PHYSICAL REVIEW E 91, 012711 (2015) Weak correlation of starch and volume in synchronized photosynthetic cells M. Michael Rading, 1 , * Michael Sandmann, 2 Martin Steup, 3 Davide Chiarugi, 1 and Angelo Valleriani 1 , † 1 Department of Theory and Bio-Systems, Max Planck Institute of Colloids and Interfaces, 14476 Potsdam, Germany 2 innoFSPEC, Institut f ¨ ur Chemie, Universit ¨ at Potsdam, Physikalische Chemie, 14476 Potsdam, Germany 3 Department of Molecular and Cellular Biology, College of Biological Science, University of Guelph, Guelph, Ontario, Canada N1G 2W1 (Received 22 August 2014; published 28 January 2015) In cultures of unicellular algae, features of single cells, such as cellular volume and starch content, are thought to be the result of carefully balanced growth and division processes. Single-cell analyses of synchronized photoautotrophic cultures of the unicellular alga Chlamydomonas reinhardtii reveal, however, that the cellular volume and starch content are only weakly correlated. Likewise, other cell parameters, e.g., the chlorophyll content per cell, are only weakly correlated with cell size. We derive the cell size distributions at the beginning of each synchronization cycle considering growth, timing of cell division and daughter cell release, and the uneven division of cell volume. Furthermore, we investigate the link between cell volume growth and starch accumulation. This work presents evidence that, under the experimental conditions of light-dark synchronized cultures, the weak correlation between both cell features is a result of a cumulative process rather than due to asymmetric partition of biomolecules during cell division. This cumulative process necessarily limits cellular similarities within a synchronized cell population. DOI: 10.1103/PhysRevE.91.012711 PACS number(s): 87.17.Ee, 87.17.Aa, 87.10.Mn I. INTRODUCTION For technical reasons, most studies in the Ohmic area require the analysis of relatively large cell populations. Essentially, this approach implies that the average values obtained from population measurements closely reflect data from single cells. However, this assumption is safe only when the cells composing a given population are homogeneous with respect to their physiological state, to their developmental states, and to the content of the analytes to be assayed. For obtaining homogeneous cell populations, experimen- talists rely on various techniques. To maintain both constant efficiency of external conditions and unchanged cell density, continuous cultures are typically used in which the culture medium is continuously renewed [1]. Under these conditions, the average growth rate can be adjusted to a constant value, i.e., the so-called steady-state or bound growth. The number of cells per suspension volume can also be kept constant if the number of daughter cells released exactly matches the number of cells lost through the outflow of the culture vessel. Uniformity of the cellular developmental state is obtained by synchronization techniques. These approaches aim at establishing cell cultures that, temporarily or permanently, will reside in the same phase of the cell cycle. To achieve this goal, various methods exist [2–4]. Nevertheless, single-cell analyses are providing increasing evidence that a truly homogeneous cell suspension is more difficult to obtain than expected. Indeed, the single cells com- posing isogenic populations can be largely different from each other even when they are grown in the same environment [5]. The reasons for the heterogeneity at the cellular level have been investigated for several model systems, including prokaryotic and eukaryotic cells, indicating that stochasticity in gene expression and cell division can play a major role [5–7]. * rading@uni-potsdam.de † angelo.valleriani@mpikg.mpg.de However, the impact of these sources of stochasticity on synchronized cell cultures is still not completely understood and must be carefully taken into consideration. We address this issue in this paper. In particular, we focus on the unicellular eukaryotic alga Chlamydomonas reinhardtii, which is one of the most studied plant model organisms. When grown photoautotrophically, the vegetative cell cycle of C. reinhardtii depends on the photoperiod. During the light period, photosynthesis drives the growth of cell volume, the DNA replication, and the biosynthesis of all other cellular constituents. The transition to the dark phase instead marks the release of daughter cells. These features are typically used for continuous synchronization of photoautotrophic cells. Exposing a cell culture of unicellular algae, such as Chlorella or Chlamydomonas, to a relatively short alternating series of light-dark phases results into a population that, at the onset of the light-dark cycle, consists essentially of young and small daughter cells. Photosynthesis-driven growth continues during illumination, leading to cell division and finally to the release of the offspring [8,9]. Readers interested in a comprehensive overview of the development of synchronized algal cells can consult Ref. [10]. Single-cell analyses, however, provide a different view on the homogeneity of synchronized cell cultures. Indeed, as reported in [11], synchronized cells of C. reinhardtii exhibit a relatively broad distribution of both cell sizes v and the cellular starch content y as shown by the coefficients of variation of cellular starch density, which ranges between 0.51 and 0.63. For the details about this issue, we refer the reader to Appendix A. Moreover, throughout the entire cell cycle, v and y are only weakly correlated [11]. Indeed, the Spearman rank correlation coefficient between relative cellular starch content and cellular volume ranges from 0.08 to 0.30, thus confirming that there is no obvious correlation. This finding is surprising because, intuitively, larger cells are expected to contain more starch than smaller cells. Furthermore, one may ask whether this counterintuitive experimental finding holds also for other cellular constituents. Our results will show that the ingredients 1539-3755/2015/91(1)/012711(11) 012711-1 ©2015 American Physical Society