ◽  
Lauren Bortolotti ◽  
Vikram Misra ◽  
...  
Keyword(s):  
Matrix Protein ◽  
Golgi Matrix ◽  
Identification And Characterization

scholarly journals 3D FIB-SEM reconstruction of microtubule–organelle interaction in whole primary mouse β cells

2020 ◽  
Vol 220 (2) ◽  
Author(s):  
Andreas Müller ◽  
Deborah Schmidt ◽  
C. Shan Xu ◽  
Song Pang ◽  
Joyson Verner D’Costa ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Cell Function ◽  
Secretory Granules ◽  
Detailed Knowledge ◽  
Β Cells ◽  
Microtubule Organization ◽  
Supportive Role ◽  
Primary Mouse ◽  
Microtubule Networks ◽  
Insulin Secretory Granule

Microtubules play a major role in intracellular trafficking of vesicles in endocrine cells. Detailed knowledge of microtubule organization and their relation to other cell constituents is crucial for understanding cell function. However, their role in insulin transport and secretion is under debate. Here, we use FIB-SEM to image islet β cells in their entirety with unprecedented resolution. We reconstruct mitochondria, Golgi apparati, centrioles, insulin secretory granules, and microtubules of seven β cells, and generate a comprehensive spatial map of microtubule–organelle interactions. We find that microtubules form nonradial networks that are predominantly not connected to either centrioles or endomembranes. Microtubule number and length, but not microtubule polymer density, vary with glucose stimulation. Furthermore, insulin secretory granules are enriched near the plasma membrane, where they associate with microtubules. In summary, we provide the first 3D reconstructions of complete microtubule networks in primary mammalian cells together with evidence regarding their importance for insulin secretory granule positioning and thus their supportive role in insulin secretion.

scholarly journals SIRT1 activity orchestrates ECM expression during hESC-chondrogenic differentiation

2020 ◽  
Author(s):  
Christopher A Smith ◽  
Paul A Humphreys ◽  
Nicola Bates ◽  
Mark A Naven ◽  
Stuart A Cain ◽  
...  
Keyword(s):  
Matrix Protein ◽  
Cell Function ◽  
Epigenetic Modification ◽  
Embryonic Stem ◽  
Type Ii Collagen ◽  
Cartilage Development ◽  
The Matrix ◽  
Col2a1 Expression ◽  
Key Driver

AbstractEpigenetic modification is a key driver of differentiation and the deacetylase Sirtuin1 (SIRT1) is an established regulator of cell function, ageing and articular cartilage homeostasis. Here we investigate the role of SIRT1 during development of chondrocytes by using human embryonic stem cells (hESCs). HESC-chondroprogenitors were treated with SIRT1 activator; SRT1720, or inhibitor; EX527, at different development stages. Activation of SIRT1 during 3D-pellet culture led to significant increases in expression of ECM genes for type-II collagen (COL2A1) and aggrecan (ACAN), and chondrogenic transcription factors SOX5 and ARID5B, with SOX5 ChIP analysis demonstrating enrichment on the ACAN –10 enhancer. Unexpectedly, while ACAN was enhanced, GAG retention in the matrix was reduced when SIRT1 was activated. Significantly, ARID5B and COL2A1 were positively correlated, with Co-IP indicating association of ARID5B with SIRT1 suggesting that COL2A1 expression is promoted by an ARID5B and SIRT1 interaction. In conclusion, SIRT1 activation positively impacts on the expression of the main ECM proteins, whilst altering ECM composition and suppressing GAG content during cartilage development. These results suggest that SIRT1 activity can be beneficial to cartilage development and matrix protein synthesis but tailored by addition of other positive GAG mediators.

Sign in / Sign up

Export Citation Format

Share Document