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Regulation of translation is required for dendritic cell function and survival during activation.

Lelouard H, Schmidt EK, Camosseto V, Clavarino G, Ceppi M, Hsu HT, Pierre P - J. Cell Biol. (2007)

Bottom Line: In addition, we show that later on, in a manner similar to viral or apoptotic stress, DC activation leads to the phosphorylation and proteolysis of important translation initiation factors, thus inhibiting cap-dependent translation.This inhibition correlates with major changes in the origin of the peptides presented by MHC class I and the ability of mature DCs to prevent cell death.Our observations have important implications in linking translation regulation with DC function and survival during the immune response.

View Article: PubMed Central - PubMed

Affiliation: Centre d'Immunologie de Marseille-Luminy, Université de la Méditerranée, Case 906, 13288 Marseille, France.

ABSTRACT
In response to inflammatory stimulation, dendritic cells (DCs) have a remarkable pattern of differentiation (maturation) that exhibits specific mechanisms to control antigen processing and presentation. Here, we show that in response to lipopolysaccharides, protein synthesis is rapidly enhanced in DCs. This enhancement occurs via a PI3K-dependent signaling pathway and is key for DC activation. In addition, we show that later on, in a manner similar to viral or apoptotic stress, DC activation leads to the phosphorylation and proteolysis of important translation initiation factors, thus inhibiting cap-dependent translation. This inhibition correlates with major changes in the origin of the peptides presented by MHC class I and the ability of mature DCs to prevent cell death. Our observations have important implications in linking translation regulation with DC function and survival during the immune response.

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Up-regulation and cleavage of eIF4GI during DC maturation. (A) Scheme of the murine eIF4GI largest isoform. Binding sites for PABP and the different translation initiation factors are boxed. The caspase-3 cleavage sites are indicated with arrowheads. Epitopes recognized by the different antibodies used throughout this study are organized according to their position in eIF4GI. (B) Conversely to eIF4GII (middle) and eIF4A (bottom), eIF4GI and associated fragments (top) are strongly increased after 8 h of maturation as monitored by immunoblot performed with α-C-FAG antibody. (C) Model of eIF4GI cleavage in mDCs. Extracts of mDCs were separated by SDS-PAGE and immunoblotted with different anti-eIF4GI antibodies. Fragment patterns were compared as follows: bands recognized by only one antibody were eliminated; other bands were analyzed according to antibody reactivity (illustrated by asterisks on the right of the immunoblot), molecular weight and comparison to the previously described cleavage products generated by caspase-3, proteasome, picornaviral, and HIV virus proteases. Three major proteolytic sensitive areas (gray boxes) and their putative products were deduced from antibody reactivity and molecular weights. NFc stands for N-terminal fragment cluster because several clustered bands are always displayed due to the existence of the multiple isoforms of eIF4GI. MF stands for middle fragment and CF for C-terminal fragment. (D) Alteration of eIF4GI during DC maturation is prevented by proteasome and pan-caspase inhibitors. Effects of protease inhibitors on eIF4GI fate in DCs were analyzed by immunoblot using the anti- C-FAG antibody. Conversely to MG132 and Z-VAD-FMK, specific caspase inhibitors (YVAD, DEVD, IETD, LEHD, WEHD) had no effect on accumulation of eIF4GI fragments.
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fig5: Up-regulation and cleavage of eIF4GI during DC maturation. (A) Scheme of the murine eIF4GI largest isoform. Binding sites for PABP and the different translation initiation factors are boxed. The caspase-3 cleavage sites are indicated with arrowheads. Epitopes recognized by the different antibodies used throughout this study are organized according to their position in eIF4GI. (B) Conversely to eIF4GII (middle) and eIF4A (bottom), eIF4GI and associated fragments (top) are strongly increased after 8 h of maturation as monitored by immunoblot performed with α-C-FAG antibody. (C) Model of eIF4GI cleavage in mDCs. Extracts of mDCs were separated by SDS-PAGE and immunoblotted with different anti-eIF4GI antibodies. Fragment patterns were compared as follows: bands recognized by only one antibody were eliminated; other bands were analyzed according to antibody reactivity (illustrated by asterisks on the right of the immunoblot), molecular weight and comparison to the previously described cleavage products generated by caspase-3, proteasome, picornaviral, and HIV virus proteases. Three major proteolytic sensitive areas (gray boxes) and their putative products were deduced from antibody reactivity and molecular weights. NFc stands for N-terminal fragment cluster because several clustered bands are always displayed due to the existence of the multiple isoforms of eIF4GI. MF stands for middle fragment and CF for C-terminal fragment. (D) Alteration of eIF4GI during DC maturation is prevented by proteasome and pan-caspase inhibitors. Effects of protease inhibitors on eIF4GI fate in DCs were analyzed by immunoblot using the anti- C-FAG antibody. Conversely to MG132 and Z-VAD-FMK, specific caspase inhibitors (YVAD, DEVD, IETD, LEHD, WEHD) had no effect on accumulation of eIF4GI fragments.

Mentions: Alternatively, cap-dependent translation can also be inhibited by the cleavage of the scaffold translation initiation factor eIF4GI (Prevot et al., 2003; Holcik and Sonenberg, 2005; Spriggs et al., 2005) (Fig. 5 A). Proteolytic cleavage of eIF4GI by viral proteases or caspase-3 occurs during picornavirus and retrovirus infection as well as during cellular stress (Holcik and Sonnenberg, 2005). Proteolytic fragments of eIF4GI have been shown to compete with the recruitment of the complete cap-binding protein complex eIF4F by the mRNAs, thereby limiting cap-dependent translation and favoring the internal ribosome entry sites (IRES)–mediated translation of a small number of specific mRNAs (Bushell et al., 2000). Thus, we investigated by immunoblotting the fate of the scaffold translation initiation factor eIF4GI using an antibody recognizing a C-terminal epitope (α-C-FAG) (Cowan and Morley, 2004). Surprisingly, between 8 and 14 h of LPS activation, a time at which cap-dependent translation is down-regulated, eIF4GI levels increased sturdily (Fig. 5 B). In contrast, the levels of the closely related homologue eIF4GII and of eIF4A remained stable. The specific up-regulation of eIF4GI does probably not reflect a slower degradation rate because several smaller fragments weakly present in iDCs also increased after 8 h of LPS treatment. Interestingly, eIF4GI translation is enhanced in a context of general cap-dependent translation inhibition.


Regulation of translation is required for dendritic cell function and survival during activation.

Lelouard H, Schmidt EK, Camosseto V, Clavarino G, Ceppi M, Hsu HT, Pierre P - J. Cell Biol. (2007)

Up-regulation and cleavage of eIF4GI during DC maturation. (A) Scheme of the murine eIF4GI largest isoform. Binding sites for PABP and the different translation initiation factors are boxed. The caspase-3 cleavage sites are indicated with arrowheads. Epitopes recognized by the different antibodies used throughout this study are organized according to their position in eIF4GI. (B) Conversely to eIF4GII (middle) and eIF4A (bottom), eIF4GI and associated fragments (top) are strongly increased after 8 h of maturation as monitored by immunoblot performed with α-C-FAG antibody. (C) Model of eIF4GI cleavage in mDCs. Extracts of mDCs were separated by SDS-PAGE and immunoblotted with different anti-eIF4GI antibodies. Fragment patterns were compared as follows: bands recognized by only one antibody were eliminated; other bands were analyzed according to antibody reactivity (illustrated by asterisks on the right of the immunoblot), molecular weight and comparison to the previously described cleavage products generated by caspase-3, proteasome, picornaviral, and HIV virus proteases. Three major proteolytic sensitive areas (gray boxes) and their putative products were deduced from antibody reactivity and molecular weights. NFc stands for N-terminal fragment cluster because several clustered bands are always displayed due to the existence of the multiple isoforms of eIF4GI. MF stands for middle fragment and CF for C-terminal fragment. (D) Alteration of eIF4GI during DC maturation is prevented by proteasome and pan-caspase inhibitors. Effects of protease inhibitors on eIF4GI fate in DCs were analyzed by immunoblot using the anti- C-FAG antibody. Conversely to MG132 and Z-VAD-FMK, specific caspase inhibitors (YVAD, DEVD, IETD, LEHD, WEHD) had no effect on accumulation of eIF4GI fragments.
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Related In: Results  -  Collection

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fig5: Up-regulation and cleavage of eIF4GI during DC maturation. (A) Scheme of the murine eIF4GI largest isoform. Binding sites for PABP and the different translation initiation factors are boxed. The caspase-3 cleavage sites are indicated with arrowheads. Epitopes recognized by the different antibodies used throughout this study are organized according to their position in eIF4GI. (B) Conversely to eIF4GII (middle) and eIF4A (bottom), eIF4GI and associated fragments (top) are strongly increased after 8 h of maturation as monitored by immunoblot performed with α-C-FAG antibody. (C) Model of eIF4GI cleavage in mDCs. Extracts of mDCs were separated by SDS-PAGE and immunoblotted with different anti-eIF4GI antibodies. Fragment patterns were compared as follows: bands recognized by only one antibody were eliminated; other bands were analyzed according to antibody reactivity (illustrated by asterisks on the right of the immunoblot), molecular weight and comparison to the previously described cleavage products generated by caspase-3, proteasome, picornaviral, and HIV virus proteases. Three major proteolytic sensitive areas (gray boxes) and their putative products were deduced from antibody reactivity and molecular weights. NFc stands for N-terminal fragment cluster because several clustered bands are always displayed due to the existence of the multiple isoforms of eIF4GI. MF stands for middle fragment and CF for C-terminal fragment. (D) Alteration of eIF4GI during DC maturation is prevented by proteasome and pan-caspase inhibitors. Effects of protease inhibitors on eIF4GI fate in DCs were analyzed by immunoblot using the anti- C-FAG antibody. Conversely to MG132 and Z-VAD-FMK, specific caspase inhibitors (YVAD, DEVD, IETD, LEHD, WEHD) had no effect on accumulation of eIF4GI fragments.
Mentions: Alternatively, cap-dependent translation can also be inhibited by the cleavage of the scaffold translation initiation factor eIF4GI (Prevot et al., 2003; Holcik and Sonenberg, 2005; Spriggs et al., 2005) (Fig. 5 A). Proteolytic cleavage of eIF4GI by viral proteases or caspase-3 occurs during picornavirus and retrovirus infection as well as during cellular stress (Holcik and Sonnenberg, 2005). Proteolytic fragments of eIF4GI have been shown to compete with the recruitment of the complete cap-binding protein complex eIF4F by the mRNAs, thereby limiting cap-dependent translation and favoring the internal ribosome entry sites (IRES)–mediated translation of a small number of specific mRNAs (Bushell et al., 2000). Thus, we investigated by immunoblotting the fate of the scaffold translation initiation factor eIF4GI using an antibody recognizing a C-terminal epitope (α-C-FAG) (Cowan and Morley, 2004). Surprisingly, between 8 and 14 h of LPS activation, a time at which cap-dependent translation is down-regulated, eIF4GI levels increased sturdily (Fig. 5 B). In contrast, the levels of the closely related homologue eIF4GII and of eIF4A remained stable. The specific up-regulation of eIF4GI does probably not reflect a slower degradation rate because several smaller fragments weakly present in iDCs also increased after 8 h of LPS treatment. Interestingly, eIF4GI translation is enhanced in a context of general cap-dependent translation inhibition.

Bottom Line: In addition, we show that later on, in a manner similar to viral or apoptotic stress, DC activation leads to the phosphorylation and proteolysis of important translation initiation factors, thus inhibiting cap-dependent translation.This inhibition correlates with major changes in the origin of the peptides presented by MHC class I and the ability of mature DCs to prevent cell death.Our observations have important implications in linking translation regulation with DC function and survival during the immune response.

View Article: PubMed Central - PubMed

Affiliation: Centre d'Immunologie de Marseille-Luminy, Université de la Méditerranée, Case 906, 13288 Marseille, France.

ABSTRACT
In response to inflammatory stimulation, dendritic cells (DCs) have a remarkable pattern of differentiation (maturation) that exhibits specific mechanisms to control antigen processing and presentation. Here, we show that in response to lipopolysaccharides, protein synthesis is rapidly enhanced in DCs. This enhancement occurs via a PI3K-dependent signaling pathway and is key for DC activation. In addition, we show that later on, in a manner similar to viral or apoptotic stress, DC activation leads to the phosphorylation and proteolysis of important translation initiation factors, thus inhibiting cap-dependent translation. This inhibition correlates with major changes in the origin of the peptides presented by MHC class I and the ability of mature DCs to prevent cell death. Our observations have important implications in linking translation regulation with DC function and survival during the immune response.

Show MeSH
Related in: MedlinePlus