Photopolarization of Fucus zygotes is determined by time sensitive vectorial addition of environmental cues during axis amplification

Abstract

Fucoid zygotes have been extensively used to study cell polarization and asymmetrical cell division. Fertilized eggs are responsive to different environmental cues (e.g., light, gravity) for a long period before the polarity is fixed and the cells germinate accordingly. First, it is commonly believed that the direction and sense of the polarization vector are established simultaneously as indicated by the formation of an F-actin patch. Secondly, upon reorientation of the zygote, a new polar gradient is formed and it is assumed that the position of the future rhizoid pole is only influenced by the latter. Here we tested these two hypotheses investigating photopolarization in Fucus zygotes by reorienting zygotes 90° relative to a unilateral light source at different time points during the first cell cycle. We conclude that fixation of direction and sense of the polarization vector is indeed established simultaneously. However, the experiments yielded a distribution of polarization axes that cannot be explained if only the last environmental cue is supposed to determine the polarization axis. We conclude that our observations, together with published findings, can only be explained by assuming imprinting of the different polarization vectors and their integration as a vectorial sum at the moment of axis fixation. This way cells will average different serially perceived cues resulting in a polarization vector representative of the dynamic intertidal environment, instead of betting exclusively on the perceived vector at the moment of axis fixation.

Keywords: Brown algae; Fucus; asymmetrical cell division; embryogenesis; intrinsic factors; patterning; polarization; positional information.

Figure 1

Experimental design and scoring method and (A) 90° reorientation, (B) 180° reorientations. Angle between the vector pointing toward the first direction of the light (black line at 0°) and the polarization vector (arrow, determined by the rhizoid outgrowth and the center of the cell) was scored by estimating its orientation on a angular scale with a 45° resolution as depicted (dashed interval). Numbers depict borders of the intervals.

Figure 2

Influence of 90° reorientation experiments on the direction and sense of polarization vector. Second order mean of the direction and sense of the polarization vector after 90° reorientation is depicted by an arrow. Length of the arrow represents the r measure of the second order mean angle. Percentages represent the fraction of the population that shows a particular angular orientation of rhizoid outgrowth relative to the center of the cell. Angles represent the relative orientation of the rhizoid relative to the orientation of the first unidirectional light source. All mean angles of polarization vectors, including sense information, were statistically significant (df = 3, P < 0.05).

Figure 3

Influence of reorientation on the fixation of direction and sense of polarization vector. Second order mean angles for the polarization vector after 90° reorientation. Upper and lower error bars are Mardia's circular standard deviation for the mean angles (n > 200 in three replicate experiments). The line depicts the regressed Gompertz sigmoid that best describes the data.

Figure 4

Influence of 180° reorientation experiment on the polarization vector for the time points 10.5, 12, and 16.5 h AF. Percentages aligned vertically represent the percentages of the population that show a particular angular orientation of rhizoid outgrowth relative to the center of the cell. Angles represent the relative orientation of the rhizoid relative to the orientation of the first unidirectional light source (n = 300 in three replicate experiments).

Figure 5

Polarization after varying 180° reorientation over time and TBO staining in zygotes developing under unchanged unilateral light. Fraction of zygotes having a polarization vector that points toward the shaded side in relation to L1 (squares). Percentage of zygotes staining asymmetrically with TBO under unilateral light (triangles). Upper and lower error bars are standard deviations (n = 300 in three replicate experiments). The lines depict the regressed Gompertz sigmoids that best describes the data (continuous, photopolarization according to L1; dotted, TBO patch staining).

Figure 6

Resultant angular distribution for each of the different polarization scenarios considered. Note that 180° reorientation yields for each of these situations a diametrically bimodal distribution (see text for explanation).

Figure 7

Proposed scenario for integration of polarization vectors as vectorial sum under after reorientation of 90° (A) or 180° (B). AV, axis amplification vector; SV, axis stabilization vector. Red block arcs depict the hypothesized intrinsic factor responsible for the imprinting of unknown nature. Blue shading denotes F2 deposition as assayed by TBO staining. Hours denote duration of illumination under different light regimes and therefore the strength of each vector. Black bars and spheres denote the polar endomembrane system and F-actin deposition symbolizing the entire axis amplification machinery.