Cryopreparation Provides New Insight into the Effects of Brefeldin A on the Structure of the HepG2 Golgi Apparatus

Hess, Michael W.; Müller, Martin; Debbage, Paul; Vetterlein, M.; Pavelka, Margit

doi:10.1006/jsbi.2000.4230

Cited by 73 publications

(45 citation statements)

References 28 publications

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“…2B). This ER resembles the specialized trans ER characterized in fast frozen͞freeze-substituted normal rat kidney and HepG2 cells (6,16). It is noteworthy that the ER, which flattens against C5, is directly continuous with that which appears to be intimately associated with mitochondria situated near to the Golgi (Figs.…”

Section: -D Reconstructionsmentioning

confidence: 53%

Organellar relationships in the Golgi region of the pancreatic beta cell line, HIT-T15, visualized by high resolution electron tomography

Marsh

Mastronarde

Buttle

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

328

299

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Section: -D Reconstructionsmentioning

confidence: 53%

Organellar relationships in the Golgi region of the pancreatic beta cell line, HIT-T15, visualized by high resolution electron tomography

Marsh

Mastronarde

Buttle

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

328

299

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“…For transmission EM the samples were subjected to standard chemical fixation (glutaraldehyde, OsO 4 ) or to cryo-fixation by means of high-pressure freezing and freeze substitution [Hess et al, 2000] that greatly improves ultrastructure preservation. EMcytochemistry for PAS-positive substances was performed according to Thiéry [1967].…”

Section: Emmentioning

confidence: 99%

Loss-of-function of MYO5B is the main cause of microvillus inclusion disease: 15 novel mutations and a CaCo-2 RNAi cell model

et al. 2010

Self Cite

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Autosomal recessive microvillus inclusion disease (MVID) is characterized by an intractable diarrhea starting within the first few weeks of life. The hallmarks of MVID are a lack of microvilli on the surface of villous enterocytes, occurrence of intracellular vacuoles lined by microvilli (microvillus inclusions), and the cytoplasmic accumulation of periodic acid-Schiff (PAS)-positive vesicles in enterocytes. Recently, we identified mutations in MYO5B, encoding the unconventional type Vb myosin motor protein, in a first cohort of nine MVID patients. In this study, we identified 15 novel nonsense and missense mutations in MYO5B in 11 unrelated MVID patients. Fluorescence microscopy, Western blotting, and electron microscopy were applied to analyze the effects of MYO5B siRNA knockdown in polarized, brush border possessing CaCo-2 cells. Loss of surface microvilli, increased formation of microvillus inclusions, and subapical enrichment of PAS-positive endomembrane compartments were induced in polarized, filter-grown CaCo-2 cells, following MYO5B knock-down. Our data indicate that MYO5B mutations are a major cause of microvillus inclusion disease and that MYO5B knockdown recapitulates most of the cellular phenotype in vitro, thus independently showing loss of MYO5B function as the cause of microvillus inclusion disease.

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“…1B, left BFA has been a useful tool in studying protein trafficking in the secretory pathway because this drug induced Golgi stacks to form enlarged compartments and prevented protein traffic from the ER to the Golgi (Jiang and Rogers, 1998;Hess et al, 2000;Ritzenthaler et al, 2002). Using transgenic tobacco BY-2 cell lines expressing Golgi and PVC reporters, we have recently demonstrated that Golgi and PVC had different sensitivity to wortmannin in which the drug induced the PVC, but not the Golgi, to dilate (Tse et al, 2004).…”

Section: Development Of the Gfp Bp-80 Transgenic Cell Linementioning

confidence: 99%

“…In addition, BFA may also act on endosomes and induce endosomes to become tubulated in mammalian cells (Hunziker et al, 1991;LippincottSchwartz et al, 1991;Wood et al, 1991). In plant cells, BFA also induced the Golgi apparatus to form ER-Golgi hybrid structures , BFA compartments (Satiat-Jeunemaitre et al, 1996;Wee et al, 1998;Baldwin et al, 2001;Tse et al, 2004), or the loss of Golgi cis-cisternae (Hess et al, 2000). In addition, BFA may induce the trans-Golgi network to incorporate into BFA compartments in Arabidopsis root cells (Dettmer et al, 2006).…”

mentioning

confidence: 99%

Dynamic Response of Prevacuolar Compartments to Brefeldin A in Plant Cells

Tse

Hillmer

et al. 2006

Plant Physiology

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Little is known about the dynamics and molecular components of plant prevacuolar compartments (PVCs) in the secretory pathway. Using transgenic tobacco (Nicotiana tabacum) Bright-Yellow-2 (BY-2) cells expressing membrane-anchored yellow fluorescent protein (YFP) reporters marking Golgi or PVCs, we have recently demonstrated that PVCs are mobile multivesicular bodies defined by vacuolar sorting receptor proteins. Here, we demonstrate that Golgi and PVCs have different sensitivity in response to brefeldin A (BFA) treatment in living tobacco BY-2 cells. BFA at low concentrations (5-10 mg mL 21 ) induced YFPmarked Golgi stacks to form both endoplasmic reticulum-Golgi hybrid structures and BFA-induced aggregates, but had little effect on YFP-marked PVCs in transgenic BY-2 cells at both confocal and immunogold electron microscopy levels. However, BFA at high concentrations (50-100 mg mL 21 ) caused both YFP-marked Golgi stacks and PVCs to form aggregates in a dose-and timedependent manner. Normal Golgi or PVC signals can be recovered upon removal of BFA from the culture media. Confocal immunofluorescence and immunogold electron microscopy studies with specific organelle markers further demonstrate that the PVC aggregates are distinct, but physically associated, with Golgi aggregates in BFA-treated cells and that PVCs might lose their internal vesicle structures at high BFA concentration. In addition, vacuolar sorting receptor-marked PVCs in root-tip cells of tobacco, pea (Pisum sativum), mung bean (Vigna radiata), and Arabidopsis (Arabidopsis thaliana) upon BFA treatment are also induced to form similar aggregates. Thus, we have demonstrated that the effects of BFA are not limited to endoplasmic reticulum and Golgi, but extend to PVC in the endomembrane system, which might provide a quick tool for distinguishing Golgi from PVC for its identification and characterization, as well as a possible new tool in studying PVC-mediated protein traffic in plant cells.

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Cryopreparation Provides New Insight into the Effects of Brefeldin A on the Structure of the HepG2 Golgi Apparatus

Cited by 73 publications

References 28 publications

Organellar relationships in the Golgi region of the pancreatic beta cell line, HIT-T15, visualized by high resolution electron tomography

Organellar relationships in the Golgi region of the pancreatic beta cell line, HIT-T15, visualized by high resolution electron tomography

Loss-of-function of MYO5B is the main cause of microvillus inclusion disease: 15 novel mutations and a CaCo-2 RNAi cell model

Dynamic Response of Prevacuolar Compartments to Brefeldin A in Plant Cells

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