Student or educator? Sign up with your school email and get 6 months of Pro free.
CoChat
Sign Up
Cited by 165

The DOI matches its Crossref record; 8 of the 8 references it cites were found in a registry or an index.

Narrow equilibrium window for complex coacervation of tau and RNA under cellular conditions

Lin, Yanxian, McCarty, James, Rauch, Jennifer N, Delaney, Kris T, Kosik, Kenneth S, Fredrickson, Glenn H, Shea, Joan-Emma, Han, Songi·2019·eLife
DOI 10.7554/elife.42571PMC PMC6450672

Published version of a preprint

Open this paper in CoChat — free account

Abstract

The mechanism that leads to liquid-liquid phase separation (LLPS) of the tau protein, whose pathological aggregation is implicated in neurodegenerative disorders, is not well understood. Establishing a phase diagram that delineates the boundaries of phase co-existence is key to understanding whether LLPS is an equilibrium or intermediate state. We demonstrate that tau and RNA reversibly form complex coacervates. While the equilibrium phase diagram can be fit to an analytical theory, a more advanced model is investigated through field theoretic simulations (FTS) that provided direct insight into the thermodynamic driving forces of tau LLPS. Together, experiment and simulation reveal that tau-RNA LLPS is stable within a narrow equilibrium window near physiological conditions over experimentally tunable parameters including temperature, salt and tau concentrations, and is entropy-driven. Guided by our phase diagram, we show that tau can be driven toward LLPS under live cell coculturing conditions with rationally chosen experimental parameters.

Overview

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Integrity signals

Registry

Crossref · 10.7554/elife.42571

Cites

8 of 8 references verified0 citation errors · 0 not checkable

Statistics

Figures

Figure 1. Steady tau dynamics and reversible droplet formation of tau-RNA complex coacervates. (A) Diagram of tau187 and polyU RNA. Tau187 is a truncated version of full-length human tau (2N4R 255-441) containing repeat domains and C terminal. At neutral pH experimental conditions, tau187 is overall positively charged; while RNA consists of a phosphate backbone and is negatively charged. (B) Scheme of tau-RNA CC preparation. Mixing clear tau187 and RNA solutions at proper conditions results in a turbid solution containing liquid droplets. (C) X-band cw-EPR spectra (solid line) of tau187 solution (tau187, red), tau187-RNA CC (blue) and tau187-heparin (green) at room temperature with different incubation time, tinc. Samples contains 500 μM tau with 20% spin-labelled. EPR simulation were performed (see Materials and methods) and the fitted spectra is shown as a dashed line. (D) Rotational correlation time, τR extracted from EPR simulation shown in (C) (see Materials and methods). (E) Turbidity of tau187-RNA suspension in consecutive heating-cooling cycles. Confocal images represented samples at 19 °C and 25 °C. Temperatures were ramped at 1 °C/min.
Figure 1. Steady tau dynamics and reversible droplet formation of tau-RNA complex coacervates. (A) Diagram of tau187 and polyU RNA. Tau187 is a truncated version of full-length human tau (2N4R 255-441) containing repeat domains and C terminal. At neutral pH experimental conditions, tau187 is overall positively charged; while RNA consists of a phosphate backbone and is negatively charged. (B) Scheme of tau-RNA CC preparation. Mixing clear tau187 and RNA solutions at proper conditions results in a turbid solution containing liquid droplets. (C) X-band cw-EPR spectra (solid line) of tau187 solution (tau187, red), tau187-RNA CC (blue) and tau187-heparin (green) at room temperature with different incubation time, tinc. Samples contains 500 μM tau with 20% spin-labelled. EPR simulation were performed (see Materials and methods) and the fitted spectra is shown as a dashed line. (D) Rotational correlation time, τR extracted from EPR simulation shown in (C) (see Materials and methods). (E) Turbidity of tau187-RNA suspension in consecutive heating-cooling cycles. Confocal images represented samples at 19 °C and 25 °C. Temperatures were ramped at 1 °C/min.CC BY · PubMed Central
Figure 1—figure supplement 1. ThT fluorescence of tau-RNA CC. 100 μM of tau187 and tau187-SL was mixed with 300 μg/mL polyU RNA (RNA) at room temperature. 100 μM tau187 with 25 μM heparin was prepared as reference. Ribbon shows the standard deviation of three replicates.
Figure 1—figure supplement 1. ThT fluorescence of tau-RNA CC. 100 μM of tau187 and tau187-SL was mixed with 300 μg/mL polyU RNA (RNA) at room temperature. 100 μM tau187 with 25 μM heparin was prepared as reference. Ribbon shows the standard deviation of three replicates.CC BY · PubMed Central
Figure 1—figure supplement 2. RNase induces turbidity decrease of tau187-RNA CC suspension. Tau187-RNA CCs were re-prepared with 100 μM tau187C291S and 300 μg/mL polyU RNA, while absorbance at 500 nm was monitored. Samples were treated with buffer (Ctrl), 1 U/μL RNase inhibitor (mainly inhibiting RNase A), 50 ng/mL RNase A, and 50 ng/mL RNase A with 1 U/μL RNase inhibitor.
Figure 1—figure supplement 2. RNase induces turbidity decrease of tau187-RNA CC suspension. Tau187-RNA CCs were re-prepared with 100 μM tau187C291S and 300 μg/mL polyU RNA, while absorbance at 500 nm was monitored. Samples were treated with buffer (Ctrl), 1 U/μL RNase inhibitor (mainly inhibiting RNase A), 50 ng/mL RNase A, and 50 ng/mL RNase A with 1 U/μL RNase inhibitor.CC BY · PubMed Central
Figure 1—figure supplement 3. Turbidity of tau-RNA CC at varying charge ratios and ionic strength. (A) PolyU RNA was titrated into 20 μM tau187C291S in the buffer of 20 mM ammonium acetate at pH 7. Charge ratio was calculated based on the estimated charge for tau,+11 per tau molecule and the estimated charge for RNA, −1 per nucleotide. Turbidity was assigned from absorbance at λ = 500 nm. Red line indicates the place where the estimated RNA:tau charge ratio equals to 1. (B) 20 μM tau was mixed with 60 μg/mL RNA so that RNA:tau charge ratio equals to 1. NaCl was titrated into the mixture while turbidity was monitored.
Figure 1—figure supplement 3. Turbidity of tau-RNA CC at varying charge ratios and ionic strength. (A) PolyU RNA was titrated into 20 μM tau187C291S in the buffer of 20 mM ammonium acetate at pH 7. Charge ratio was calculated based on the estimated charge for tau,+11 per tau molecule and the estimated charge for RNA, −1 per nucleotide. Turbidity was assigned from absorbance at λ = 500 nm. Red line indicates the place where the estimated RNA:tau charge ratio equals to 1. (B) 20 μM tau was mixed with 60 μg/mL RNA so that RNA:tau charge ratio equals to 1. NaCl was titrated into the mixture while turbidity was monitored.CC BY · PubMed Central
Figure 1—figure supplement 4. Tau-RNA CC upon addition of heparin. (A) 100 μM tau187C291S-SL and 300 μg/mL polyA RNA was mixed in the buffer of 20 mM ammonium acetate at pH 7, with 19 vol % glycerol. Images were taken after 10 min at room temperature. (B) Sample in A was re-prepared and incubated for 10 min. 25 μM heparin was added afterwards, immediately followed by imaging. The scale bars in both A and B are 50 μm long. (C) ThT fluorescences of samples in A and B were monitored with additional 10 μM ThT. (D) TEM images of sample in A after overnight incubation, showing no structures similar to amyloid fibrils. (E) TEM images of sample in B after overnight incubation, showing structures with length and width consistent with tau fibrils. The scale bars in both D and E are 200 nm long.
Figure 1—figure supplement 4. Tau-RNA CC upon addition of heparin. (A) 100 μM tau187C291S-SL and 300 μg/mL polyA RNA was mixed in the buffer of 20 mM ammonium acetate at pH 7, with 19 vol % glycerol. Images were taken after 10 min at room temperature. (B) Sample in A was re-prepared and incubated for 10 min. 25 μM heparin was added afterwards, immediately followed by imaging. The scale bars in both A and B are 50 μm long. (C) ThT fluorescences of samples in A and B were monitored with additional 10 μM ThT. (D) TEM images of sample in A after overnight incubation, showing no structures similar to amyloid fibrils. (E) TEM images of sample in B after overnight incubation, showing structures with length and width consistent with tau fibrils. The scale bars in both D and E are 200 nm long.CC BY · PubMed Central
Figure 2. FH-VO modeling of tau-RNA complex coacervation. (A). Turbidity of tau187-RNA CC upon heating (Expt.1 ([tau], [NaCl]) = (50 μM, 120 mM), green dots; Expt.2 ([tau], [NaCl]) = (40 μM, 30 mM), purple dots). Absorbance at λ = 500 nm were normalized and used as turbidity value. Turbidity-temperature data of the heating curves were fitted with a sigmoidal function (solid line) as described in (Materials and methods), and the temperature at which normalized turbidity reaches 0.5 was assigned to cloud point, Tcp. (B)-(C). Experimental phase diagram (points) showing [tau] vs. Tcp and [NaCl] vs. Tcp along with the binodal curve generated from fitting the data to the FH-VO model with χ = χ(Tcp) (solid line). (D). Diagram of Flory Huggins lattice. Tau and RNA are represented by consecutively occupied lattice sites. (E). Each experimental condition in (A) was independently fit to the FH-VO model (solid lines) to obtain an empirical χ value. (E) shows two representative curves. These empirically determined values of χ are shown as points in (F). The solid line in (F) is a linear regression, generating χ = χ(Tcp), which is then used to generate the binodal lines in (B) and (C).
Figure 2. FH-VO modeling of tau-RNA complex coacervation. (A). Turbidity of tau187-RNA CC upon heating (Expt.1 ([tau], [NaCl]) = (50 μM, 120 mM), green dots; Expt.2 ([tau], [NaCl]) = (40 μM, 30 mM), purple dots). Absorbance at λ = 500 nm were normalized and used as turbidity value. Turbidity-temperature data of the heating curves were fitted with a sigmoidal function (solid line) as described in (Materials and methods), and the temperature at which normalized turbidity reaches 0.5 was assigned to cloud point, Tcp. (B)-(C). Experimental phase diagram (points) showing [tau] vs. Tcp and [NaCl] vs. Tcp along with the binodal curve generated from fitting the data to the FH-VO model with χ = χ(Tcp) (solid line). (D). Diagram of Flory Huggins lattice. Tau and RNA are represented by consecutively occupied lattice sites. (E). Each experimental condition in (A) was independently fit to the FH-VO model (solid lines) to obtain an empirical χ value. (E) shows two representative curves. These empirically determined values of χ are shown as points in (F). The solid line in (F) is a linear regression, generating χ = χ(Tcp), which is then used to generate the binodal lines in (B) and (C).CC BY · PubMed Central
Figure 2—figure supplement 1. Turbidity-temperature data and cloud points determinations of various [tau] and [NaCl]. Tau187-RNA CCs were prepared at various concentration of [tau] and [NaCl] with fixed ratio of [RNA]:[tau], shown as strip text (e.g. ‘2 6 30’ refers to [tau]=2 μM, [RNA]=6 μg/mL, [NaCl]=30 mM). Samples were kept at 4°C before ramping up temperature at 1°C/min. Absorbance at 500 nm was monitored and used as turbidity (grey points). Turbidity-temperature data were fit to a sigmoid curve (black solid line), where the cloud point, Tcp, were determined (blue vertical line). [tau], [NaCl] and Tcp were used for theory and simulation modeling.
Figure 2—figure supplement 1. Turbidity-temperature data and cloud points determinations of various [tau] and [NaCl]. Tau187-RNA CCs were prepared at various concentration of [tau] and [NaCl] with fixed ratio of [RNA]:[tau], shown as strip text (e.g. ‘2 6 30’ refers to [tau]=2 μM, [RNA]=6 μg/mL, [NaCl]=30 mM). Samples were kept at 4°C before ramping up temperature at 1°C/min. Absorbance at 500 nm was monitored and used as turbidity (grey points). Turbidity-temperature data were fit to a sigmoid curve (black solid line), where the cloud point, Tcp, were determined (blue vertical line). [tau], [NaCl] and Tcp were used for theory and simulation modeling.CC BY · PubMed Central
Figure 2—figure supplement 2. Phase diagrams of tau114 vs tau187. Tau-RNA CC were prepared with 20 μM tau187 with 60 μg/mL RNA (tau187) or 28 μM tau114 with 84 μg/mL RNA (tau114). (A) Turbidity was recorded with varying NaCl concentration. [NaCl] where turbidity vanished were determined by linear fitting to be 131 mM for tau187 and 150 mM for tau114. (B) [NaCl] vs. Tcp vs. [NaCl] curves for 20 μM tau187 or 28 μM tau114 were computed by FH-VO with χ = χ(Tcp) obtained from Figure 2D, shown together with the experimental data points obtained from (A).
Figure 2—figure supplement 2. Phase diagrams of tau114 vs tau187. Tau-RNA CC were prepared with 20 μM tau187 with 60 μg/mL RNA (tau187) or 28 μM tau114 with 84 μg/mL RNA (tau114). (A) Turbidity was recorded with varying NaCl concentration. [NaCl] where turbidity vanished were determined by linear fitting to be 131 mM for tau187 and 150 mM for tau114. (B) [NaCl] vs. Tcp vs. [NaCl] curves for 20 μM tau187 or 28 μM tau114 were computed by FH-VO with χ = χ(Tcp) obtained from Figure 2D, shown together with the experimental data points obtained from (A).CC BY · PubMed Central
Figure 2—figure supplement 3. Full phase diagram of tau187-RNA CC. Experimental data showing [tau] vs Tcp was replot from Figure 2B (points). Binodal curves was generated by fitting the data to the FH-VO model (solid lines).
Figure 2—figure supplement 3. Full phase diagram of tau187-RNA CC. Experimental data showing [tau] vs Tcp was replot from Figure 2B (points). Binodal curves was generated by fitting the data to the FH-VO model (solid lines).CC BY · PubMed Central
Figure 3. Schematic depiction of the tau and RNA polyelectrolyte models. Tau and RNA molecules are represented as bead-spring polymers with segment length b in implicit solvent. Tau is modeled as a polyampholyte with the charge of each monomer determined from the amino acid charge at pH = 7. RNA is modeled as a fully charged polyelectrolyte. In addition to chain connectivity, all monomers interact with an excluded volume repulsive potential, and charged monomers interact with a long-ranged Coulomb potential.
Figure 3. Schematic depiction of the tau and RNA polyelectrolyte models. Tau and RNA molecules are represented as bead-spring polymers with segment length b in implicit solvent. Tau is modeled as a polyampholyte with the charge of each monomer determined from the amino acid charge at pH = 7. RNA is modeled as a fully charged polyelectrolyte. In addition to chain connectivity, all monomers interact with an excluded volume repulsive potential, and charged monomers interact with a long-ranged Coulomb potential.CC BY · PubMed Central
Figure 4. Polymer density field of single phase solution and two phase coacervate from FTS. Left: Polymer density profile showing a single solution phase for the condition of relatively weak electrostatic strength lB = 0.16 b and relatively high excluded volume (good solvent conditions) v = 0.02 b3. The solution phase is characterized by near homogeneous low polymer density (white/light blue) throughout the entire simulation box. Right: Polymer density profile showing complex coacervation upon increasing the electrostatic strength to lB = 3.25 b and decreasing the solvent quality by lowering the excluded volume to v = 0.0068 b3. The two phase region is characterized by a distinct region of high polymer density (dark red) and a surrounding region of low polymer density (white) within the same simulation box. The total polymer concentration is the same in both simulations.
Figure 4. Polymer density field of single phase solution and two phase coacervate from FTS. Left: Polymer density profile showing a single solution phase for the condition of relatively weak electrostatic strength lB = 0.16 b and relatively high excluded volume (good solvent conditions) v = 0.02 b3. The solution phase is characterized by near homogeneous low polymer density (white/light blue) throughout the entire simulation box. Right: Polymer density profile showing complex coacervation upon increasing the electrostatic strength to lB = 3.25 b and decreasing the solvent quality by lowering the excluded volume to v = 0.0068 b3. The two phase region is characterized by a distinct region of high polymer density (dark red) and a surrounding region of low polymer density (white) within the same simulation box. The total polymer concentration is the same in both simulations.CC BY · PubMed Central
Figure 5. Phase diagram of the tau-RNA model obtained from FTS. Left: Binodal points as a function of the Bjerrum length at fixed excluded volume of v = 0.0068 b3. For comparison three concentrations at 300 K are indicated (arrows) assuming ϵr = 80 and b = 4 Å. Right: Binodal points as a function of the excluded volume at fixed Bjerrum length lB = 1.79 b.
Figure 5. Phase diagram of the tau-RNA model obtained from FTS. Left: Binodal points as a function of the Bjerrum length at fixed excluded volume of v = 0.0068 b3. For comparison three concentrations at 300 K are indicated (arrows) assuming ϵr = 80 and b = 4 Å. Right: Binodal points as a function of the excluded volume at fixed Bjerrum length lB = 1.79 b.CC BY · PubMed Central
Figure 5—figure supplement 1. Determination of phase coexistence points from FTS. (Left). Parametric plot of FTS simulation data points of the osmotic pressure vs the chemical potential. The dilute phase is shown in blue and the concentrated phase (complex coacervate) is shown in red. The intersection point of the two branches is the condition for simultaneous equivalence of pressure and chemical potential for the two phases. (Right) The chemical potential vs. the total polymer density from FTS simulation (red points). The horizontal dashed line is the chemical potential at the phase coexistence point. Intersection of the data points with the horizontal dashed lines determines the supernatant (I) and complex-coacervate (II) coexisting polymer densities.
Figure 5—figure supplement 1. Determination of phase coexistence points from FTS. (Left). Parametric plot of FTS simulation data points of the osmotic pressure vs the chemical potential. The dilute phase is shown in blue and the concentrated phase (complex coacervate) is shown in red. The intersection point of the two branches is the condition for simultaneous equivalence of pressure and chemical potential for the two phases. (Right) The chemical potential vs. the total polymer density from FTS simulation (red points). The horizontal dashed line is the chemical potential at the phase coexistence point. Intersection of the data points with the horizontal dashed lines determines the supernatant (I) and complex-coacervate (II) coexisting polymer densities.CC BY · PubMed Central
Figure 6. LCST phase behavior recapitulated from FTS. (A) Temperature dependence of the reduced Bjerrum length (εr = 80 for water) shown in red. (B) Temperature dependence of the excluded volume obtained by adjusting the excluded volume parameter until FTS agrees with a subset of the experimental data (points). The solid line shows a linear fit to the data which is used to obtain the temperature dependent excluded volume for subsequent simulations. (C) FTS coexistence points (filled green squares) obtained by using a temperature dependent Bjerrum length (A) and excluded volume (B). Experiments performed at 20 mM NaCl are shown in red for comparison. Upon introducing excess salt ions in FTS with a fixed concentration of [NaCl] = 120 mM, the binodal shifts upwards (open green squares). For comparison experiments performed at 120 mM NaCl are shown in blue.
Figure 6. LCST phase behavior recapitulated from FTS. (A) Temperature dependence of the reduced Bjerrum length (εr = 80 for water) shown in red. (B) Temperature dependence of the excluded volume obtained by adjusting the excluded volume parameter until FTS agrees with a subset of the experimental data (points). The solid line shows a linear fit to the data which is used to obtain the temperature dependent excluded volume for subsequent simulations. (C) FTS coexistence points (filled green squares) obtained by using a temperature dependent Bjerrum length (A) and excluded volume (B). Experiments performed at 20 mM NaCl are shown in red for comparison. Upon introducing excess salt ions in FTS with a fixed concentration of [NaCl] = 120 mM, the binodal shifts upwards (open green squares). For comparison experiments performed at 120 mM NaCl are shown in blue.CC BY · PubMed Central
Figure 6—figure supplement 1. Full phase diagram of tau187-RNA CC from FTS. Low-density points (left) are reproduced from Figure 6C in the main text. Filled green (low salt) and open green (120 mM NaCl) are obtain from FTS, and red and blue circles are corresponding experimental data. The right side of the figure shows the corresponding high-density binodal curve predicted from FTS.
Figure 6—figure supplement 1. Full phase diagram of tau187-RNA CC from FTS. Low-density points (left) are reproduced from Figure 6C in the main text. Filled green (low salt) and open green (120 mM NaCl) are obtain from FTS, and red and blue circles are corresponding experimental data. The right side of the figure shows the corresponding high-density binodal curve predicted from FTS.CC BY · PubMed Central
Figure 6—figure supplement 2. Tau in vitro phosphorylation. Tau (tau187C291S) were prepared as described in Materials and methods. Tau was subjected to in vitro phosphorylation using mouse brain extract as previously described (Despres et al., 2017). Phosphorylation was confirmed using SDS-PAGE and western blot analysis. Ponceau S staining of the blot shows equal loading of samples before (Tau) and after (P-Tau) the phosphorylation reaction, while reactivity with PHF-1 antibody indicates phosphorylation at S396/404.
Figure 6—figure supplement 2. Tau in vitro phosphorylation. Tau (tau187C291S) were prepared as described in Materials and methods. Tau was subjected to in vitro phosphorylation using mouse brain extract as previously described (Despres et al., 2017). Phosphorylation was confirmed using SDS-PAGE and western blot analysis. Ponceau S staining of the blot shows equal loading of samples before (Tau) and after (P-Tau) the phosphorylation reaction, while reactivity with PHF-1 antibody indicates phosphorylation at S396/404.CC BY · PubMed Central
Figure 6—figure supplement 3. Phosphorylation on tau-RNA complex coacervation. (A) 20 μM Tau and P-Tau was titrated with PolyU RNA. Absorbance at 500 nm was monitored and plotted as turbidity. (B) Corresponding representative bright field microscope images of samples in (A). (C) 20 μM Tau and P-Tau was mixed with 50 μg/mL and 100 μg/mL PolyU RNA, respectively. Mixtures were titrated with NaCl. Turbidity was plot (solid points) and fit using linear regression (dashed lines). (D) Corresponding representative bright-field microscope images of samples in (C). (Experiments were performed in 20 mM HEPES, pH = 7.0 with 1 mM TCEP. P-Tau concentration was determined by BCA assay. Tau concentration was determined by UV-Vis spectroscopy as described in Materials and methods).
Figure 6—figure supplement 3. Phosphorylation on tau-RNA complex coacervation. (A) 20 μM Tau and P-Tau was titrated with PolyU RNA. Absorbance at 500 nm was monitored and plotted as turbidity. (B) Corresponding representative bright field microscope images of samples in (A). (C) 20 μM Tau and P-Tau was mixed with 50 μg/mL and 100 μg/mL PolyU RNA, respectively. Mixtures were titrated with NaCl. Turbidity was plot (solid points) and fit using linear regression (dashed lines). (D) Corresponding representative bright-field microscope images of samples in (C). (Experiments were performed in 20 mM HEPES, pH = 7.0 with 1 mM TCEP. P-Tau concentration was determined by BCA assay. Tau concentration was determined by UV-Vis spectroscopy as described in Materials and methods).CC BY · PubMed Central
Figure 6—figure supplement 4. Concentration of protein in the solution phase and coacervate phase predicted from FTS simulation performed at 20°C on Tau (blue) and P-tau (green) in excess salt. Phosphorylation is seen to decrease the concentration difference between the two phases indicating a narrowing of the two phase window.
Figure 6—figure supplement 4. Concentration of protein in the solution phase and coacervate phase predicted from FTS simulation performed at 20°C on Tau (blue) and P-tau (green) in excess salt. Phosphorylation is seen to decrease the concentration difference between the two phases indicating a narrowing of the two phase window.CC BY · PubMed Central
Figure 7. Tuning experimental conditions to catch tau-RNA complex coacervates in the presence of living cells. Bright field images and superimposing fluorescence images of tau-RNA CC coculturing with H4 cells, with 10 μM tau (left), 100 μM tau (middle) and 10 μM tau with 10% v.v. PEG (right). Samples at 37 °C (first row) and 18 °C (second row) were images with representative images showing the co-presence of living cells and tau-RNA CCs. Tau187 (Top) and tau114 (Bottom) were used showing tau114 with higher propensity at CC formation. Alexa Fluor 488 was used to prepare fluorescent labeled tau. 3 μg/mL polyU RNA per 1 μM tau was used to prepared samples. Scale bar is 20 μm long.
Figure 7. Tuning experimental conditions to catch tau-RNA complex coacervates in the presence of living cells. Bright field images and superimposing fluorescence images of tau-RNA CC coculturing with H4 cells, with 10 μM tau (left), 100 μM tau (middle) and 10 μM tau with 10% v.v. PEG (right). Samples at 37 °C (first row) and 18 °C (second row) were images with representative images showing the co-presence of living cells and tau-RNA CCs. Tau187 (Top) and tau114 (Bottom) were used showing tau114 with higher propensity at CC formation. Alexa Fluor 488 was used to prepare fluorescent labeled tau. 3 μg/mL polyU RNA per 1 μM tau was used to prepared samples. Scale bar is 20 μm long.CC BY · PubMed Central
Author response image 1. pH changes of buffer. The pH values of 20 mM ammonium acetate and 20 mM HEPES were recorded upon addition of 20 μM tau or 60 μg/mL RNA.
Author response image 1. pH changes of buffer. The pH values of 20 mM ammonium acetate and 20 mM HEPES were recorded upon addition of 20 μM tau or 60 μg/mL RNA.CC BY · PubMed Central

Verification

Claims the paper makes

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

The self-check needs the paper’s full text.

Citations

Cites

Show all 8 references

Research works printed but not linked: 99

  1. Alexander-Katz, 2005, Field-theoretic simulations of polymer solutions: finite-size and discretization effects, The Journal of Chemical Physics, 122, 10.1063/1.1827211 · DOI
  2. Arneodo, 1988, Interfacial-Tension behavior of citrus oils against phases formed by complex coacervation of gelatin, ACS Symposium Series, 370, 132, 10.1021/bk-1988-0370.ch015 · DOI
  3. Banjade, 2015, Conserved Interdomain Linker promotes phase separation of the multivalent adaptor protein nck, PNAS, 112, E6426, 10.1073/pnas.1508778112 · DOI
  4. Banjade, 2014, Phase transitions of multivalent proteins can promote clustering of membrane receptors, eLife, 3, 10.7554/eLife.04123 · DOI
  5. Berry, 2015, RNA transcription modulates phase transition-driven nuclear body assembly, PNAS, 112, E5237, 10.1073/pnas.1509317112 · DOI · Paper page
  6. Bianconi, 2012, Temperature and solvent dependence of the dynamical landscape of tau protein conformations, Journal of Biological Physics, 38, 169, 10.1007/s10867-011-9244-6 · DOI
  7. Biesheuvel, 2004, Electrostatic free energy of weakly charged macromolecules in solution and intermacromolecular complexes consisting of oppositely charged polymers, Langmuir, 20, 2785, 10.1021/la036204l · DOI
  8. Biesheuvel, 2004, Cylindrical cell model for the electrostatic free energy of polyelectrolyte complexes, Langmuir, 20, 4764, 10.1021/la0496789 · DOI
  9. Boeynaems, 2017, Phase separation of C9orf72 dipeptide repeats perturbs stress granule dynamics, Molecular Cell, 65, 1044, 10.1016/j.molcel.2017.02.013 · DOI
  10. Borue, 1988, A statistical theory of weakly charged polyelectrolytes: fluctuations, equation of state and microphase separation, Macromolecules, 21, 3240, 10.1021/ma00189a019 · DOI
  11. Borue, 1990, A statistical theory of globular polyelectrolyte complexes, Macromolecules, 23, 3625, 10.1021/ma00217a015 · DOI
  12. Brady, 2017, Structural and hydrodynamic properties of an intrinsically disordered region of a germ cell-specific protein on phase separation, PNAS, 114, E8194, 10.1073/pnas.1706197114 · DOI · Paper page
  13. Brangwynne, 2009, Germline P granules are liquid droplets that localize by controlled dissolution/condensation, Science, 324, 1729, 10.1126/science.1172046 · DOI
  14. Brangwynne, 2015, Polymer physics of intracellular phase transitions, Nature Physics, 11, 899, 10.1038/nphys3532 · DOI
  15. Bungenberg de Jong, 1949, Crystallisation–coacervation–flocculation, Colloid Sci, 2, 232
  16. Burke, 2015, Vitro FUS granules that bind the C-Terminal domain of RNA polymerase II, Molecular Cell, 60, 231, 10.1016/j.molcel.2015.09.006 · DOI · Paper page
  17. Castelnovo, 2000, Formation of polyelectrolyte multilayers, Langmuir, 16, 7524, 10.1021/la000211h · DOI
  18. Castelnovo, 2001, Complexation between oppositely charged polyelectrolytes: beyond the random phase approximation, The European Physical Journal, 6, 377, 10.1007/s10189-001-8051-7 · DOI
  19. Chang, 2017, Sequence and entropy-based control of complex coacervates, Nature Communications, 8, 10.1038/s41467-017-01249-1 · DOI
  20. Choi, 2015, Entropic mechanism for the lower critical solution temperature of poly(ethylene oxide) in a room temperature ionic liquid, ACS Macro Letters, 4, 799, 10.1021/acsmacrolett.5b00355 · DOI
  21. Chollakup, 2010, Phase behavior and coacervation of aqueous poly(acrylic acid)−poly(allylamine) Solutions, Macromolecules, 43, 2518, 10.1021/ma902144k · DOI
  22. Das, 2018, A lattice model of Charge-Pattern-Dependent polyampholyte phase separation, The Journal of Physical Chemistry B, 122, 5418, 10.1021/acs.jpcb.7b11723 · DOI
  23. de la Cruz, 1995, Precipitation of highly charged polyelectrolyte solutions in the presence of multivalent salts, The Journal of Chemical Physics, 103, 5781, 10.1063/1.470459 · DOI
  24. Delaney, 2016, Recent developments in fully fluctuating Field-Theoretic simulations of polymer melts and solutions, The Journal of Physical Chemistry B, 120, 7615, 10.1021/acs.jpcb.6b05704 · DOI
  25. Delaney, 2017, Theory of polyelectrolyte complexation-Complex coacervates are self-coacervates, The Journal of Chemical Physics, 146, 10.1063/1.4985568 · DOI
  26. Despres, 2017, Identification of the tau phosphorylation pattern that drives its aggregation, PNAS, 114, 9080, 10.1073/pnas.1708448114 · DOI
  27. Dias, 2014, Pressure-Dependent properties of elementary hydrophobic interactions: ramifications for activation properties of protein folding, The Journal of Physical Chemistry B, 118, 7488, 10.1021/jp501935f · DOI
  28. Dill, 1989, Thermal stabilities of globular proteins, Biochemistry, 28, 5439, 10.1021/bi00439a019 · DOI
  29. Dobrynin, 2001, Counterion condensation and phase separation in solutions of hydrophobic polyelectrolytes, Macromolecules, 34, 1964, 10.1021/ma001619o · DOI
  30. Doi, 1988, The Theory of Polymer Dynamics
  31. Düchs, 2014, A multi-species exchange model for fully fluctuating Polymer field theory simulations, The Journal of Chemical Physics, 141, 10.1063/1.4900574 · DOI
  32. Elbaum-Garfinkle, 2015, The disordered P granule protein LAF-1 drives phase separation into droplets with tunable viscosity and dynamics, PNAS, 112, 7189, 10.1073/pnas.1504822112 · DOI · Paper page
  33. Eschmann, 2017, Signature of an aggregation-prone conformation of tau, Scientific Reports, 7, 10.1038/srep44739 · DOI
  34. Feil, 1993, Effect of comonomer hydrophilicity and ionization on the lower critical solution temperature of N-isopropylacrylamide copolymers, Macromolecules, 26, 2496, 10.1021/ma00062a016 · DOI
  35. Finka, 2013, Proteomic data from human cell cultures refine mechanisms of chaperone-mediated protein homeostasis, Cell Stress and Chaperones, 18, 591, 10.1007/s12192-013-0413-3 · DOI · Paper page
  36. Flory, 1969, Statistical mechanics of chain molecules, Biopolymers, 8, 699, 10.1002/bip.1969.360080514 · DOI
  37. Fredrickson, 2006, The Equilibrium Theory of Inhomogeneous Polymers
  38. Fu, 2016, Driving forces for oppositely charged polyion association in aqueous solutions: enthalpic, entropic, but not electrostatic, Journal of the American Chemical Society, 138, 980, 10.1021/jacs.5b11878 · DOI
  39. Gennes, 1979, Scaling Concepts in Polymer Physics
  40. Gummel, 2007, Counterions release from electrostatic complexes of polyelectrolytes and proteins of opposite charge: a direct measurement, Journal of the American Chemical Society, 129, 5806, 10.1021/ja070414t · DOI
  41. Holehouse, 2015, Quantitative assessments of the distinct contributions of polypeptide backbone amides versus side chain groups to chain expansion via chemical denaturation, Journal of the American Chemical Society, 137, 2984, 10.1021/ja512062h · DOI
  42. Hone, 2000, A Small-Angle neutron scattering study of the structure of gelatin/Polyelectrolyte complexes, Macromolecules, 33, 1206, 10.1021/ma9911750 · DOI
  43. Hückel, 1923, The theory of electrolytes: I. lowering of freezing point and related phenomena, Phys Z, 24, 185
  44. Hyman, 2014, Liquid-liquid phase separation in biology, Annual Review of Cell and Developmental Biology, 30, 39, 10.1146/annurev-cellbio-100913-013325 · DOI · Paper page
  45. Jeon, 2016, Effects of molecular crowding and confinement on the spatial organization of a biopolymer, Soft Matter, 12, 9436, 10.1039/C6SM01184E · DOI
  46. Kim, 2016, Complexation and coacervation of like-charged polyelectrolytes inspired by mussels, PNAS, 113, E847, 10.1073/pnas.1521521113 · DOI
  47. Klauder, 1983, A Langevin approach to fermion and quantum spin correlation functions, Journal of Physics A: Mathematical and General, 16, L317, 10.1088/0305-4470/16/10/001 · DOI
  48. Kudlay, 2004, Complexation of oppositely charged polyelectrolytes: Effect of Ion Pair Formation, Macromolecules, 37, 9231, 10.1021/ma048519t · DOI
  49. Kudlay, 2004, Precipitation of oppositely charged polyelectrolytes in salt solutions, The Journal of Chemical Physics, 120, 404, 10.1063/1.1629271 · DOI
  50. Kwon, 2014, Poly-dipeptides encoded by the C9orf72 repeats bind nucleoli, impede RNA biogenesis, and kill cells, Science, 345, 1139, 10.1126/science.1254917 · DOI
  51. Kwon, 2015, Theoretical analysis of multiple phase coexistence in polyelectrolyte blends, Macromolecules, 48, 6008, 10.1021/acs.macromol.5b00901 · DOI
  52. Lacombe, 1976, Statistical thermodynamics of fluid mixtures, The Journal of Physical Chemistry, 80, 2568, 10.1021/j100564a009 · DOI
  53. Lee, 2008, Complex coacervation: a field theoretic simulation study of polyelectrolyte complexation, The Journal of Chemical Physics, 128, 10.1063/1.2936834 · DOI
  54. Lee, 2016, C9orf72 dipeptide repeats impair the assembly, dynamics, and function of Membrane-Less organelles, Cell, 167, 774, 10.1016/j.cell.2016.10.002 · DOI
  55. Li, 2013, Stress granules as crucibles of ALS pathogenesis, The Journal of Cell Biology, 201, 361, 10.1083/jcb.201302044 · DOI
  56. Li, 2018, Phase behavior and salt partitioning in polyelectrolyte complex coacervates, Macromolecules, 51, 2988, 10.1021/acs.macromol.8b00238 · DOI
  57. Li, 2018, TAR DNA-binding protein 43 (TDP-43) liquid-liquid phase separation is mediated by just a few aromatic residues, The Journal of Biological Chemistry, 293, 10.1074/jbc.AC117.001037 · DOI
  58. Lin, 2016, Sequence-Specific polyampholyte phase separation in membraneless organelles, Physical Review Letters, 117, 10.1103/PhysRevLett.117.178101 · DOI
  59. Lin, 2017, Random-phase-approximation theory for sequence-dependent, biologically functional liquid-liquid phase separation of intrinsically disordered proteins, Journal of Molecular Liquids, 228, 176, 10.1016/j.molliq.2016.09.090 · DOI
  60. Lin, 2018, Theories for Sequence-Dependent phase behaviors of biomolecular condensates, Biochemistry, 57, 2499, 10.1021/acs.biochem.8b00058 · DOI
  61. Lytle, 2017, Transfer matrix theory of polymer complex coacervation, Soft Matter, 13, 7001, 10.1039/C7SM01080J · DOI
  62. Mair, 2016, FLEXITau: quantifying Post-translational modifications of tau protein in Vitro and in Human Disease, Analytical Chemistry, 88, 3704, 10.1021/acs.analchem.5b04509 · DOI
  63. Makhatadze, 1993, Contribution of hydration to protein folding thermodynamics. I. the enthalpy of hydration, Journal of Molecular Biology, 232, 639, 10.1006/jmbi.1993.1416 · DOI
  64. Matubayasi, 2017, Free-energy analysis of protein solvation with all-atom molecular dynamics simulation combined with a theory of solutions, Current Opinion in Structural Biology, 43, 45, 10.1016/j.sbi.2016.10.005 · DOI
  65. Molliex, 2015, Phase separation by low complexity domains promotes stress granule assembly and drives pathological fibrillization, Cell, 163, 123, 10.1016/j.cell.2015.09.015 · DOI · Paper page
  66. Murakami, 2015, ALS/FTD Mutation-Induced phase transition of FUS liquid droplets and reversible hydrogels into irreversible hydrogels impairs RNP granule function, Neuron, 88, 678, 10.1016/j.neuron.2015.10.030 · DOI · Paper page
  67. Muthukumar, 2004, Theory of counter-ion condensation on flexible polyelectrolytes: adsorption mechanism, The Journal of Chemical Physics, 120, 9343, 10.1063/1.1701839 · DOI
  68. Nakajima, 1972, Phase relationships of an equivalent mixture of sulfated polyvinyl alcohol and aminoacetalyzed polyvinyl alcohol in microsalt aqueous solution, Biopolymers, 11, 1345, 10.1002/bip.1972.360110704 · DOI
  69. Nott, 2015, Phase transition of a disordered nuage protein generates environmentally responsive membraneless organelles, Molecular Cell, 57, 936, 10.1016/j.molcel.2015.01.013 · DOI · Paper page
  70. Overbeek, 1957, Phase separation in polyelectrolyte solutions. Theory of complex coacervation, Journal of Cellular and Comparative Physiology, 49, 7, 10.1002/jcp.1030490404 · DOI
  71. Parisi, 1983, On complex probabilities, Physics Letters B, 131, 393, 10.1016/0370-2693(83)90525-7 · DOI
  72. Patel, 2015, A Liquid-to-Solid phase transition of the ALS protein FUS accelerated by disease mutation, Cell, 162, 1066, 10.1016/j.cell.2015.07.047 · DOI · Paper page
  73. Pavlova, 2009, Site-specific dynamic nuclear polarization of hydration water as a generally applicable approach to monitor protein aggregation, Physical Chemistry Chemical Physics, 11, 6833, 10.1039/b906101k · DOI
  74. Pavlova, 2016, Protein structural and surface water rearrangement constitute major events in the earliest aggregation stages of tau, PNAS, 113, E127, 10.1073/pnas.1504415113 · DOI
  75. Peterson, 2008, A soluble oligomer of tau associated with fiber formation analyzed by NMR, Biochemistry, 47, 7393, 10.1021/bi702466a · DOI
  76. Popov, 2007, Field-theoretic simulations of polyelectrolyte complexation, Journal of Polymer Science Part B: Polymer Physics, 45, 3223, 10.1002/polb.21334 · DOI
  77. Qin, 2016, Criticality and connectivity in macromolecular charge complexation, Macromolecules, 49, 8789, 10.1021/acs.macromol.6b02113 · DOI
  78. Rubinstein, 2003, Polymer Physics, 10.1093/oso/9780198520597.001.0001 · DOI · Paper page
  79. Sanchez, 1978, Statistical thermodynamics of polymer solutions, Macromolecules, 11, 1145, 10.1021/ma60066a017 · DOI
  80. Shen, 2017, Electrostatic correlations and the polyelectrolyte self energy, The Journal of Chemical Physics, 146, 10.1063/1.4975777 · DOI
  81. Shen, 2018, Polyelectrolyte chain structure and solution phase behavior, Macromolecules, 51, 1706, 10.1021/acs.macromol.7b02685 · DOI
  82. Silverstein, 2000, The strength of hydrogen bonds in liquid water and around nonpolar solutes, Journal of the American Chemical Society, 122, 8037, 10.1021/ja000459t · DOI
  83. Sing, 2017, Development of the modern theory of polymeric complex coacervation, Advances in Colloid and Interface Science, 239, 2, 10.1016/j.cis.2016.04.004 · DOI
  84. Siow, 1972, Cloud-Point curves in polymer solutions with adjacent upper and lower critical solution temperatures, Macromolecules, 5, 29, 10.1021/ma60025a008 · DOI
  85. Spruijt, 2010, Binodal compositions of polyelectrolyte complexes, Macromolecules, 43, 6476, 10.1021/ma101031t · DOI
  86. Stockmayer, 1955, Chain dimensions near the flory temperature, Journal of Polymer Science, 15, 595, 10.1002/pol.1955.120158028 · DOI
  87. Suzuki, 1982, An estimation of the temperature dependence of the excluded-volume integral, European Polymer Journal, 18, 247, 10.1016/0014-3057(82)90115-X · DOI
  88. Tainaka, 1979, Study of complex coacervation in low concentration by virial expansion method. I. salt free systems, Journal of the Physical Society of Japan, 46, 1899, 10.1143/JPSJ.46.1899 · DOI
  89. Tainaka, 1980, Effect of counterions on complex coacervation, Biopolymers, 19, 1289, 10.1002/bip.1980.360190705 · DOI
  90. Thirumalai, 2012, Role of water in protein aggregation and amyloid polymorphism, Accounts of Chemical Research, 45, 83, 10.1021/ar2000869 · DOI
  91. Uversky, 2015, Intrinsically disordered proteins as crucial constituents of cellular aqueous two phase systems and coacervates, FEBS Letters, 589, 15, 10.1016/j.febslet.2014.11.028 · DOI
  92. Veis, 1963, Phase separation in polyelectrolyte systems. iii. effect of aggregation and molecular weight heterogeneity, The Journal of Physical Chemistry, 67, 1960, 10.1021/j100804a004 · DOI
  93. Veis, 1960, Phase separation in polyelectrolyte systems. I. complex coacervates of gelatin, The Journal of Physical Chemistry, 64, 1203, 10.1021/j100838a022 · DOI
  94. Villet MC. 2012. Advanced computational field theory methods for fluctuating polymer solutions. ProQuest. https://search.proquest.com/openview/4c8b8e54782d4d6d5964834f0bdc88ec/1?pq-origsite=gscholar&cbl=18750&diss=y [Accessed June 16, 2018].
  95. Villet, 2014, Efficient field-theoretic simulation of polymer solutions, The Journal of Chemical Physics, 141, 10.1063/1.4902886 · DOI
  96. Water, 2014, Complex coacervates of hyaluronic acid and lysozyme: effect on protein structure and physical stability, European Journal of Pharmaceutics and Biopharmaceutics, 88, 325, 10.1016/j.ejpb.2014.09.001 · DOI
  97. Wegmann, 2018, Tau protein liquid-liquid phase separation can initiate tau aggregation, The EMBO Journal, 37, e98049, 10.15252/embj.201798049 · DOI · Paper page
  98. Wippich, 2013, Dual specificity kinase DYRK3 couples stress granule condensation/dissolution to mTORC1 signaling, Cell, 152, 791, 10.1016/j.cell.2013.01.033 · DOI
  99. Zhang, 2017, RNA stores tau reversibly in complex coacervates, PLOS Biology, 15, 10.1371/journal.pbio.2002183 · DOI · Paper page

Cited by

Metadata

Authors
Lin, Y., McCarty, J., Rauch, J. N., Delaney, K. T., Kosik, K. S., Fredrickson, G. H., Shea, J.-E., & Han, S.
Affiliations
  • Biomolecular Science and Engineering, University of California Santa Barbara, Santa Barbara, United States
  • Department of Chemistry and Biochemistry, University of California Santa Barbara, Santa Barbara, United States
  • Department of Molecular, Cellular and Developmental Biology, University of California Santa Barbara, Santa Barbara, United States
  • Materials Research Laboratory, University of California Santa Barbara, Santa Barbara, United States
ORCID
Venue
eLife
Publisher
eLife Sciences Publications, Ltd
Published
2019-04-05
Volume
8
ISSN
2050-084X
DOI
10.7554/elife.42571
PMC
PMC6450672
Cited by
165
References
8
Language
en
Licence
Funders
  • National Institute on Aging (RO1AG05605)
  • National Science Foundation (MRSEC Program, Award No. DMR 1720256, MCB-1716956)
  • Santa Barbara Foundation (Tri-Counties Blood Bank Fellowship)
  • Rainwater Foundation (Tau Consortium)

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review

Data sources

Bibliographic data: Semantic Scholar, OpenAlex, Crossref.

Contains information from the Semantic Scholar Academic Graph Datasets, made available under the ODC Attribution License (ODC-BY 1.0).

Semantic Scholar’s data is described in: Kinney, R., et al. (2023). The Semantic Scholar Open Data Platform. arXiv:2301.10140.