A mammalian homolog of SEC61p and SECYp is associated with ribosomes and nascent polypeptides during translocation

Görlich, Dirk; Prehn, Siegfried; Hartmann, Enno; Kalies, Kai‐Uwe; Rapoport, Tom A.

doi:10.1016/0092-8674(92)90517-g

Cited by 437 publications

(381 citation statements)

References 52 publications

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“…Such a domain might play an important role in the gating of the translocation pore, either during the initiation of translocation or perhaps to permit the lateral diffusion of hydrophobic domains into the lipid bilayer during the assembly of integral membrane proteins. The potential importance of this putative amphipathic structure is underlined by its conservation both in Sec61␣ (19) and in a Schizosaccharomyces pombe homologue. 4 Our data indicate that both HS2 and HS5 require C-terminal sequences for their topogenesis.…”

Section: Discussionmentioning

confidence: 99%

“…Cross-linking studies indicate that Sec61p is in close proximity to prepro-␣-factor at different stages of its translocation through the bilayer (17,18), suggesting that Sec61p may play a direct role in the formation of a protein-conducting channel in the ER membrane. A mammalian homologue of Sec61p, Sec61␣ (19), also contacts nascent polypeptides during their membrane transfer (19 -21) and exists in a homologous Sec61 complex together with Sec61␤ (Sbh1p) and Sec61␥ (Sss1p) (22).…”

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confidence: 99%

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Determination of the Transmembrane Topology of Yeast Sec61p, an Essential Component of the Endoplasmic Reticulum Translocation Complex

Wilkinson

Critchley

Stirling

1996

Journal of Biological Chemistry

142

147

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Sec61p is a highly conserved integral membrane protein that plays a role in the formation of a proteinconducting channel required for the translocation of polypeptides into, and across, the membrane of the endoplasmic reticulum. As a major step toward elucidating the structure of the endoplasmic reticulum translocation apparatus, we have determined the transmembrane topology of Sec61p using a combination of C-terminal reporter-domain fusions and the in situ digestion of specifically inserted factor Xa protease cleavage sites. Our data indicate the presence of 10 transmembrane domains, including several with surprisingly limited hydrophobicity. Furthermore, we provide evidence for complex intramolecular interactions in which these weakly hydrophobic domains require C-terminal sequences for their correct topogenesis. The incorporation of sequences with limited hydrophobicity into the bilayer may play a vital role in the formation of an aqueous membrane channel required for the translocation of hydrophilic polypeptide chains.

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Determination of the Transmembrane Topology of Yeast Sec61p, an Essential Component of the Endoplasmic Reticulum Translocation Complex

Wilkinson

Critchley

Stirling

1996

Journal of Biological Chemistry

142

147

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“…In eukaryotes, mRNA encoding secreted and membrane proteins can be targeted to the ER co-translationally by the signal recognition particle 1,2 and can be maintained on the ER via the direct interactions between ribosomes and translocons during the translation 3,4 . However, whether mRNAs can be targeted and maintained on the ER independent of either ribosomes or translation was unclear until very recently.…”

Section: Introductionmentioning

confidence: 99%

Visualization of Endoplasmic Reticulum Localized mRNAs in Mammalian Cells

Cui

Palazzo

2012

JoVE

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In eukaryotes, most of the messenger RNAs (mRNAs) that encode secreted and membrane proteins are localized to the surface of the endoplasmic reticulum (ER). However, the visualization of these mRNAs can be challenging. This is especially true when only a fraction of the mRNA is ER-associated and their distribution to this organelle is obstructed by non-targeted (i.e. "free") transcripts. In order to monitor ER-associated mRNAs, we have developed a method in which cells are treated with a short exposure to a digitonin extraction solution that selectively permeabilizes the plasma membrane, and thus removes the cytoplasmic contents, while simultaneously maintaining the integrity of the ER. When this method is coupled with fluorescent in situ hybridization (FISH), one can clearly visualize ER-bound mRNAs by fluorescent microscopy. Using this protocol the degree of ER-association for either bulk poly(A) transcripts or specific mRNAs can be assessed and even quantified. In the process, one can use this assay to investigate the nature of mRNA-ER interactions. Video LinkThe video component of this article can be found at

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“…By virtue of an interaction with its ER-localized receptor, signal recognition particle brings the ribosome-nascent chain complex to the ER membrane (2). Once at the ER, the ribosome is thought to facilitate the assembly or stabilization of the translocation channel, composed of the heterotrimeric Sec61 complex (3,4). The 35-kDa polytopic Sec61␣ subunit appears to form the channel walls, whereas the smaller bitopic ␤-and ␥-subunits play an as yet undetermined role, perhaps in facilitating the insertion of the nascent chain into the channel (5)(6)(7).…”

mentioning

confidence: 99%

Signal Sequences Initiate the Pathway of Maturation in the Endoplasmic Reticulum Lumen

Rutkowski

Ott

Polansky

et al. 2003

Journal of Biological Chemistry

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An interaction between an N-terminal signal sequence and the translocon leads to the initiation of protein translocation into the endoplasmic reticulum lumen. Subsequently, folding and modification of the substrate rapidly ensue. The close temporal coordination of these processes suggests that they may be structurally and functionally coordinated as well. Here we show that information encoded in the hydrophobic domain of a signal sequence influences the timing and efficiency of at least two steps in maturation, namely N-linked glycosylation and signal sequence cleavage. We demonstrate that these consequences correlate with and likely stem from the nature of the initial association made between the signal sequence and the translocon during the initiation of translocation. We propose a model by which these maturational events are controlled by the signal sequence-translocon interaction. Our work demonstrates that the pathway taken by a nascent chain through post-translational maturation depends on information encoded in its signal sequence.N-terminal signal sequences enable nascent secretory and transmembrane proteins to be targeted to the endoplasmic reticulum (ER) 1 for translocation. The signal sequence, newly emerged from the translating ribosome, is recognized in the cytoplasm by the signal recognition particle (1). By virtue of an interaction with its ER-localized receptor, signal recognition particle brings the ribosome-nascent chain complex to the ER membrane (2). Once at the ER, the ribosome is thought to facilitate the assembly or stabilization of the translocation channel, composed of the heterotrimeric Sec61 complex (3, 4). The 35-kDa polytopic Sec61␣ subunit appears to form the channel walls, whereas the smaller bitopic ␤-and ␥-subunits play an as yet undetermined role, perhaps in facilitating the insertion of the nascent chain into the channel (5-7).An important advance in the understanding of the initiation of translocation was the discovery of a second step of signal sequence recognition. In addition to being recognized by the signal recognition particle, the signal sequence also interacts with the translocation channel itself (8,9). Although the mechanism is not yet clear, this event is thought to stimulate the initiation of translocation. For the simplest secretory proteins, when the signal sequence is recognized by the channel, the ribosome assumes a tight interaction with the channel that shields the protein from the cytoplasm (8, 10). Concomitantly, the channel opens toward the ER lumen, either via a conformational change in the channel or removal of a molecular plug covering the luminal aperture (11,12).Not all signal sequences initiate translocation in the same way. Beyond merely stimulating the initiation of translocation, signal sequences can regulate the association between the ribosome and translocon and the exposure of the nascent chain to the cytoplasm or ER lumen (13-16). In at least one case, that of the prion protein, the consequence of this regulatory step is the governance of the ...

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A mammalian homolog of SEC61p and SECYp is associated with ribosomes and nascent polypeptides during translocation

Cited by 437 publications

References 52 publications

Determination of the Transmembrane Topology of Yeast Sec61p, an Essential Component of the Endoplasmic Reticulum Translocation Complex

Determination of the Transmembrane Topology of Yeast Sec61p, an Essential Component of the Endoplasmic Reticulum Translocation Complex

Visualization of Endoplasmic Reticulum Localized mRNAs in Mammalian Cells

Signal Sequences Initiate the Pathway of Maturation in the Endoplasmic Reticulum Lumen

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