Resource plasticity-driven carbon-nitrogen budgeting enables specialization and division of labor in a clonal community
- InStem - Institute for Stem Cell Science and Regenerative Medicine, India
- Simons Centre for the Study of Living Machines, National Center for Biological Sciences, Tata Institute for Fundamental Research, India
- Manipal Academy of Higher Education, India
Figures
Figure 1 with 1 supplement
Amino acid driven gluconeogenesis is critical for the emergence of phenotypic heterogeneity.
(A) External trehalose controls the emergence of light cells. Trehalose synthesized by the dark cells fuels glycolysis and pentose phosphate pathway in light cells. (B) Gluconeogenesis is required …
Figure 1—figure supplement 1
Amino acid dependent gluconeogenesis drives the development of complex colonies exhibiting specialized cell states.
(A) Gluconeogenesis is required for development of structural morphology in the colonies. The panel shows the morphology of mature wild-type and gluconeogenesis defective (∆pck1 and ∆fbp1) yeast …
Figure 2 with 1 supplement
Aspartate enables light cell emergence by fueling trehalose synthesis.
(A) Comparative steady-state amounts of trehalose measured in wild-type, ∆pck1 (gluconeogenesis defective) and ∆tps1 (trehalose synthesis defective) colonies grown in minimal medium, or minimal …
Figure 2—figure supplement 1
Amino acids, in particular aspartate is critical for the maintenance of light cell populations.
(A) The light cell population was calculated by spotting wild-type cells harboring the PPP reporter plasmid either in minimal media or minimal media supplemented with either all amino acids or …
Figure 3 with 5 supplements
An agent-based model for carbon-nitrogen budgeting reveals principles of metabolic heterogeneity via self-organization.
(A) A model schematic based on an experimental understanding of aspartate utilization by the two cell types in the system. Dark and light cells are colored accordingly. Dark cells take in aspartate, …
Figure 3—figure supplement 1
A histogram of the amount of trehalose secreted per unit time per dark cell throughout a simulation.
Dark cells secrete a fraction, Pf, of their internal carbon levels as trehalose. However, we place an upper limit on the absolute amount of secreted trehalose at 0.12 units/Time. This limit is meant …
Figure 3—figure supplement 2
Normalized histograms of dark and light cell block division times through one simulation with default parameters.
The bins along the x-axis measure the time steps elapsed between a cell block’s birth and when it divides. The cell blocks have the same probability of division once they build up sufficient N and C …
Figure 3—figure supplement 3
Different final colony compositions for different combinations of the main model parameters, ‘f’ and ‘AspU’.
Each panel is a generated colony using the values of ‘f’ and ‘AspU’ corresponding to its location. Note: the axes values are not on a linear or logarithmic scale but are chosen for representative …
Figure 3—figure supplement 4
Comparing simulated colonies generated with a low-switching rule against the older no-switching rule (light switching to dark).
(A) A colony where light cell blocks are not allowed to switch to dark cell blocks during the simulation. This is similar to the original model implemented in Varahan et al., 2019. (B) A colony …
Figure 3—figure supplement 5
A flowchart of the simulation algorithm highlighting all the processes in the mathematical model.
A step-by-step description is provided in the Materials and methods section of the main text.
Figure 4
Aspartate is differentially utilized as a carbon or nitrogen currency in light and dark cells.
(A) Metabolic fates of aspartate: The carbon of aspartate (green) can be used for synthesis of molecules like trehalose via gluconeogenesis (green boxed). The nitrogen of aspartate (red) is …
Figure 5
Dark and light cells exhibit division of labor and confer distinct survival and collective growth advantages to the whole colony.
(A) Equal numbers of light and dark cells were subjected to multiple freeze-thaw cycles, and survival estimated by spotting onto rich media plates and allowing growth for 18 hr. Cells grown in …
Figure 6
Model: Metabolite plasticity allowing differential carbon/nitrogen resource budgeting of aspartate drives metabolic specialization resulting in division of labor.
Cells in low glucose perform gluconeogenesis (Dark cells), as would be required in low glucose medium. During this process, dark cells predominantly budget aspartate for their carbon needs to …
Videos
Video 1
Development of a simulated wild-type colony.
A simulation movie of the WT colony over 750 time-steps (~6 days in real time). The colony starts with 95–99% dark cells, which go through switching and growth phases as observed. This colony is …
Video 2
Aspartate is allocated equally for Carbon and Nitrogen by dark cells.
A simulation movie where the budgeting fraction ‘f’ is 50%, i.e., 50% of the aspartate flux is allocated towards nitrogen reserves. The dark cell blocks cannot allocate sufficient carbon for …
Video 3
Aspartate uptake rate by both types of cells is equal to trehalose uptake rate by light cells.
A simulation movie where the relative rate of aspartate uptake is equal to the trehalose uptake rate. (i.e. AspU = 1.0). In this case, the aspartate uptake by dark cells is much slower, which also …
Video 4
Aspartate uptake rate by both types of cells is much higher than trehalose uptake rate by light cells.
A simulation movie where the relative rate of aspartate uptake is high. (i.e. AspU = 8.0). In this case, the dark cells allocate adequate aspartate for both nitrogen and carbon requirements rapidly …
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Gene (Saccharomyces cerevisiae) | pck1 | Saccharomyces genome database (SGD) | SGD:S000001805 | |
| Gene (Saccharomyces cerevisiae) | fbp1 | Saccharomyces genome database (SGD) | SGD:S000004369 | |
| Gene (Saccharomyces cerevisiae) | nth1 | Saccharomyces genome database (SGD) | SGD:S000002408 | |
| Strain, strain background (Saccharomyces cerevisiae) | Prototrophic sigma1278b, MATa (WT) | Isolate from Fink Lab. | YBC16G1 | Wild-type strain. |
| Strain, strain background (Saccharomyces cerevisiae) | Δpck1 | This study | sigma1278b MAT a pck1::kanMX6 | |
| Strain, strain background (Saccharomyces cerevisiae) | Δfbp1 | This study | sigma1278b MAT a fbp1::kanMX6 | |
| Strain, strain background (Saccharomyces cerevisiae) | Δnth1 | Varahan et al., 2019 | sigma1278b MAT a nth1::kanMX6 | |
| Strain, strain background (Saccharomyces cerevisiae) | WT (pTKL1-mCherry) | Varahan et al., 2019 | Wild-type strain with pentose phosphate pathway reporter plasmid (mCherry with TKL1 promoter) | |
| Strain, strain background (Saccharomyces cerevisiae) | Δpck1 (pTKL1-mCherry) | This study | Δpck1 strain with pentose phosphate pathway reporter plasmid (mCherry with TKL1 promoter) | |
| Strain, strain background (Saccharomyces cerevisiae) | Δfbp1 (pTKL1-mCherry) | This study | Δfbp1 strain with pentose phosphate pathway reporter plasmid (mCherry with TKL1 promoter) | |
| Recombinant DNA reagent | pTKL1-mCherry | Varahan et al., 2019 | mCherry under the TKL1 promoter and CYC1 terminator. p417 centromeric plasmid backbone, G418R. | |
| Commercial assay or kit | Glucose (GO) Assay Kit | Sigma Aldrich | Cat. #: GAGO20-1KT | Kit used for the biochemical measurement of trehalose from cells. |
| Chemical compound, drug | 15N Aspartate | Cambridge isotope laboratories | Cat. #: NLM-718-PK | |
| Chemical compound, drug | 15N Ammonium sulphate | Cambridge isotope laboratories | Cat. #: NLM-713-PK | |
| Chemical compound, drug | 13C Aspartate | Cambridge isotope laboratories | Cat. #: CLM-1801-PK |
Table 1
Mass transitions used for LC-MS/MS experiments.
| Nucleotides | Formula | Parent/Product (positive polarity) | Comment (for 15N experiment) |
|---|---|---|---|
| AMP | C10H14N5O7P | 348/136 | Product has all N |
| 15N_AMP_1 | 349/137 | ||
| 15N_AMP_2 | 350/138 | ||
| 15N_AMP_3 | 351/139 | ||
| 15N_AMP_4 | 352/140 | ||
| 15N_AMP_5 | 353/141 | ||
| GMP | C10H14N5O8P | 364/152 | Product has all N |
| 15N_GMP_1 | 365/153 | ||
| 15N_GMP_2 | 366/154 | ||
| 15N_GMP_3 | 367/155 | ||
| 15N_GMP_4 | 368/156 | ||
| 15N_GMP_5 | 369/157 | ||
| CMP | C9H14N3O8P | 324/112 | Product has all N |
| 15N_CMP_1 | 325/113 | ||
| 15N_CMP_2 | 326/114 | ||
| 15N_CMP_3 | 327/115 | ||
| UMP | C9H13N2O9P | 325/113 | Product has all N |
| 15N_UMP_1 | 326/114 | ||
| 15N_UMP_2 | 327/115 | ||
| Trehalose and sugar phosphates | Formula | Parent/Product (negative polarity) | Comment (for 13C experiment) |
| Trehalose | C12H22O11 | 341.3/179.3 | |
| 13C_Trehalose_12 | 353.3/185.3 | Product has 6 C all of which are labeled | |
| 13C_3 PG_3 | 188/97 | ||
| G6P | C6H13O9P | 259/97 | Monitoring the phosphate release |
| 13C_G6P_6 | 265/97 | ||
| 6 PG | C6H13O10P | 275/97 | Monitoring the phosphate release |
Table 2
Model parameters.
| Main parameters | Notation | Default Value | Range of Variation |
|---|---|---|---|
| Fraction of aspartate flux allocated to N in dark cell blocks | f | 0.125 | 0.0–1.0 (0–100%) |
| Relative rate of aspartate uptake compared to trehalose uptake rate | AspU | 4.0 | 1.0–8.0 |
| Additional parameters | |||
| Yield (converting N to C) | Y | 0.31 C/N | |
| Fraction secreted as trehalose, per dark cell block | Pf | 0.049/Time | -- |
| Max secreted trehalose, per dark cell block | -- | 0.12 units/Time | -- |
| Extra N for light cells | ExN | 4.0 | -- |
| Aspartate consumed by dark and light cell blocks | AspU*Cmax | 0.2/Time | |
| Parameters from previous model | |||
| Growth rate (light and dark cell block) | g | 0.04/Time | -- |
| Max trehalose consumed by a light cell block | Cmax | 0.05 units/Time | |
| Switching threshold (dark to light) | TDL | 1.5 units | -- |
| Switching probability (dark to light) | PDL | 0.5/Time | |
| Switching threshold (light to dark) | TLD | 0.0001 units | |
| Switching probability (light to dark) | PLD | 0.0001/Time | |
| Scaled diffusion constant of trehalose | Deff | 0.24 L2/Time |
