Double-spanning Plant Viral Movement Protein Integration into theEndoplasmic Reticulum Membrane Is Signal Recognition Particle-dependent,Translocon-mediated, andConcerted

Saurı́, Ana; Saksena, Suraj; Salgado, Jesús; Johnson, Arthur E.; Mingarro, Ismael

doi:10.1074/jbc.m412476200

Cited by 44 publications

(63 citation statements)

References 35 publications

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“…More recently, we have demonstrated that the protein integrates into the ER membrane cotranslationally through the Sec61 translocon 13 . But in contrast to what has been observed for other double-spanning proteins both in mammalian and prokaryotic systems 15; 16 , p9 remained adjacent to Sec61α until translation was terminated.…”

Section: Plant Viral Membrane Protein P9 Is Sequentially Adjacent To mentioning

confidence: 99%

“…A quantitative comparison of the relative amounts of photoadducts containing Sec61α and TRAM was made by splitting each sample and then immunoprecipitating in parallel one half with an affinity-purified antiserum specific for Sec61α 13 and the other half with an affinity-purified antiserum for TRAM. When the probe was positioned in TM1, fewer crosslinks were formed to TRAM than to Sec61α, but the number of TRAM photoadducts increased with increasing RNC lengths (Fig.…”

Section: Accepted Manuscriptmentioning

confidence: 99%

“…To accomplish this, the extramembraneous P2 domain of the Escherichia coli leader peptidase (Lep) was fused to the C-terminal end of p9 (see reference 13 for cloning details), and the resulting plasmid was used to generate DNA fragments of different lengths by PCR using 3' primers that anneal at selected positions (Fig. 1).…”

Section: Accepted Manuscriptmentioning

confidence: 99%

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Sec61α and TRAM are Sequentially Adjacent to a Nascent Viral Membrane Protein during its ER Integration

Saurı́

McCormick

Johnson

et al. 2007

Journal of Molecular Biology

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Section: Plant Viral Membrane Protein P9 Is Sequentially Adjacent To mentioning

confidence: 99%

Section: Accepted Manuscriptmentioning

confidence: 99%

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Sec61α and TRAM are Sequentially Adjacent to a Nascent Viral Membrane Protein during its ER Integration

Saurı́

McCormick

Johnson

et al. 2007

Journal of Molecular Biology

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“…In viruses with genomes that include three partially overlapping open reading frames, termed the triple-gene block (TGB), all three TGB proteins are required for movement where the two smaller proteins, TGBp2 and TGBp3, are also TM proteins (24). Furthermore, cross-linking experiments with carnation mottle virus p9 protein demonstrated that its membrane insertion occurs cotranslationally in a signal recognition particle-dependent manner and throughout the cellular membrane integration components, the translocon (33,34).…”

mentioning

confidence: 99%

Plant Virus Cell-to-Cell Movement Is Not Dependent on the Transmembrane Disposition of Its Movement Protein

Martínez-Gil

Sánchez‐Navarro

Cruz

et al. 2009

J Virol

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The cell-to-cell transport of plant viruses depends on one or more virus-encoded movement proteins (MPs). Some MPs are integral membrane proteins that interact with the membrane of the endoplasmic reticulum, but a detailed understanding of the interaction between MPs and biological membranes has been lacking. The cell-to-cell movement of the Prunus necrotic ringspot virus (PNRSV) is facilitated by a single MP of the 30K superfamily. Here, using a myriad of biochemical and biophysical approaches, we show that the PNRSV MP contains only one hydrophobic region (HR) that interacts with the membrane interface, as opposed to being a transmembrane protein. We also show that a proline residue located in the middle of the HR constrains the structural conformation of this region at the membrane interface, and its replacement precludes virus movement.Plant viruses encode movement proteins (MPs) that mediate the intra-and intercellular spread of the viral genome via plasmodesmata, membranous channels that traverse the walls of plant cells and enable intercellular transport and communication. There is a range of diversity in the number and type of viral proteins required for viral movement (21). Research on tobacco mosaic virus (TMV) has played a leading role in understanding MP activity (2). The genome of TMV encodes a single 30-kDa multidomain protein, the namesake of the 30K superfamily (7). Viral RNA is associated with the membrane of the endoplasmic reticulum (ER) and microtubules in the presence of this MP (23,30).A large number of plant viruses have 30K MPs, which share common abilities, including binding nucleic acids, localizing and increasing the size exclusion limit of plasmodesmata, and interacting with the ER membrane. A topological model has been proposed in which the TMV MP has two putative transmembrane (TM) helices, both the N and C termini oriented toward the cytoplasm, and a short loop exposed in the ER lumen (4). There is less experimental information for other 30K MPs, but they are likely to have some membrane interaction.Direct experimental evidence of the integration of MPs into the membrane has been obtained only for small hydrophobic MPs that do not belong to the 30K superfamily. There are two TM segments in the p9 protein of carnation mottle virus (41), whereas the p6 protein of beet yellow virus (29) and the p7B protein of melon necrotic spot virus (22) have a single TM segment. In viruses with genomes that include three partially overlapping open reading frames, termed the triple-gene block (TGB), all three TGB proteins are required for movement where the two smaller proteins, TGBp2 and TGBp3, are also TM proteins (24). Furthermore, cross-linking experiments with carnation mottle virus p9 protein demonstrated that its membrane insertion occurs cotranslationally in a signal recognition particle-dependent manner and throughout the cellular membrane integration components, the translocon (33, 34).Prunus necrotic ringspot virus (PNRSV) is a tripartite, positive-strand RNA virus in the genus Ilarvirus ...

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“…In this regard, two TM segments have been found to accumulate within or adjacent to the translocon before final integration into the lipid bilayer. [39][40][41][42][43] Furthermore, molecular dynamics simulations of the translocon in a fully solvated lipid bilayer showed that two helices can coexist within the translocon (J. Gumbart, personal communication). An interesting possibility arising from these observations is that the translocon may facilitate membrane protein integration by allowing efficient polar interactions between consecutive TM segments harboring charged residues and thereby stabilizing them in the nascent polypeptide.…”

Section: Discussionmentioning

confidence: 99%

Charge Pair Interactions in Transmembrane Helices and Turn Propensity of the Connecting Sequence Promote Helical Hairpin Insertion

Bañó‐Polo

Martínez-Gil

Wallner

et al. 2013

Journal of Molecular Biology

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α-Helical hairpins, consisting of a pair of closely spaced transmembrane (TM) helices that are connected by a short interfacial turn, are the simplest structural motifs found in multi-spanning membrane proteins. In naturally occurring hairpins, the presence of polar residues is common and predicted to complicate membrane insertion. We postulate that the pre-packing process offsets any energetic cost of allocating polar and charged residues within the hydrophobic environment of biological membranes. Consistent with this idea, we provide here experimental evidence demonstrating that helical hairpin insertion into biological membranes can be driven by electrostatic interactions between closely separated, poorly hydrophobic sequences. Additionally, we observe that the integral hairpin can be stabilized by a short loop heavily populated by turn-promoting residues. We conclude that the combined effect of TM-TM electrostatic interactions and tight turns plays an important role in generating the functional architecture of membrane proteins and propose that helical hairpin motifs can be acquired within the context of the Sec61 translocon at the early stages of membrane protein biosynthesis. Taken together, these data further underline the potential complexities involved in accurately predicting TM domains from primary structures.

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Double-spanning Plant Viral Movement Protein Integration into theEndoplasmic Reticulum Membrane Is Signal Recognition Particle-dependent,Translocon-mediated, andConcerted

Cited by 44 publications

References 35 publications

Sec61α and TRAM are Sequentially Adjacent to a Nascent Viral Membrane Protein during its ER Integration

Sec61α and TRAM are Sequentially Adjacent to a Nascent Viral Membrane Protein during its ER Integration

Plant Virus Cell-to-Cell Movement Is Not Dependent on the Transmembrane Disposition of Its Movement Protein

Charge Pair Interactions in Transmembrane Helices and Turn Propensity of the Connecting Sequence Promote Helical Hairpin Insertion

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