Abstract:
Syncytial nuclei found in fungi, plant endosperm, insect embryos and muscles maintain
an autonomous 'nuclear-cytoplasmic region of influence', thereby generating
compartments within a seemingly shared cytoplasm. Molecular diffusion across
syncytial nucleo-cytoplasmic domains may be constrained by binding to different
components of the cytoarchitecture. The syncytial Drosophila blastoderm embryo
provides a paradigm to test the impact of binding to the cytoarchitecture on cytoplasmic
diffusion. We have analyzed the organization of the cytoplasm and further tested the
extent of diffusion of molecules that bind differentially to the cytoarchitecture across
nucleo-cytoplasmic domains. We find that the cytoplasm is enriched in a phase
separated manner at the cortex of the syncytial Drosophila embryo above the yolk.
Photobleaching analysis shows that molecules are more constrained above the nuclei
as compared to below the nuclei in the syncytial embryo. We generated ectopic
diffusion sources of photoactivatable-GFP (PA-GFP) or PA-GFP-Tubulin by
photoactivation in the antero-posterior axis.We found that PA-GFP-Tubulin which is
incorporated in the tubulin cytoskeleton, spreads less than PA-GFP in the anterior-
posterior direction. Using anteriorly localized ectopic probes,we have tested how
molecules of different sizes or with different cyto-architectural interactions spread in
embryo. Expression of heavier fluorescent molecules at the anterior showed a shallow
linear gradient in the antero-posterior axis as compared to RFP that spread uniformly
throughout the embryo. Expression of anteriorly PH-PLC-CFP that has an attachment to
plasma membrane phospholipids however, created an exponential gradient. We have
further tested what cyto-architectural components can restrict these gradients, using
Bicoid gradient as a functional readout. Mutants affecting the cytoarchitecture of the
syncytial embryo differentially affect these gradients and our study proposes a role of
the cytoarchitecture in differentially constraining gradients based on their molecular
properties.
