Quantifying Absolute Protein Synthesis Rates Reveals Principles Underlying Allocation of Cellular Resources

Li, Gene-Wei; Burkhardt, David H; Gross, Carol A.; Weissman, Jonathan S.

doi:10.1016/j.cell.2014.02.033

Cited by 1,155 publications

(1,430 citation statements)

References 81 publications

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“…These results are similar to the one obtained with ribosome profiling [30] that measure 360 to 560 channels per cell. These numbers take the channel density close to or above the threshold for cluster formation at low tensions.…”

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confidence: 78%

Cooperative response and clustering: Consequences of membrane-mediated interactions among mechanosensitive channels

2017

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Mechanosensitive (MS) channels are ion channels which act as cells' safety valves, opening when the osmotic pressure becomes too high and making cells avoid damage by releasing ions. They are found on the cellular membrane of a large number of organisms. They interact with each other by means of deformations they induce in the membrane. We show that collective dynamics arising from the inter-channel interactions lead to first and second-order phase transitions in the fraction of open channels in equilibrium relating to the formation of channel clusters. We show that this results in a considerable delay of the response of cells to osmotic shocks, and to an extreme cell-to-cell stochastic variations in their response times, despite the large numbers of channels present in each cell. We discuss how our results are relevant for E. coli.Abrupt changes in the osmolarity of the environment is a hazard most organisms are subject to at one time or another [1][2][3][4][5][6]. A sudden drop in osmolarity (an osmotic shock ) will cause water to rush into a living cell, and requires an immediate response by the cell to prevent it from getting damaged or undergoing lysis from the increased tension on the cellular membrane. Mechanosensitive channels (or MS channels) are ion channels located on the cell membrane, which open when the membrane tension becomes too high [7,8], and play a crucial role in the cell's defence mechanism against osmotic shocks [9,10]. They act as safety valves, releasing ions and decreasing the osmotic pressure and the membrane tension. Mechanosensitive channels are found in many organisms, and have been well characterised in the bacterium E. coli [11-13].The cellular membrane in which the mechanosensitive channels are inserted is a lipid bilayer. The interior of the bilayer is hydrophobic, making it energetically favourable for it to thicken or compress to match the hydrophobic parts of the channel proteins inserted in the membrane [14]. This results in a deformation profile around each channel, with the thickness of the bilayer being a function of position. This deformation mediates a shortrange effective force between two neighbouring channels, similar to the force between two nearby corks floating on water, which interact through the deformation they induce on the surface of water. This interaction can be attractive or repulsive, depending on the shapes of the two molecules. Furthermore, a theoretical analysis suggests that the interaction between two neighbouring channels lowers the tension needed to open them during an osmotic shock [15], raising the possibility that their function could be influenced by their spatial distribution on the membrane (as already noticed for other membrane proteins [16,17]). This is reinforced by the fact that the channels' attractive forces suggest that they may agglomerate into clusters. Our goal in this paper is to determine the consequences that the inter-channel interaction has on the dynamics of this system, focusing in particular on channel clustering and its con...

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confidence: 78%

Cooperative response and clustering: Consequences of membrane-mediated interactions among mechanosensitive channels

2017

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“…Do these complexes also have uneven stoichiometry within the cell? Recent studies have demonstrated increased translational efficiency for the higher stoichiometry subunits within a complex 33,34 , suggesting that in vivo protein expression levels are often optimized for the same uneven stoichiometry observed in vitro. In another study, a high proportion of the pairwise interactions from complexes purified from human cells was estimated to have uneven stoichiometry, although such proteomic measurements are only approximate 3 .…”

Section: Resultsmentioning

confidence: 99%

Structural and evolutionary versatility in protein complexes with uneven stoichiometry

et al. 2015

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Proteins assemble into complexes with diverse quaternary structures. Although most heteromeric complexes of known structure have even stoichiometry, a significant minority have uneven stoichiometry-that is, differing numbers of each subunit type. To adopt this uneven stoichiometry, sequence-identical subunits must be asymmetric with respect to each other, forming different interactions within the complex. Here we first investigate the occurrence of uneven stoichiometry, demonstrating that it is common in vitro and is likely to be common in vivo. Next, we elucidate the structural determinants of uneven stoichiometry, identifying six different mechanisms by which it can be achieved. Finally, we study the frequency of uneven stoichiometry across evolution, observing a significant enrichment in bacteria compared with eukaryotes. We show that this arises due to a general increased tendency for bacterial proteins to self-assemble and form homomeric interactions, even within the context of a heteromeric complex.

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“…Cellular levels of FtsZ are predicted to be~5000 molecules per cell and changes in FtsZ intracellular concentrations falling below or increasing above the threshold amounts lead to gross division defects or cell death (Wang et al, 1991;Ward & Lutkenhaus, 1985). ZapC is predicted to be a low-abundance protein with~200 copies per cell and overexpression of ZapC is thought to hyperstabilize aberrant FtsZ assemblies leading to lethal filamentation (Li et al, 2014). Therefore, the relatively slow ClpP-mediated turnover of FtsZ and ZapC per generation suggests that maintaining a fine balance of the FtsZ monomer/polymer ratio and the amounts of the stabilizer ZapC in the cell is critical to Z-ring assembly.…”

Section: Discussionmentioning

confidence: 99%

“…Prior work has shown that overexpression of ZapC leads to hyperstable Zring assemblies and filamentation without affecting steadystate FtsZ levels (Durand-Heredia et al, 2011;Hale et al, 2011). Little is known about the regulation of ZapC itself, but the intracellular levels are considered to be low and it is predicted to be a substrate of ClpXP in E. coli (Flynn et al, 2003;Li et al, 2014). Consistent with this latter observation, the ZapC sequence encodes both an N-terminal ClpX recognition sequence ( 1 MRIK-X 6 -W 11 ) and a C-terminal ssrAlike sequence ( 178 QAV 180 ) (Flynn et al, 2003).…”

Section: Introductionmentioning

confidence: 99%

ClpXP and ClpAP control the Escherichia coli division protein ZapC by proteolysis

2016

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The bacterial FtsZ-ring is an essential cytokinetic structure under tight spatiotemporal regulation. In Escherichia coli, FtsZ polymerization and assembly into the Z-ring is controlled on multiple levels through interactions with positive and negative regulators. Among these regulatory factors are ZapC, a Z-ring stabilizer, and the conserved protease ClpXP, which has been shown to degrade FtsZ protofilaments in preference to FtsZ monomers. Here we report that ZapC and ClpX interact in a protein-protein interaction assay, and that ZapC is degraded in a ClpXP-dependent manner in vivo. The SspB adaptor protein is not required for targeting ZapC to the ClpXP proteolytic machinery. A mutation disrupting the zapC ssrAlike sequence (zapC DD ) stabilizes ZapC consistent with a reduction in ClpXP-mediated ZapC degradation. ZapC DD retains the ability to interact with FtsZ and to promote bundling in vitro indicating that WT ZapC contains discrete FtsZ and ClpX recognition motifs. Additionally, ClpAP complexes are sufficient for degradation of ZapC in the absence of ClpX in vivo. Further, chromosomal expression of zapC DD suppresses filamentation of the temperature-sensitive ftsZ84 mutant, confirming the role of ZapC as a Z-ring stabilizer. Lastly, changes in ClpXP and ZapC levels lead to cell division effects, likely through their roles in modulating FtsZ assembly dynamics. Taken together, our results indicate that the Zring stabilizer ZapC is a substrate of both ClpXP and ClpAP in vivo. Our data also point to a more complex regulatory circuit that integrates FtsZ, ClpXP and ZapC in achieving Z-ring stability in E. coli and related species.

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Quantifying Absolute Protein Synthesis Rates Reveals Principles Underlying Allocation of Cellular Resources

Cited by 1,155 publications

References 81 publications

Cooperative response and clustering: Consequences of membrane-mediated interactions among mechanosensitive channels

Cooperative response and clustering: Consequences of membrane-mediated interactions among mechanosensitive channels

Structural and evolutionary versatility in protein complexes with uneven stoichiometry

ClpXP and ClpAP control the Escherichia coli division protein ZapC by proteolysis

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