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Escherichia coli SRP, its protein subunit Ffh, and the Ffh M domain are able to selectively limit membrane protein expression when overexpressed.

Yosef I, Bochkareva ES, Bibi E - MBio (2010)

Bottom Line: The results show that SRP, Ffh, and the M domain are all able to selectively inhibit the expression of membrane proteins.We observed no apparent changes in the steady-state mRNA levels or membrane protein stability, suggesting that inhibition may occur at the level of translation, possibly through the interaction between Ffh and ribosome-hydrophobic nascent chain complexes.Since E. coli SRP does not have a eukaryote-like translation arrest domain, we discuss other possible mechanisms by which this SRP might regulate membrane protein translation when overexpressed.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, Israel.

ABSTRACT
The Escherichia coli signal recognition particle (SRP) system plays an important role in membrane protein biogenesis. Previous studies have suggested indirectly that in addition to its role during the targeting of ribosomes translating membrane proteins to translocons, the SRP might also have a quality control role in preventing premature synthesis of membrane proteins in the cytoplasm. This proposal was studied here using cells simultaneously overexpressing various membrane proteins and either SRP, the SRP protein Ffh, its 4.5S RNA, or the Ffh M domain. The results show that SRP, Ffh, and the M domain are all able to selectively inhibit the expression of membrane proteins. We observed no apparent changes in the steady-state mRNA levels or membrane protein stability, suggesting that inhibition may occur at the level of translation, possibly through the interaction between Ffh and ribosome-hydrophobic nascent chain complexes. Since E. coli SRP does not have a eukaryote-like translation arrest domain, we discuss other possible mechanisms by which this SRP might regulate membrane protein translation when overexpressed.

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Related in: MedlinePlus

A model for the interaction of Ffh domains with the E.¬†coli 50S ribosomal subunit (Protein Data Bank [PDB] code 2AWB). The large-subunit rRNA (23S) is shown as a view from the ribosome exit tunnel (‚ėÜ). Two of its helices are shown, h24 and h59. Large-subunit proteins are shown as a sphere presentation. A double line across the figure approximately separates the M and NG domain binding sites on the ribosome. This figure was created by the software program PyMol (DeLano Scientific LLC, San Carlos, CA [http://www.pymol.org/]).
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f9: A model for the interaction of Ffh domains with the E.¬†coli 50S ribosomal subunit (Protein Data Bank [PDB] code 2AWB). The large-subunit rRNA (23S) is shown as a view from the ribosome exit tunnel (‚ėÜ). Two of its helices are shown, h24 and h59. Large-subunit proteins are shown as a sphere presentation. A double line across the figure approximately separates the M and NG domain binding sites on the ribosome. This figure was created by the software program PyMol (DeLano Scientific LLC, San Carlos, CA [http://www.pymol.org/]).

Mentions: In mammalian cells, SRP binds to the elongation-factors’ binding site(s) of the ribosome through the SRP-RNA Alu domain and the accompanied SRP9/14 proteins, as observed by cryoelectron microscopy (38), and thereby inhibits translation. In contrast, the E. coli SRP does not have an Alu domain, and therefore, the mechanism by which E. coli utilizes the SRP to regulate translation of membrane proteins in the cytosol must be different from that proposed for the eukaryotic system. One possibility is that the SRP imposes an inhibitory conformational effect upon binding at the ribosome region flanking the nascent peptide exit site. Cryoelectron microscopy and single-particle analysis (39) revealed that the E. coli SRP interacts with a translation-arrested 70S ribosome via both its NG and its M domains (depicted in Fig. 9). NG interacts through the N domain with the ribosomal proteins L23/L29 in the large subunit, and the M domain interacts with several rRNA helices and possibly with the ribosomal proteins L22 and L24. The pairs L22/L24 and L23/L29 are located on opposite sides of the ribosome exit tunnel (Fig. 9). In the context of our results, this may suggest that the conformational response of L22/L24 upon the binding to the M domain might lead to regulation of translation by slowing down the movement of the nascent chain through the tunnel. Particularly interesting is the postulated interaction with L22, because this protein was shown to be involved in translation regulation under various conditions (40). Elucidation at high resolution of the precise contacts of the M domain with the ribosome large subunit is crucial for understanding the mechanism of the postulated translation regulation by the SRP and its further evaluation.


Escherichia coli SRP, its protein subunit Ffh, and the Ffh M domain are able to selectively limit membrane protein expression when overexpressed.

Yosef I, Bochkareva ES, Bibi E - MBio (2010)

A model for the interaction of Ffh domains with the E.¬†coli 50S ribosomal subunit (Protein Data Bank [PDB] code 2AWB). The large-subunit rRNA (23S) is shown as a view from the ribosome exit tunnel (‚ėÜ). Two of its helices are shown, h24 and h59. Large-subunit proteins are shown as a sphere presentation. A double line across the figure approximately separates the M and NG domain binding sites on the ribosome. This figure was created by the software program PyMol (DeLano Scientific LLC, San Carlos, CA [http://www.pymol.org/]).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2921155&req=5

f9: A model for the interaction of Ffh domains with the E.¬†coli 50S ribosomal subunit (Protein Data Bank [PDB] code 2AWB). The large-subunit rRNA (23S) is shown as a view from the ribosome exit tunnel (‚ėÜ). Two of its helices are shown, h24 and h59. Large-subunit proteins are shown as a sphere presentation. A double line across the figure approximately separates the M and NG domain binding sites on the ribosome. This figure was created by the software program PyMol (DeLano Scientific LLC, San Carlos, CA [http://www.pymol.org/]).
Mentions: In mammalian cells, SRP binds to the elongation-factors’ binding site(s) of the ribosome through the SRP-RNA Alu domain and the accompanied SRP9/14 proteins, as observed by cryoelectron microscopy (38), and thereby inhibits translation. In contrast, the E. coli SRP does not have an Alu domain, and therefore, the mechanism by which E. coli utilizes the SRP to regulate translation of membrane proteins in the cytosol must be different from that proposed for the eukaryotic system. One possibility is that the SRP imposes an inhibitory conformational effect upon binding at the ribosome region flanking the nascent peptide exit site. Cryoelectron microscopy and single-particle analysis (39) revealed that the E. coli SRP interacts with a translation-arrested 70S ribosome via both its NG and its M domains (depicted in Fig. 9). NG interacts through the N domain with the ribosomal proteins L23/L29 in the large subunit, and the M domain interacts with several rRNA helices and possibly with the ribosomal proteins L22 and L24. The pairs L22/L24 and L23/L29 are located on opposite sides of the ribosome exit tunnel (Fig. 9). In the context of our results, this may suggest that the conformational response of L22/L24 upon the binding to the M domain might lead to regulation of translation by slowing down the movement of the nascent chain through the tunnel. Particularly interesting is the postulated interaction with L22, because this protein was shown to be involved in translation regulation under various conditions (40). Elucidation at high resolution of the precise contacts of the M domain with the ribosome large subunit is crucial for understanding the mechanism of the postulated translation regulation by the SRP and its further evaluation.

Bottom Line: The results show that SRP, Ffh, and the M domain are all able to selectively inhibit the expression of membrane proteins.We observed no apparent changes in the steady-state mRNA levels or membrane protein stability, suggesting that inhibition may occur at the level of translation, possibly through the interaction between Ffh and ribosome-hydrophobic nascent chain complexes.Since E. coli SRP does not have a eukaryote-like translation arrest domain, we discuss other possible mechanisms by which this SRP might regulate membrane protein translation when overexpressed.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, Israel.

ABSTRACT
The Escherichia coli signal recognition particle (SRP) system plays an important role in membrane protein biogenesis. Previous studies have suggested indirectly that in addition to its role during the targeting of ribosomes translating membrane proteins to translocons, the SRP might also have a quality control role in preventing premature synthesis of membrane proteins in the cytoplasm. This proposal was studied here using cells simultaneously overexpressing various membrane proteins and either SRP, the SRP protein Ffh, its 4.5S RNA, or the Ffh M domain. The results show that SRP, Ffh, and the M domain are all able to selectively inhibit the expression of membrane proteins. We observed no apparent changes in the steady-state mRNA levels or membrane protein stability, suggesting that inhibition may occur at the level of translation, possibly through the interaction between Ffh and ribosome-hydrophobic nascent chain complexes. Since E. coli SRP does not have a eukaryote-like translation arrest domain, we discuss other possible mechanisms by which this SRP might regulate membrane protein translation when overexpressed.

Show MeSH
Related in: MedlinePlus