Collective nuclear behavior shapes bilateral nuclear symmetry for subsequent left-right asymmetric morphogenesis in Drosophila

概要

Directional left-right (LR) asymmetry, which is evident in many animals’ external and internal morphology, is genetically determined. Recent studies show that the mechanisms determining LR-asymmetry are evolutionarily divergent. In vertebrates, several different mechanisms contribute to LR-asymmetric development, including nodal flow, LR-asymmetric proton influx, and LR-asymmetric cell migration; some of these mechanisms have parallel functions. In Lophotrochozoa and Ecdysozoa, intrinsic cell chirality plays a key role in LR-asymmetric development. For example, cell chirality in snail and nematode blastomeres determines their subsequent LR-asymmetric organ and body development. In Drosophila, the LR-asymmetrical development of several organs also relies on cell chirality, which is controlled by the myosin 1D gene. Importantly, chiral cells are also found in vertebrates and are thought to contribute to their LR-asymmetric development. However, the molecular mechanisms of LR-asymmetric development in invertebrates remain largely unclear.
Proper organ development often requires nuclei to move to a specific position within the cell. To determine how nuclear positioning affects left-right (LR) development in the Drosophila anterior midgut (AMG), I developed a surface-modeling method to measure and describe nuclear behavior at stages 13-14, captured in three-dimensional time-lapse movies.
I found that the nuclei of the visceral muscles were positioned LR-symmetrically in distinct regions along the anterior–posterior axis in wild-type embryos; I refer to this distribution as proper nuclear positioning (PNP) hereafter. The densely crowded nuclei in these regions actively rearranged their positions relative to neighboring nuclei; I refer to this as collective nuclear behavior (CNB) hereafter. Dally-like protein (dlp), a component of Wnt signaling, was essential for both PNP and CNB. MyoII and LINC complex were required for PNP but not for CNB. Unexpectedly, however, the nuclei aligned LR-asymmetrically in mutants with disrupted MyoII or LINC complex, although the AMG developed LR-symmetrically. My results show that the positioning of the nuclei in the visceral muscles is accomplished via multiple regulatory machineries, including Wnt signaling, MyoII, and LINC complex, and that the LR-symmetric positioning of the nuclei is important for the LR-asymmetric development of the AMG.
I demonstrated that the bilaterally symmetric arrangement of the nuclei in the visceral muscles of the AMG is required for this organ’s LR-asymmetric development. In the absence of MyoII or a LINC-complex component, the nuclei align LR-asymmetrically but the AMG develops LR-symmetrically. Thus, MyoII and LINC complex play important roles in the LR-symmetric rearrangement of the nuclei, which is required for or coupled with the subsequent LR-asymmetric morphogenesis.
I revealed two distinct events that control nuclear location PNP and CNB, both of which require Wnt4 signaling. However, LINC complex and MyoII are required for PNP but not for CNB, demonstrating that the two events depend on distinct underlying mechanisms.
I also found that Wnt4 signaling is required for the CNB in the visceral muscles. In the wild-type embryo, the nuclei are densely packed into a limited area in each lateral half of the ventral region of the AMG. However, when Wnt4 signaling was interrupted, as in dlp mutants, the nuclei were sparsely distributed over a larger area and migrated more actively. This observation suggests that Wnt4 signaling might organize the collective movement of the nuclei in wild-type embryos by downregulating nuclear migration.
In this analysis, I demonstrated that nuclear position is crucial in forming LR asymmetry. In wild-type embryo, nuclear position is bilateral, and the morphology of the AMG is LR-asymmetric. However, in the absence of kash and MyoII, nuclear positioning become LR-asymmetric but the AMG morphology stays bilateral. Therefore, LR-symmetric nuclear arrangement is required for the subsequent LR-asymmetric development of the AMG.
Considering that non-skeletal muscles—which are, like Drosophila visceral muscles, formed of multi-nucleated cells—contribute to LR-asymmetric organs and tissues such as the heart, blood vessels, and digestive organs in vertebrates and other organisms, the contribution of nuclear positioning to LR-asymmetric development may be evolutionarily conserved.