The Drosophila anterior-posterior axis is polarized by asymmetric myosin activation

Context and Background:

The formation of the main body axis is a tightly regulated process, fundamental for successful embryonic development. In Drosophila, body axis patterning is established earlier on during oogenesis, well before egg fertilization and deposition. The process requires a complex cross talk between the oocyte and the surrounding somatic epithelium, and it is guided by multiple hierarchical steps of symmetry-breaking events within the oocyte. The first polarising signal is detectable at stage 6-7 of egg development (Fig.1). At this stage, the release of the morphogen Gurken by the oocyte induces the follicle cells to commit towards a posterior fate through the activation of the EGF-like receptor Torpedo [1] (Fig.1, step1, red arrows). The posterior follicle cells induce the establishment of cortical polarity in the oocyte, sending back a signal of which the molecular nature is still unknown. (Fig.1, step1, green arrows). Under the influence of this signal, the oocyte recruits the polarity protein PAR-1 to the posterior cortex in an actin-dependent manner [2, 3] (Fig.1, step2, green crescent). Meanwhile, aPKC and Par-6 are actively excluded from the posterior cortex and recruited and maintained at the anterior (Fig.1, step 2, purple crescent). The establishment of cortical polarity is reinforced by mutual exclusion between the anterior and posterior PAR complexes. This series of events leads to the polarisation of the oocyte microtubule network (Fig.1, step 3) and the consequent asymmetric distribution of cell fate determinants (Fig.1, step 4). Additionally, the oocyte nucleus (Fig.1, step3, in orange) migrates to a more antero-dorsal position as a consequence of microtubule polarisation, setting the basis for the formation of the dorsal-ventral (D/V) axis of the future embryo.

 

Figure 1: Schematic summary of the main steps leading to the establishment of A/P and D/V polarity in Drosophila.

 

In this preprint, Doerflinger et al. investigate how the signal behind the establishment of oocyte cortical polarity is transduced at a molecular level and identify the first event of polarity establishment in the asymmetric activation of non-muscle Myosin II.

 

Main findings:

By performing a germline clone screen for mutants with disrupted A/P polarity, the authors identified a complementation group of three mutant alleles which showed clear signs of compromised anterior-posterior polarization: oocyte microtubule network failed to polarize, PAR-1 crescent at the posterior end failed to form and the oocyte nucleus at stage 10 of egg development failed to occupy the classical antero-lateral localization. The mutant alleles were mapped to the unc-45 locus which encodes for a chaperone protein known to fold and stabilize myosins in general and specifically non-muscle Myosin-II (Myo-II) in this particular context. Therefore, they explored whether activation of non-muscle Myo-II was required for establishment of polarity. They found that the di-phosphorylated active form of Myo-II was enriched in a posterior crescent, from stage 7 of egg chamber development onwards. Activation of Myo-II was dependent on the polarising signal Gurken but was upstream of Par-1 cortical recruitment. Moreover, the authors showed that di-phosphorylation of Myosin regulatory light chain (known as Spaghetti squah ‘sqh’ in flies) was required for the polarisation of the oocyte. Indeed, 50% of sqh null mutantant egg chambers overexpressing a mutant form of Sqh in which threonine 20 was mutated into alanine (sqhAS) failed to localise Par-1 at the posterior cortex and consequently, to transport Staufen to the posterior pole of the oocyte.

Were Myo-II activation and cortical contraction continuously required to maintain polarisation? Pharmacological treatment of egg chambers with ML-7, an inhibitor of myosin light chain kinase, resulted in the fast and complete loss of di-phosphorylated Myo-II and Par-1 from the posterior cortex, confirming the central role played by Myo-II phosphorylation in the establishment and maintenance of polarity.

Relevance:

This work provides the first evidence that di-phosphorylation of Myo-II is required for the establishment and maintenance of cortical polarity in the Drosophila oocyte. Preventing myosin-II activation disrupts the recruitment of Par-1 to the posterior cortex and compromises axis formation. The Myo-II dependent localisation of Par-1 in the oocyte resembles the acto-myosin dependent mechanism of localisation of Miranda during the asymmetric division of the neuroblasts [4], pointing to a more general role for Myo-II in the establishment of cellular asymmetry.

Questions to the authors:

1. How is asymmetric Myosin-II distribution achieved? Is Myosin transported along microtubules in a kinesin-dependent manner? Is the EGF signalling modulating the activity of Unc-45 as well Myosin-II activation?
2. Is there evidence of a difference lipid composition between the anterior and the posterior cortical domains? If yes, how do local changes in lipid composition affect establishment of cortical polarity?

 

References:

1. Gonzalez-Reyes, A., H. Elliott, and D. St Johnston, Polarization of both major body axes in Drosophila by gurken-torpedo signalling. Nature, 1995. 375(6533): p. 654-8.
2. Shulman, J.M., R. Benton, and D. St Johnston, The Drosophila homolog of C. elegans PAR-1 organizes the oocyte cytoskeleton and directs oskar mRNA localization to the posterior pole. Cell, 2000. 101(4): p. 377-88.
3. Doerflinger, H., et al., The role of PAR-1 in regulating the polarised microtubule cytoskeleton in the Drosophila follicular epithelium. Development, 2003. 130(17): p. 3965-75.
4. Hannaford, M.R., et al., aPKC-mediated displacement and actomyosin-mediated retention polarize Miranda in Drosophila neuroblasts. Elife, 2018. 7.

 

doi: https://doi.org/10.1242/prelights.29535

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