Cell-to-Cell Diversity in a Synchronized Chlamydomonas Culture As Revealed by Single-Cell Analyses

Abstract

In a synchronized photoautotrophic culture of Chlamydomonas reinhardtii, cell size, cell number, and the averaged starch content were determined throughout the light-dark cycle. For single-cell analyses, the relative cellular starch was quantified by measuring the second harmonic generation (SHG). In destained cells, amylopectin essentially represents the only biophotonic structure. As revealed by various validation procedures, SHG signal intensities are a reliable relative measure of the cellular starch content. During photosynthesis-driven starch biosynthesis, synchronized Chlamydomonas cells possess an unexpected cell-to-cell diversity both in size and starch content, but the starch-related heterogeneity largely exceeds that of size. The cellular volume, starch content, and amount of starch/cell volume obey lognormal distributions. Starch degradation was initiated by inhibiting the photosynthetic electron transport in illuminated cells or by darkening. Under both conditions, the averaged rate of starch degradation is almost constant, but it is higher in illuminated than in darkened cells. At the single-cell level, rates of starch degradation largely differ but are unrelated to the initial cellular starch content. A rate equation describing the cellular starch degradation is presented. SHG-based three-dimensional reconstructions of Chlamydomonas cells containing starch granules are shown.

Double Knockout Mutants of Arabidopsis Grown under Normal Conditions Reveal that the Plastidial Phosphorylase Isozyme Participates in Transitory Starch Metabolism

“…6, A-C and M-P). Similar results have previously been reported for leaves of Arabidopsis plants (CrumptonTaylor et al, 2012;Ingkasuwan et al, 2012) as well as for unicellular green alga C. reinhardtii (Garz et al, 2012). Secondly, when grown under a light-dark regime (12-h-light, 12-h-dark), the ultrastructural integrity of mesophyll cells of the two double mutants differed.…”

Systems-Wide Analysis of Acclimation Responses to Long-Term Heat Stress and Recovery in the Photosynthetic Model OrganismChlamydomonas reinhardtii

“…Prior to DNA staining, cells were destained and fixed with ethanol (Garz et al, 2012). Fixed cells were washed two times with ice-cold PBS buffer and were kept in PBS at 4°C.…”

Weak correlation of starch and volume in synchronized photosynthetic cells

“…[10], Single-cell analyses, however, provide a different view on the hom ogeneity o f synchronized cell cultures. Indeed, as reported in [11], synchronized cells o f C. reinhardtii exhibit a relatively broad distribution o f both cell sizes v and the cellular starch content y as shown by the coefficients o f variation o f cellular starch density, w hich ranges betw een 0.51 and 0.63. For the details about this issue, we refer the reader to A ppendix A. M oreover, throughout the entire cell cycle, v and y are only w eakly correlated [11].…”

“…Indeed, as reported in [11], synchronized cells o f C. reinhardtii exhibit a relatively broad distribution o f both cell sizes v and the cellular starch content y as shown by the coefficients o f variation o f cellular starch density, w hich ranges betw een 0.51 and 0.63. For the details about this issue, we refer the reader to A ppendix A. M oreover, throughout the entire cell cycle, v and y are only w eakly correlated [11]. Indeed, the Spearm an rank correlation coefficient betw een relative cellular starch content and cellular volum e ranges from 0.08 to 0.30, thus confirm ing that there is no obvious correlation.…”