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Nestin modulates glucocorticoid receptor function by cytoplasmic anchoring.

Reimer R, Helmbold H, Szalay B, Hagel C, Hohenberg H, Deppert W, Bohn W - PLoS ONE (2009)

Bottom Line: The reaction pattern with phospho-GR specific antibodies and the presence of the chaperone HSC70 suggest that specifically the unliganded receptor is anchored to the IF system.Ligand addition releases GR from IFs and shifts the receptor into the nucleus.The data give evidence that nestin/vimentin specific anchoring modulates growth suppression by GR.

View Article: PubMed Central - PubMed

Affiliation: Heinrich-Pette-Institute for Experimental Virology and Immunology at the University of Hamburg, Hamburg, Germany.

ABSTRACT
Nestin is the characteristic intermediate filament (IF) protein of rapidly proliferating progenitor cells and regenerating tissue. Nestin copolymerizes with class III IF-proteins, mostly vimentin, into heteromeric filaments. Its expression is downregulated with differentiation. Here we show that a strong nestin expression in mouse embryo tissue coincides with a strong accumulation of the glucocorticoid receptor (GR), a key regulator of growth and differentiation in embryonic development. Microscopic studies on cultured cells show an association of GR with IFs composed of vimentin and nestin. Cells lacking nestin, but expressing vimentin, or cells expressing vimentin, but lacking nestin accumulate GR in the nucleus. Completing these networks with an exogenous nestin, respectively an exogenous vimentin restores cytoplasmic anchoring of GR to the IF system. Thus, heteromeric filaments provide the basis for anchoring of GR. The reaction pattern with phospho-GR specific antibodies and the presence of the chaperone HSC70 suggest that specifically the unliganded receptor is anchored to the IF system. Ligand addition releases GR from IFs and shifts the receptor into the nucleus. Suppression of nestin by specific shRNA abolishes anchoring of GR, induces its accumulation in the nucleus and provokes an irreversible G1/S cell cycle arrest. Suppression of GR prior to that of nestin prevents entry into the arrest. The data give evidence that nestin/vimentin specific anchoring modulates growth suppression by GR. We hypothesize that expression of nestin is a major determinant in suppression of anti-proliferative activity of GR in undifferentiated tissue and facilitates activation of this growth control in a precise tissue and differentiation dependent manner.

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Related in: MedlinePlus

GR redistributes with shortened IFs composed of a tailless vimentin protein.(A–D) Confocal images of cytoskeletons prepared from B8 VimMT3B4 cells, which stably express a tailless vimentin protein; tailless vimentin forms a fragmented IF network (A, B); the filaments contain also nestin (C); GR colocalizes with vimentin/nestin filament fragments (A, C); Bars in A and C = 1 µm; B = 5 µm.
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pone-0006084-g007: GR redistributes with shortened IFs composed of a tailless vimentin protein.(A–D) Confocal images of cytoskeletons prepared from B8 VimMT3B4 cells, which stably express a tailless vimentin protein; tailless vimentin forms a fragmented IF network (A, B); the filaments contain also nestin (C); GR colocalizes with vimentin/nestin filament fragments (A, C); Bars in A and C = 1 µm; B = 5 µm.

Mentions: Further to substantiate the association of GR with IFs we asked whether altering the structure of the vimentin filament network would induce a coincident rearrangement of GR. We transfected the vimentin deficient C6D10 cells with an expression vector coding for a C-terminal truncated vimentin protein (ct-vimentin). Although not essential for the polymerization itself the tails seem to stabilize the lateral interaction of vimentin filament subunits. In addition, they seem to mediate the structural interaction of vimentin with other major cytoskeleton filament systems [34]. As shown in Figure 7A–C tailless vimentin was still capable of forming a filament network, but this was composed of shortened filaments and less well organized. GR colocalized with these shortened vimentin filaments. High resolution confocal images showed that the GR label did not evenly decorate the vimentin filaments but exhibited a spotty distribution (Figure 7, A3). A similar staining pattern was obtained in double labeling of nestin and GR (Figure 7C).


Nestin modulates glucocorticoid receptor function by cytoplasmic anchoring.

Reimer R, Helmbold H, Szalay B, Hagel C, Hohenberg H, Deppert W, Bohn W - PLoS ONE (2009)

GR redistributes with shortened IFs composed of a tailless vimentin protein.(A–D) Confocal images of cytoskeletons prepared from B8 VimMT3B4 cells, which stably express a tailless vimentin protein; tailless vimentin forms a fragmented IF network (A, B); the filaments contain also nestin (C); GR colocalizes with vimentin/nestin filament fragments (A, C); Bars in A and C = 1 µm; B = 5 µm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2698154&req=5

pone-0006084-g007: GR redistributes with shortened IFs composed of a tailless vimentin protein.(A–D) Confocal images of cytoskeletons prepared from B8 VimMT3B4 cells, which stably express a tailless vimentin protein; tailless vimentin forms a fragmented IF network (A, B); the filaments contain also nestin (C); GR colocalizes with vimentin/nestin filament fragments (A, C); Bars in A and C = 1 µm; B = 5 µm.
Mentions: Further to substantiate the association of GR with IFs we asked whether altering the structure of the vimentin filament network would induce a coincident rearrangement of GR. We transfected the vimentin deficient C6D10 cells with an expression vector coding for a C-terminal truncated vimentin protein (ct-vimentin). Although not essential for the polymerization itself the tails seem to stabilize the lateral interaction of vimentin filament subunits. In addition, they seem to mediate the structural interaction of vimentin with other major cytoskeleton filament systems [34]. As shown in Figure 7A–C tailless vimentin was still capable of forming a filament network, but this was composed of shortened filaments and less well organized. GR colocalized with these shortened vimentin filaments. High resolution confocal images showed that the GR label did not evenly decorate the vimentin filaments but exhibited a spotty distribution (Figure 7, A3). A similar staining pattern was obtained in double labeling of nestin and GR (Figure 7C).

Bottom Line: The reaction pattern with phospho-GR specific antibodies and the presence of the chaperone HSC70 suggest that specifically the unliganded receptor is anchored to the IF system.Ligand addition releases GR from IFs and shifts the receptor into the nucleus.The data give evidence that nestin/vimentin specific anchoring modulates growth suppression by GR.

View Article: PubMed Central - PubMed

Affiliation: Heinrich-Pette-Institute for Experimental Virology and Immunology at the University of Hamburg, Hamburg, Germany.

ABSTRACT
Nestin is the characteristic intermediate filament (IF) protein of rapidly proliferating progenitor cells and regenerating tissue. Nestin copolymerizes with class III IF-proteins, mostly vimentin, into heteromeric filaments. Its expression is downregulated with differentiation. Here we show that a strong nestin expression in mouse embryo tissue coincides with a strong accumulation of the glucocorticoid receptor (GR), a key regulator of growth and differentiation in embryonic development. Microscopic studies on cultured cells show an association of GR with IFs composed of vimentin and nestin. Cells lacking nestin, but expressing vimentin, or cells expressing vimentin, but lacking nestin accumulate GR in the nucleus. Completing these networks with an exogenous nestin, respectively an exogenous vimentin restores cytoplasmic anchoring of GR to the IF system. Thus, heteromeric filaments provide the basis for anchoring of GR. The reaction pattern with phospho-GR specific antibodies and the presence of the chaperone HSC70 suggest that specifically the unliganded receptor is anchored to the IF system. Ligand addition releases GR from IFs and shifts the receptor into the nucleus. Suppression of nestin by specific shRNA abolishes anchoring of GR, induces its accumulation in the nucleus and provokes an irreversible G1/S cell cycle arrest. Suppression of GR prior to that of nestin prevents entry into the arrest. The data give evidence that nestin/vimentin specific anchoring modulates growth suppression by GR. We hypothesize that expression of nestin is a major determinant in suppression of anti-proliferative activity of GR in undifferentiated tissue and facilitates activation of this growth control in a precise tissue and differentiation dependent manner.

Show MeSH
Related in: MedlinePlus