TY - JOUR
T1 - SRP, FtsY, DnaK and YidC Are Required for the Biogenesis of the E. coli Tail-Anchored Membrane Proteins DjlC and Flk
AU - Peschke, Markus
AU - Le Goff, Mélanie
AU - Koningstein, Gregory M.
AU - Karyolaimos, Alexandros
AU - de Gier, Jan Willem
AU - van Ulsen, Peter
AU - Luirink, Joen
PY - 2017/1/1
Y1 - 2017/1/1
N2 - Tail-anchored membrane proteins (TAMPs) are relatively simple membrane proteins characterized by a single transmembrane domain (TMD) at their C-terminus. Consequently, the hydrophobic TMD, which acts as a subcellular targeting signal, emerges from the ribosome only after termination of translation precluding canonical co-translational targeting and membrane insertion. In contrast to the well-studied eukaryotic TAMPs, surprisingly little is known about the cellular components that facilitate the biogenesis of bacterial TAMPs. In this study, we identify DjlC and Flk as bona fide Escherichia coli TAMPs and show that their TMDs are necessary and sufficient for authentic membrane targeting of the fluorescent reporter mNeonGreen. Using strains conditional for the expression of known E. coli membrane targeting and insertion factors, we demonstrate that the signal recognition particle (SRP), its receptor FtsY, the chaperone DnaK and insertase YidC are each required for efficient membrane localization of both TAMPs. A close association between the TMD of DjlC and Flk with both the Ffh subunit of SRP and YidC was confirmed by site-directed in vivo photo-crosslinking. In addition, our data suggest that the hydrophobicity of the TMD correlates with the dependency on SRP for efficient targeting.
AB - Tail-anchored membrane proteins (TAMPs) are relatively simple membrane proteins characterized by a single transmembrane domain (TMD) at their C-terminus. Consequently, the hydrophobic TMD, which acts as a subcellular targeting signal, emerges from the ribosome only after termination of translation precluding canonical co-translational targeting and membrane insertion. In contrast to the well-studied eukaryotic TAMPs, surprisingly little is known about the cellular components that facilitate the biogenesis of bacterial TAMPs. In this study, we identify DjlC and Flk as bona fide Escherichia coli TAMPs and show that their TMDs are necessary and sufficient for authentic membrane targeting of the fluorescent reporter mNeonGreen. Using strains conditional for the expression of known E. coli membrane targeting and insertion factors, we demonstrate that the signal recognition particle (SRP), its receptor FtsY, the chaperone DnaK and insertase YidC are each required for efficient membrane localization of both TAMPs. A close association between the TMD of DjlC and Flk with both the Ffh subunit of SRP and YidC was confirmed by site-directed in vivo photo-crosslinking. In addition, our data suggest that the hydrophobicity of the TMD correlates with the dependency on SRP for efficient targeting.
KW - E. coli
KW - Membrane insertion
KW - Membrane protein
KW - Membrane targeting
KW - Tail-anchored
UR - http://www.scopus.com/inward/record.url?scp=85038878102&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85038878102&partnerID=8YFLogxK
U2 - 10.1016/j.jmb.2017.12.004
DO - 10.1016/j.jmb.2017.12.004
M3 - Article
AN - SCOPUS:85038878102
SN - 0022-2836
VL - 430
SP - 389
EP - 403
JO - Journal of Molecular Biology
JF - Journal of Molecular Biology
IS - 3
ER -