Dataset Information

The SecYEG interactome of E. coli: non-competitive binding of PpiD and YidC to the SecYEG translocon

ABSTRACT: The SecYEG translocon constitutes the major protein transporting channel in bacteria and transports an enormous variety of different secretory and inner membrane proteins. The minimal core of the SecYEG translocon consists of the three inner membrane proteins SecY, SecE and SecG, which together with appropriate targeting factors are sufficient for protein transport in vitro. However, in vivo the SecYEG translocon has been shown to associate with multiple partner proteins, which likely allow the SecYEG translocon to manage diverse substrates. For obtaining a global view on SecYEG plasticity, a label free proteomics approach was executed, which identified known SecYEG interacting proteins, verified the interaction with quality control proteins and identified several novel putative SecYEG interactors. Surprisingly, the chaperone complex PpiD/YfgM was identified as the most pronounced contact partner of SecYEG and was much dominant than the established partner protein YidC. Detailed analyses of the PpiD and YidC contact sites on SecY by site directed cross-linking revealed that PpiD and YidC use almost completely overlapping binding sites on SecY and contact the lateral gate, the plug domain and the periplasmic cavity of SecY. Still, despite having almost identical binding sites, their binding to SecY is non-competitive as determined by quantitative mass spectrometry and cross-linking. This implies that the SecYEG translocon forms different substrate-independent sub-assemblies, in which SecYEG is either in contact with YidC or with the PpiD/YfgM complex.

INSTRUMENT(S): LTQ FT

ORGANISM(S): Escherichia Coli

SUBMITTER: Friedel Drepper

LAB HEAD: Bettina Warscheid

PROVIDER: PXD015974 | Pride | 2019-11-07

REPOSITORIES: Pride

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Publications

Noncompetitive binding of PpiD and YidC to the SecYEG translocon expands the global view on the SecYEG interactome in Escherichia coli.

Jauss Benjamin B Petriman Narcis-Adrian NA Drepper Friedel F Franz Lisa L Sachelaru Ilie I Welte Thomas T Steinberg Ruth R Warscheid Bettina B Koch Hans-Georg HG

The Journal of biological chemistry 20191107 50

The SecYEG translocon constitutes the major protein transport channel in bacteria and transfers an enormous variety of different secretory and inner-membrane proteins. The minimal core of the SecYEG translocon consists of three inner-membrane proteins, SecY, SecE, and SecG, which, together with appropriate targeting factors, are sufficient for protein transport in vitro However, in vivo the SecYEG translocon has been shown to associate with multiple partner proteins, likely allowin ...[more]

PMID: 31699901

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Project description:Disruptions of protein homeostasis in the endoplasmic reticulum (ER) elicit activation of the unfolded protein response (UPR), a translation- and transcription-coupled proteostatic stress response pathway. The primary translational control arm of the UPR is initiated by the PERK-dependent phosphorylation of eIF2Î±, leading to a large-scale reprogramming of translation and subsequent activation of an ATF4-mediated transcriptional program. It has remained challenging, however, to accurately evaluate the contribution of each component of the eIF2Î±/ATF4 pathway to the remodelling of transcription and translation. Here, we have used mouse embryonic fibroblasts containing either a knock-in of the non-phosphorylatable eIF2Î± S51A mutant or knock-out for ATF4 by ribosome profiling and mRNA-seq to define the specific contributions of eIF2Î± phosphoryation and ATF4 in controlling the translational and transcriptional components of the UPR. These studies show that the transcriptional and translational targets of each P-eIF2Î±, ATF4, and the other UPR gene expression programs overlapped extensively, leading to a core set of UPR genes whose robust expression in response to ER stress is driven by multiple mechanisms. The identification of other, more factor-specific targets illustrated the potential for functional specialization of the UPR. As the UPR progressed temporally, cells employed distinct combinations of transcriptional and translational mechanisms, initiated by different factors, to adapt to ongoing stress. These effects were accompanied by a buffering effect where changes in mRNA levels were coupled to opposing changes in ribosome loading, a property which makes cooperation between transcription and translation essential to confer robust protein expression. Translational analysis by ribosome profiling and mRNA-seq of PERK pathways mutants during the UPR. Mouse embryonic fibroblasts (MEFs) lacking components of the PERK pathway (eIF2a phosphorylation and ATF4) were subjected to ER stress and analyzed by ribosome profiling.

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Dataset Information

The SecYEG interactome of E. coli: non-competitive binding of PpiD and YidC to the SecYEG translocon

Dataset's files

Publications

Noncompetitive binding of PpiD and YidC to the SecYEG translocon expands the global view on the SecYEG interactome in <i>Escherichia coli</i>.

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