High mobility of bicoid captured by fluorescence correlation spectroscopy: implication for the rapid establishment of its gradient

Abstract

The Bicoid (Bcd) morphogen is essential for pattern formation in fruit flies. It forms an exponential concentration gradient along the embryo AP axis and turns on cascades of target genes in distinct anterior domains. The most commonly accepted model for gradient formation assumes that Bcd travels by simple diffusion and is uniformly degraded across syncytial embryos, yet several recent studies have challenged these ideas. Here, the question of Bcd mobility was investigated using fluorescence correlation spectroscopy in live Drosophila melanogaster embryos. Bcd-EGFP molecules were found to be highly mobile in the cytoplasm during cycles 12-14, with a diffusion coefficient approximately 7 microm(2)/s. This value is large enough to explain the stable establishment of the Bcd gradient simply by diffusion before cycle 8, i.e., before the onset of zygotic transcription.

Figure 1

(A) Bcd-EGFP fluorescence during nuclear cycle 13 (NC 13, lower panel) and the ensuing mitosis (M 13, upper panel) (average of three confocal images taken 2 μm apart). (B) Bcd-EGFP fluorescence gradient for the embryo shown in panel A. (Inset) Detail of a single confocal image of the embryo in both GFP and RFP channels, with arrows pointing at in-focus nuclei. (C) Amplitude and (D) decay length of the Bcd-EGFP gradient in interphase nuclei (NC 10–14) and mitotic cytoplasm (M 10–13) (mean ± SD, n = 5).

Figure 2

Normalized average autocorrelation functions obtained for Bcd-EGFP and NLS-EGFP in the anterior cortical cytoplasm of stage 12–14 embryos (continuous colored lines, fit assuming two independent diffusing species). Error bars represent the standard error. (Dashed blue lines) Expected autocorrelation functions for one diffusing species and different values of D.