A Lipid Droplet-Specific Capsule Is Present in Rat Adrenal-Cells: Evidence from a Monoclonal Antibody

Wang, Seu Mei; Fong, Tsorng Harn

doi:10.1006/bbrc.1995.2748

Cited by 22 publications

(39 citation statements)

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“…To date, a very limited group of proteins have been shown to associate with LBs; perilipins in adipocytes and steroidogenic tissues (Greenberg et al, 1991), adipophilin in most mammalian cells (Heid et al, 1998), oleosins in plants (Qu and Huang, 1990), eicosanoid synthesizing enzymes in various types of cells (Weller and Dvorak, 1994), vimentin (Franke et al, 1987), "capsular proteins" in adrenal cells (Wang and Fong, 1995), and more recently, caveolins (Fujimoto et al, 2001;Ostermeyer et al, 2001;Pol et al, 2001). Although the LB targeting domain of some of these proteins has been examined, a consensus sequence or molecular determinant involved in LB association has not been described.…”

Section: Introductionmentioning

confidence: 99%

Dynamic and Regulated Association of Caveolin with Lipid Bodies: Modulation of Lipid Body Motility and Function by a Dominant Negative Mutant

Pol

Martin

Fernández

et al. 2004

MBoC

186

175

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Caveolins are a crucial component of caveolae but have also been localized to the Golgi complex, and, under some experimental conditions, to lipid bodies (LBs). The physiological relevance and dynamics of LB association remain unclear. We now show that endogenous caveolin-1 and caveolin-2 redistribute to LBs in lipid loaded A431 and FRT cells. Association with LBs is regulated and reversible; removal of fatty acids causes caveolin to rapidly leave the lipid body. We also show by subcellular fractionation, light and electron microscopy that during the first hours of liver regeneration, caveolins show a dramatic redistribution from the cell surface to the newly formed LBs. At later stages of the regeneration process (when LBs are still abundant), the levels of caveolins in LBs decrease dramatically. As a model system to study association of caveolins with LBs we have used brefeldin A (BFA). BFA causes rapid redistribution of endogenous caveolins to LBs and this association was reversed upon BFA washout. Finally, we have used a dominant negative LB-associated caveolin mutant (cav DGV ) to study LB formation and to examine its effect on LB function. We now show that the cav DGV mutant inhibits microtubule-dependent LB motility and blocks the reversal of lipid accumulation in LBs.

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Section: Introductionmentioning

confidence: 99%

Dynamic and Regulated Association of Caveolin with Lipid Bodies: Modulation of Lipid Body Motility and Function by a Dominant Negative Mutant

Pol

Martin

Fernández

et al. 2004

MBoC

186

175

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“…Primary cultures of rat adrenocortical cells were established as previously described, with a few modifications [Wang and Fong, 1995]. Briefly, isolated cells were obtained by incubating adrenal gland fragments in serum-free M199 medium, containing 1 mg/ml of collagenase (Type II; Sigma, St. Louis, MO), for 30 min at 37°C, with gentle shaking, followed by mechanical dispersion by aspiration using a glass pipette.…”

Section: Materials and Methods Cell Culturementioning

confidence: 99%

“…The method used and the preparation of antibodies were as previously described [Wang and Fong, 1995].…”

Section: Immunofluorescence Microscopymentioning

confidence: 99%

“…We have reported a specific monoclonal antibody (mAb), A2, which identifies a 160-kD capsular protein on lipid droplets in rat adrenocortical cells [Wang and Fong, 1995]. Interestingly, the capsular staining of the lipid droplet surface is lost in response to ACTH stimulation [Wang and Fong, 1995].…”

mentioning

confidence: 97%

“…Interestingly, the capsular staining of the lipid droplet surface is lost in response to ACTH stimulation [Wang and Fong, 1995]. In the present study, our aims were to elucidate the signal transduction pathway for this ACTH-induced decapsulation of lipid droplets and the relationship between decapsulation of lipid droplets and lipolysis.…”

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Dissection of the signaling mechanism for capsule detachment of lipid droplets in rat adrenocortical cells

Fong

Wang

1997

J. Cell. Biochem.

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In a previous study, we used a monoclonal antibody, A2, to demonstrate the presence of the lipid droplet-specific capsule in adrenocortical cells and the stimulation of steroid secretion with adrenocorticotrophic hormone (ACTH), resulting in the detachment of this capsule from the droplet surface into the cytosol. To investigate the signaling pathway for this event, we tested the role of adenylate cyclase, cAMP, and protein kinases A and C (PKA and PKC) in this response. ACTH-induced decapsulation of lipid droplets was blocked by either adenylate cyclase inhibitor or PKA inhibitor and stimulated by Bt2cAMP. We conclude that the signaling mechanism involved in lipid droplet decapsulation is the cascade consisting of adenylate cyclase activation, cAMP elevation, and subsequent PKA activation. Furthermore, the cytosolic detached capsular protein was able to relocate to the lipid droplet surface on cessation of ACTH or Bt2cAMP stimulation. In addition to PKA-mediated decapsulation, inhibition of PKC by calphostin C alone was enough to induce decapsulation, a process that was independent of PKA activity, whereas activation of PKC could prevent Bt2cAMP-induced decapsulation. A cAMP radioimmunoassay also confirmed that ACTH caused a marked increase in intracellular levels of cAMP, while PMA or calphostin C caused no significant changes. We conclude that PKA and PKC are reciprocally operated to regulate the decapsulation of lipid droplets, the same mechanism adopted in steroidogenesis. A time-course study also indicates that decapsulation of lipid droplets was accompanied by detectable changes in the size and the area of lipid droplets upon the stimulation of Bt2cAMP or calphostin C, implying a possible coupling between the capsule detachment and steroidogenesis.

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Immunocytochemical demonstration of a lipid droplet-specific capsule in cultured Leydig cells of the golden hamsters

1996

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In this report, we provide direct evidence for the presence of a lipid droplet-associated capsule in hamster steroidogenic Leydig cells by using a monoclonal antibody A2. Leydig cells are characterized by containing many lipid droplets and having 3 beta-hydroxysteroid dehydrogenase activity. Immunofluorescence staining with this antibody demonstrated a rim or capsule surrounding the lipid droplets in Leydig cells, a pattern not seen with anti-vimentin antibody. Immunogold labelling confirmed ultrastructurally that antibody binding was distributed on the lipid droplet surface. In order to investigate the possible function of the capsule, we examined the morphological changes induced in the capsule following stimulation with LH or dibutyryl cAMP; the fluorescent intensity of the capsule was seen to gradually decrease, accompanied by a decrease in number and size of lipid droplets, and the response to both reagents was time- and concentration-dependent. We thus conclude that hormonal stimulation resulting in the detachment of certain capsular proteins from the surface of lipid droplets is mediated via the cAMP signaling pathway and may allow cholesterol ester hydrolytic enzyme direct access to its substrate in the lipid droplet.

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A Lipid Droplet-Specific Capsule Is Present in Rat Adrenal-Cells: Evidence from a Monoclonal Antibody

Cited by 22 publications

References 0 publications

Dynamic and Regulated Association of Caveolin with Lipid Bodies: Modulation of Lipid Body Motility and Function by a Dominant Negative Mutant

Dynamic and Regulated Association of Caveolin with Lipid Bodies: Modulation of Lipid Body Motility and Function by a Dominant Negative Mutant

Dissection of the signaling mechanism for capsule detachment of lipid droplets in rat adrenocortical cells

Immunocytochemical demonstration of a lipid droplet-specific capsule in cultured Leydig cells of the golden hamsters

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