INVESTIGADORES
ANGEL Sergio Oscar
artículos
Título:
Plant Hsp90 proteins interact with B-cells and stimulate their proliferation
Autor/es:
CORIGLIANO M; MAGLIOCO A; LAGUIA BECHER M; GOLDMAN A; MARTIN V; ANGEL SO; CLEMENTE, M.
Revista:
PLOS ONE
Editorial:
PUBLIC LIBRARY SCIENCE
Referencias:
Año: 2011 vol. 6 p. 21231 - 21244
ISSN:
1932-6203
Resumen:
Abstract
Background: The molecular chaperone heat shock protein 90 (Hsp90) plays an important role in folding stabilization and
activation of client proteins. Besides, Hsp90 of mammals and mammalian pathogens displays immunostimulatory properties.
Here, we investigated the role of plant-derived Hsp90s as B-cell mitogens by measuring their proliferative responses in vitro.
activation of client proteins. Besides, Hsp90 of mammals and mammalian pathogens displays immunostimulatory properties.
Here, we investigated the role of plant-derived Hsp90s as B-cell mitogens by measuring their proliferative responses in vitro.
activation of client proteins. Besides, Hsp90 of mammals and mammalian pathogens displays immunostimulatory properties.
Here, we investigated the role of plant-derived Hsp90s as B-cell mitogens by measuring their proliferative responses in vitro.
The molecular chaperone heat shock protein 90 (Hsp90) plays an important role in folding stabilization and
activation of client proteins. Besides, Hsp90 of mammals and mammalian pathogens displays immunostimulatory properties.
Here, we investigated the role of plant-derived Hsp90s as B-cell mitogens by measuring their proliferative responses in vitro.in vitro.
Methodology: Plant cytosolic Hsp90 isoforms from Arabidopsis thaliana (AtHsp81.2) and Nicotiana benthamianaPlant cytosolic Hsp90 isoforms from Arabidopsis thaliana (AtHsp81.2) and Nicotiana benthamiana
(NbHsp90.3) were expressed in E. coli. Over-expression of recombinant plant Hsp90s (rpHsp90s) was confirmed by SDSPAGE
and western blot using and anti-AtHsp81.2 polyclonal anti-body. Both recombinant proteins were purified by Ni-NTA
affinity chromatography and their identity confirmed by MALDI-TOF-TOF. Recombinant AtHsp81.2 and NbHsp90.3 proteins
induced prominent proliferative responses in spleen cells form BALB/c mice. Polymyxin-B, a potent inhibitor of
lipopolysaccharide (LPS), did not eliminate the rpHsp90-induced proliferation. In addition, in vitro incubation of spleen cells
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
and western blot using and anti-AtHsp81.2 polyclonal anti-body. Both recombinant proteins were purified by Ni-NTA
affinity chromatography and their identity confirmed by MALDI-TOF-TOF. Recombinant AtHsp81.2 and NbHsp90.3 proteins
induced prominent proliferative responses in spleen cells form BALB/c mice. Polymyxin-B, a potent inhibitor of
lipopolysaccharide (LPS), did not eliminate the rpHsp90-induced proliferation. In addition, in vitro incubation of spleen cells
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
and western blot using and anti-AtHsp81.2 polyclonal anti-body. Both recombinant proteins were purified by Ni-NTA
affinity chromatography and their identity confirmed by MALDI-TOF-TOF. Recombinant AtHsp81.2 and NbHsp90.3 proteins
induced prominent proliferative responses in spleen cells form BALB/c mice. Polymyxin-B, a potent inhibitor of
lipopolysaccharide (LPS), did not eliminate the rpHsp90-induced proliferation. In addition, in vitro incubation of spleen cells
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
E. coli. Over-expression of recombinant plant Hsp90s (rpHsp90s) was confirmed by SDSPAGE
and western blot using and anti-AtHsp81.2 polyclonal anti-body. Both recombinant proteins were purified by Ni-NTA
affinity chromatography and their identity confirmed by MALDI-TOF-TOF. Recombinant AtHsp81.2 and NbHsp90.3 proteins
induced prominent proliferative responses in spleen cells form BALB/c mice. Polymyxin-B, a potent inhibitor of
lipopolysaccharide (LPS), did not eliminate the rpHsp90-induced proliferation. In addition, in vitro incubation of spleen cells
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
in vitro incubation of spleen cells
with rpHsp90 led to the expansion of CD19-bearing populations, suggesting a direct effect of these proteins on B
lymphocytes. This effect was confirmed by immunofluorescence analysis, where a direct binding of rpHsp90 to B- but not to
T-cells was observed in cells from BALB/c and C3H/HeN mice. Finally, we examined the involvement of Toll Like Receptor 4
(TLR4) molecules in the rpHsp90s induction of B-cell proliferation. Spleen cells from C3H/HeJ mice, which carry a point
mutation in the cytoplasmic region of TLR4, responded poorly to prAtHsp90. However, the interaction between rpHsp90
and B-cells from C3H/HeJ mice was not altered, suggesting that the mutation on TLR4 would be affecting the signal cascade
but not the rpHsp90-TLR4 receptor interaction.
Conclusions: Our results show for the first time that spleen cell proliferation can be stimulated by a non-pathogen-derived
Hsp90. Furthermore, our data provide a new example of a non-pathogen-derived ligand for TLRs.
Hsp90. Furthermore, our data provide a new example of a non-pathogen-derived ligand for TLRs.
Hsp90. Furthermore, our data provide a new example of a non-pathogen-derived ligand for TLRs.
Our results show for the first time that spleen cell proliferation can be stimulated by a non-pathogen-derived
Hsp90. Furthermore, our data provide a new example of a non-pathogen-derived ligand for TLRs.