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Inflating bacterial cells by increased protein synthesis.

Basan M, Zhu M, Dai X, Warren M, Sévin D, Wang YP, Hwa T - Mol. Syst. Biol. (2015)

Bottom Line: Here, we characterized cell sizes in a set of orthogonal growth limitations.We report that cell size and mass exhibit positive or negative dependences with growth rate depending on the growth limitation applied.In particular, synthesizing large amounts of "useless" proteins led to an inversion of the canonical, positive relation, with slow growing cells enlarged 7- to 8-fold compared to cells growing at similar rates under nutrient limitation.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Systems Biology, ETH Zürich, Zürich, Switzerland basan@imsb.biol.ethz.ch hwa@ucsd.edu.

No MeSH data available.


Related in: MedlinePlus

Cell counts from Coulter counter and platingCell counts for the three types of growth limitations were obtained by using a Coulter counter on the one hand and plating on the other hand. In general, there is a very good agreement between the results of the Coulter counter and plating counts. For LacZ overexpression (OE) in glucose minimal medium, plating (red asterisks) resulted in only slightly lower values (10%) than the Coulter counter method (filled red diamonds), demonstrating that almost all cells in the culture are viable even at large cell sizes. For the nutrient limitation series, the results of the two methods give good agreement except for the very slow growth conditions (GR < 0.7 h−1), where the Coulter counter method (green filled circles) resulted in a lower cell number as compared to plating (green asterisks). This results from the small cell sizes under these conditions, which are close to the detection limit of the Coulter counter. For translation inhibition with Cm, the values of plating (blue asterisks) are lower compared to those obtained from the Coulter counter method (filled blue triangles) by 20%. This indicates that a fraction of cells in these conditions is not viable when subjected to plating. Throughout this manuscript, we use the cell count in order to determine per cell quantities. In these calculations, due to the small deviations discussed above, for the actual cell count, we employ the values of plating for nutrient limitation at slow growth rates, but use the values of the Coulter counter methods for both translation inhibition and LacZ overexpression.
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fig02ev: Cell counts from Coulter counter and platingCell counts for the three types of growth limitations were obtained by using a Coulter counter on the one hand and plating on the other hand. In general, there is a very good agreement between the results of the Coulter counter and plating counts. For LacZ overexpression (OE) in glucose minimal medium, plating (red asterisks) resulted in only slightly lower values (10%) than the Coulter counter method (filled red diamonds), demonstrating that almost all cells in the culture are viable even at large cell sizes. For the nutrient limitation series, the results of the two methods give good agreement except for the very slow growth conditions (GR < 0.7 h−1), where the Coulter counter method (green filled circles) resulted in a lower cell number as compared to plating (green asterisks). This results from the small cell sizes under these conditions, which are close to the detection limit of the Coulter counter. For translation inhibition with Cm, the values of plating (blue asterisks) are lower compared to those obtained from the Coulter counter method (filled blue triangles) by 20%. This indicates that a fraction of cells in these conditions is not viable when subjected to plating. Throughout this manuscript, we use the cell count in order to determine per cell quantities. In these calculations, due to the small deviations discussed above, for the actual cell count, we employ the values of plating for nutrient limitation at slow growth rates, but use the values of the Coulter counter methods for both translation inhibition and LacZ overexpression.

Mentions: We also characterized the macromolecular content of the culture, namely protein, RNA, and DNA, as well as the total dry mass and cell count (Appendix Table S4). The sum of protein, RNA, and DNA was found to account for ∼90% of dry mass (Appendix Fig S5), over two-third of which is protein for each of the growth limitations (Appendix Fig S6A–D). Cellular dry mass and cellular protein content, shown in Fig1D and Appendix Fig S7, respectively, displayed quantitatively similar trends as those exhibited by the physical cell size (Fig1C) for each growth perturbation, with a 7- to 8-fold overall difference between nutrient limitation and LacZ OE at the slowest GR. A priori, one might expect LacZ OE to result in an increased dry mass density and molecular crowding. Instead, a tight correlation was found between dry mass (also cellular protein) and physical cell size under all tested growth limitations (Figs1G and EV1A), despite large changes in cell size (Fig1C) and macromolecular composition (Appendix Fig S8). Correspondingly, the GR dependence of cell number (per volume of culture at constant OD600), as determined by Coulter counter and colony count (FigEV2), followed just the opposite trends as cell size. While most of the increase in cellular dry mass under LacZ OE was attributable to an increase in cellular protein (Appendix Fig S7), cellular RNA also exhibited a significant increase (Fig1E).


Inflating bacterial cells by increased protein synthesis.

Basan M, Zhu M, Dai X, Warren M, Sévin D, Wang YP, Hwa T - Mol. Syst. Biol. (2015)

Cell counts from Coulter counter and platingCell counts for the three types of growth limitations were obtained by using a Coulter counter on the one hand and plating on the other hand. In general, there is a very good agreement between the results of the Coulter counter and plating counts. For LacZ overexpression (OE) in glucose minimal medium, plating (red asterisks) resulted in only slightly lower values (10%) than the Coulter counter method (filled red diamonds), demonstrating that almost all cells in the culture are viable even at large cell sizes. For the nutrient limitation series, the results of the two methods give good agreement except for the very slow growth conditions (GR < 0.7 h−1), where the Coulter counter method (green filled circles) resulted in a lower cell number as compared to plating (green asterisks). This results from the small cell sizes under these conditions, which are close to the detection limit of the Coulter counter. For translation inhibition with Cm, the values of plating (blue asterisks) are lower compared to those obtained from the Coulter counter method (filled blue triangles) by 20%. This indicates that a fraction of cells in these conditions is not viable when subjected to plating. Throughout this manuscript, we use the cell count in order to determine per cell quantities. In these calculations, due to the small deviations discussed above, for the actual cell count, we employ the values of plating for nutrient limitation at slow growth rates, but use the values of the Coulter counter methods for both translation inhibition and LacZ overexpression.
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig02ev: Cell counts from Coulter counter and platingCell counts for the three types of growth limitations were obtained by using a Coulter counter on the one hand and plating on the other hand. In general, there is a very good agreement between the results of the Coulter counter and plating counts. For LacZ overexpression (OE) in glucose minimal medium, plating (red asterisks) resulted in only slightly lower values (10%) than the Coulter counter method (filled red diamonds), demonstrating that almost all cells in the culture are viable even at large cell sizes. For the nutrient limitation series, the results of the two methods give good agreement except for the very slow growth conditions (GR < 0.7 h−1), where the Coulter counter method (green filled circles) resulted in a lower cell number as compared to plating (green asterisks). This results from the small cell sizes under these conditions, which are close to the detection limit of the Coulter counter. For translation inhibition with Cm, the values of plating (blue asterisks) are lower compared to those obtained from the Coulter counter method (filled blue triangles) by 20%. This indicates that a fraction of cells in these conditions is not viable when subjected to plating. Throughout this manuscript, we use the cell count in order to determine per cell quantities. In these calculations, due to the small deviations discussed above, for the actual cell count, we employ the values of plating for nutrient limitation at slow growth rates, but use the values of the Coulter counter methods for both translation inhibition and LacZ overexpression.
Mentions: We also characterized the macromolecular content of the culture, namely protein, RNA, and DNA, as well as the total dry mass and cell count (Appendix Table S4). The sum of protein, RNA, and DNA was found to account for ∼90% of dry mass (Appendix Fig S5), over two-third of which is protein for each of the growth limitations (Appendix Fig S6A–D). Cellular dry mass and cellular protein content, shown in Fig1D and Appendix Fig S7, respectively, displayed quantitatively similar trends as those exhibited by the physical cell size (Fig1C) for each growth perturbation, with a 7- to 8-fold overall difference between nutrient limitation and LacZ OE at the slowest GR. A priori, one might expect LacZ OE to result in an increased dry mass density and molecular crowding. Instead, a tight correlation was found between dry mass (also cellular protein) and physical cell size under all tested growth limitations (Figs1G and EV1A), despite large changes in cell size (Fig1C) and macromolecular composition (Appendix Fig S8). Correspondingly, the GR dependence of cell number (per volume of culture at constant OD600), as determined by Coulter counter and colony count (FigEV2), followed just the opposite trends as cell size. While most of the increase in cellular dry mass under LacZ OE was attributable to an increase in cellular protein (Appendix Fig S7), cellular RNA also exhibited a significant increase (Fig1E).

Bottom Line: Here, we characterized cell sizes in a set of orthogonal growth limitations.We report that cell size and mass exhibit positive or negative dependences with growth rate depending on the growth limitation applied.In particular, synthesizing large amounts of "useless" proteins led to an inversion of the canonical, positive relation, with slow growing cells enlarged 7- to 8-fold compared to cells growing at similar rates under nutrient limitation.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Systems Biology, ETH Zürich, Zürich, Switzerland basan@imsb.biol.ethz.ch hwa@ucsd.edu.

No MeSH data available.


Related in: MedlinePlus