Targeting and translocation of the two lipoproteins in Escherichia coli via the SRP/Sec/YidC pathway.

In Escherichia coli, two main protein targeting pathways to the inner membrane exist: the SecB pathway for the essentially posttranslational targeting of secretory proteins and the SRP pathway for cotranslational targeting of inner membrane proteins (IMPs). At the inner membrane both pathways converge at the Sec translocase, which is capable of both linear transport into the periplasm and lateral transport into the lipid bilayer. The Sec-associated YidC appears to assist the lateral transport of IMPs from the Sec translocase into the lipid bilayer. It should be noted that targeting and translocation of only a handful of secretory proteins and IMPs have been studied. These model proteins do not include lipoproteins. Here, we have studied the targeting and translocation of two secretory lipoproteins, the murein lipoprotein and the bacteriocin release protein, using a combined in vivo and in vitro approach. The data indicate that both murein lipoprotein and bacteriocin release protein require the SRP pathway for efficient targeting to the Sec translocase. Furthermore, we show that YidC plays an important role in the targeting/translocation of both lipoproteins

Is GPR146 really the receptor for proinsulin C-peptide?

Proinsulin C-peptide has previously been proposed to interact with a G-protein coupled receptor (GPCR), specifically the orphan receptor GPR146. To investigate the potential of C-peptide in treating complications of diabetes, such as kidney damage, it is necessary to understand its mode of action. We used CHO-K1 cells expressing human GPR146 to study human and murine C-peptide in dynamic mass redistribution and GPCR beta-arrestin assays, as well as with fluorescence confocal microscopy. Neither assay revealed any significant intracellular response to C-peptide at concentrations of up to 33 mu M. We observed no internalisation of C-peptide by fluorescence microscopy. Our results do not support GPR146 as the receptor for C-peptide, but suggest that further investigations of the mode of action of C-peptide should be undertaken

Detection of cross-links between FtsH, YidC, HflK/C suggests a linked role for these proteins in quality control upon insertion of bacterial inner membrane proteins

Little is known about the quality control of proteins upon integration in the inner membrane of Escherichia coli. Here, we demonstrate that YidC and FtsH are adjacent to a nascent, truncated membrane protein using in vitro photo cross-linking. YidC plays a critical but poorly understood role in the biogenesis of E. coli inner membrane proteins (IMPs). FtsH functions as a membrane chaperone and protease. Furthermore, we show that FtsH and its modulator proteins HflK and HflC copurify with tagged YidC and, vice versa, that YidC copurifies with tagged FtsH. These results suggest that FtsH and YidC have a linked role in the quality control of IMPs. STRUCTURED SUMMARY

Sec-dependent membrane protein insertion: sequential interaction of nascent FtsQ with SecY and YidC

Recent studies identified YidC as a novel membrane factor that may play a key role in membrane insertion of inner membrane proteins (IMPs), both in conjunction with the Sec-translocase and as a separate entity. Here, we show that the type II IMP FtsQ requires both the translocase and, to a lesser extent, YidC in vivo. Using photo-crosslinking we demonstrate that the transmembrane (TM) domain of the nascent IMP FtsQ inserts into the membrane close to SecY and lipids, and moves to a combined YidC/lipid environment upon elongation. These data are consistent with a crucial role for YidC in the lateral transfer of TM domains from the Sec translocase into the lipid bilayer

Versatility of inner membrane protein biogenesis in Escherichia coli.

To further our understanding of inner membrane protein (IMP) biogenesis in Escherichia coli, we have accomplished the widest in vivo IMP assembly screen so far. The biogenesis of a set of model IMPs covering most IMP structures possible has been studied in a variety of signal recognition particle (SRP), Sec and YidC mutant strains. We show that the assembly of the complete set of model IMPs is assisted (i.e. requires the aid of proteinaceous factors), and that the requirements for assembly of the model IMPs into the inner membrane differ significantly from each other. This indicates that IMP assembly is much more versatile than previously thought