Role of Sec61α in the Regulated Transfer of the Ribosome–Nascent Chain Complex from the Signal Recognition Particle to the Translocation Channel

Song, Weiqun; Raden, David; Mandon, Elisabet C.; Gilmore, Reid

doi:10.1016/s0092-8674(00)80669-8

Cited by 131 publications

(129 citation statements)

References 39 publications

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“…Chymotrypsin digestion can be used to selectively cleave Sec61␣ molecules that are not covered by ribosomes (Kalies et al, 1994); cleavage occurs in the loops between transmembrane segments 6 and 7 and 8 and 9 Song et al, 2000). Although only the free Sec61 population was degraded by chymotrypsin (our unpublished data), this led to the dissociation of ϳ60% of the prebound ribosomes ( Figure 5D), suggesting that Sec61 molecules that previously were beneath a ribosome became accessible to the protease.…”

Section: Interconvertible Sec61 Populationsmentioning

confidence: 89%

Ribosome Binding to and Dissociation from Translocation Sites of the Endoplasmic Reticulum Membrane

Schaletzky

Rapoport

2006

MBoC

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We have addressed how ribosome-nascent chain complexes (RNCs), associated with the signal recognition particle (SRP), can be targeted to Sec61 translocation channels of the endoplasmic reticulum (ER) membrane when all binding sites are occupied by nontranslating ribosomes. These competing ribosomes are known to be bound with high affinity to tetramers of the Sec61 complex. We found that the membrane binding of RNC-SRP complexes does not require or cause the dissociation of prebound nontranslating ribosomes, a process that is extremely slow. SRP and its receptor target RNCs to a free population of Sec61 complex, which associates with nontranslating ribosomes only weakly and is conformationally different from the population of ribosome-bound Sec61 complex. Taking into account recent structural data, we propose a model in which SRP and its receptor target RNCs to a Sec61 subpopulation of monomeric or dimeric state. This could explain how RNC-SRP complexes can overcome the competition by nontranslating ribosomes. INTRODUCTIONCotranslational translocation is the major pathway in mammals by which proteins are transported across or integrated into the endoplasmic reticulum (ER) membrane. The synthesis of these proteins starts with a free ribosome in the cytosol. Once a hydrophobic signal sequence or transmembrane (TM) segment has emerged from the ribosome, the signal recognition particle (SRP) binds, forming a ribosomenascent chain (RNC)-SRP complex (for reviews, see Walter and Johnson, 1994;Egea et al., 2005). This complex binds to the ER membrane by interactions between SRP and the SRP receptor (SR) and between the ribosome and the Sec61 complex, a heterotrimeric membrane protein that forms a proteinconducting channel (Van den Berg et al., 2004). The nascent polypeptide chain forms a loop when inserting into the channel, with the signal or TM sequence intercalated into the walls of the channel and the following mature region in the pore proper (for reviews, see Rapoport et al., 2004;Osborne et al., 2005). For secretory proteins, this part of the elongating chain is moved through the channel into the ER lumen, and the signal sequence is eventually cleaved off. In membrane proteins, TM sequences are partitioned sideways through the walls of the channel into the lipid phase, leaving some parts of the polypeptide chain in the cytosol and others in the ER lumen. For both classes of proteins, the translating ribosome stays bound to the channel during the entire translocation process (Mothes et al., 1997).Previous work has shown that the Sec61 complex can bind not only RNCs but also nontranslating ribosomes with nanomolar affinity (Borgese et al., 1974;Kalies et al., 1994;Prinz et al., 2000a). Ribosomes remain bound to ER membranes after nascent chain release (Adelman et al., 1973) and do not easily dissociate from the membrane (Borgese et al., 1973;Potter and Nicchitta, 2002). Given that the concentration of free ribosomes in a secretory cell is ϳ0.3 M, 95% of the Sec61 binding sites should be occupied (Blobel and Potter, ...

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Section: Interconvertible Sec61 Populationsmentioning

confidence: 89%

Ribosome Binding to and Dissociation from Translocation Sites of the Endoplasmic Reticulum Membrane

Schaletzky

Rapoport

2006

MBoC

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“…This may indicate that additional factors are required for high (catalytic) rates of GTP hydrolysis. In bacteria, lipid interactions stimulate the GTPase activity of FtsY (39), and interaction of available translocase components with targeting machinery initiates GTP hydrolysis in targeting to the ER (40). In that context, the low rate of GTP hydrolysis by cpSRP54/cpFtsY in solution is likely of biological significance because high rates of GTP hydrolysis in solution would lead to a potentially deleterious futile cycle of GTP hydrolysis in stroma.…”

Section: Fig 6 Functional Analysis Of the C-terminal 26 Amino Acidsmentioning

confidence: 99%

Regulation of the GTPase Cycle in Post-translational Signal Recognition Particle-based Protein Targeting Involves cpSRP43

Goforth¹,

Peterson²,

Yuan³

et al. 2004

Journal of Biological Chemistry

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“…What interactions allow other components of the SRP targeting machinery to communicate that cargo has been delivered, stimulating GTP hydrolysis and disengaging the targeting complex? 16 As a first step towards addressing the structural basis for progression from assembly of the Ffh:FtsY heterodimeric complex to activation for nucleotide hydrolysis, we crystallized the complex of the NG domains of the SRP GTPases Ffh and FtsY from T. aquaticus in the presence of Mg:GDP:AlF 4 . The GDP:AlF 4 functions as a putative transition-state analog, 17 and has been exploited to determine the structures of other GTPases with their activating proteins, including Ras:RasGAP, 18 Gα:RGS4, 19 and Rho:RhoGAP.…”

Section: Introductionmentioning

confidence: 99%

Structure of a GDP:AlF4 Complex of the SRP GTPases Ffh and FtsY, and Identification of a Peripheral Nucleotide Interaction Site

Focia

Gawronski-Salerno

Coon

et al. 2006

Journal of Molecular Biology

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The signal recognition particle (SRP) GTPases Ffh and FtsY play a central role in co-translational targeting of proteins, assembling in a GTP-dependent manner to generate the SRP targeting complex at the membrane. A suite of residues in FtsY have been identified that are essential for the hydrolysis of GTP that accompanies disengagement. We have argued previously on structural grounds that this region mediates interactions that serve to activate the complex for disengagement and term it the activation region. We report here the structure of a complex of the SRP GTPases formed in the presence of GDP:AlF 4 . This complex accommodates the putative transition-state analog without undergoing significant change from the structure of the ground-state complex formed in the presence of the GTP analog GMPPCP. However, small shifts that do occur within the shared catalytic chamber may be functionally important. Remarkably, an external nucleotide interaction site was identified at the activation region, revealed by an unexpected contaminating GMP molecule bound adjacent to the catalytic chamber. This site exhibits conserved sequence and structural features that suggest a direct interaction with RNA plays a role in regulating the activity of the SRP targeting complex.

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Role of Sec61α in the Regulated Transfer of the Ribosome–Nascent Chain Complex from the Signal Recognition Particle to the Translocation Channel

Cited by 131 publications

References 39 publications

Ribosome Binding to and Dissociation from Translocation Sites of the Endoplasmic Reticulum Membrane

Ribosome Binding to and Dissociation from Translocation Sites of the Endoplasmic Reticulum Membrane

Regulation of the GTPase Cycle in Post-translational Signal Recognition Particle-based Protein Targeting Involves cpSRP43

Structure of a GDP:AlF4 Complex of the SRP GTPases Ffh and FtsY, and Identification of a Peripheral Nucleotide Interaction Site

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