Jiang Group - Ascidian notochord morphogenesis
(May 2006 - December 2014)

Jiang Group June 2007

We are interested in how differentiated cells are organized into complex tissues and organs, and how this process evolves. The ascidian notochord provides an elegant system for studying this process. In contrast to the vertebrate notochord, which consists of thousands of still dividing cells whose positions and movements during development are difficult to trace, ascidian notochord has only 40 cells, which are post-mitotic at the beginning of morphogenesis. During a process called convergent extension, a four by ten sheet of differentiated notochord cells are organized into a single column of 40-stacked cells. Following convergent extension, notochord development continues as individual cells elongate and produce extracellular matrix along the anterior/posterior axis. Finally, these extracellular lumens fuse to form a tube that runs the length of the larval tail. Ascidian notochord at the swimming larval stage functions as a hydrostatic “skeleton” essential for the locomotion of the tadpoles.

In addition to this simple and stereotypic mode of development and morphogenesis in notochord, ascidian offers several other advantages for developmental genetic and cell biology studies. First, ascidians belong to proto-chordates that have diverged from vertebrate lineage before whole genome duplications; therefore they reveal a minimal gene repertoire for building a chordate. Second, genomes of both Ciona intestinalis and Ciona savignyi, two commonly used species in the laboratory, have been sequenced, providing a platform for genomic and genetic studies. Third, electroporation provides an efficient method for routine gene manipulation and generation of transgenic animals. Fourth, large scale culture of Ciona and their being self-fertilizing hermaphrodites makes ascidians attractive animal models for classic genetic study. Finally, fast and external embryonic development and transparent embryos allows sub-cellular level imaging and experimental manipulation of specific cell biology processes.

We have used molecular markers and confocal imaging to describe tubulogenesis in the developing Ciona notochord. During tubulogenesis each notochord cell establishes de novo apical domains, and undergoes a mesenchymal–epithelial transition to become an unusual epithelial cell with two opposing apical domains. Concomitantly, extracellular luminal matrix is produced and deposited between notochord cells (Fig. 1 and Fig. 2). Subsequently, each notochord cell simultaneously executes two types of crawling movements bidirectionally along the anterior/posterior axis on the inner surface of notochordal sheath. Lamellipodia-like protrusions results in cell lengthening along the anterior/posterior axis, while the retraction of trailing edges of the same cell leads to the merging of the two apical domains (Fig. 3). As a result, the notochord cells acquire endothelial-like shape and forms the wall of the central lumen. Inhibition of actin polymerization prevents the cell movement and tube formation. Ciona notochord tube formation utilizes an assortment of common and fundamental cellular processes including cell shape change, apical membrane biogenesis, cell/cell adhesion remodeling, dynamic cell crawling, and lumen matrix secretion.

Our group now uses Ciona notochord tubulogenesis as experimental model to address the molecular mechanisms of tube formation. We perform functionally analyses of various cell biology processes and aim to provide mechanistic insights as how each contributes to, and how their actions are coordinated during, the construction of the tube. Specific questions and projects include:

  1. Polarity establishment; interaction between apical/basal polarity and planar cell polarity pathways, and functional significance of cell polarities in organizing cell shape changes, cell movement, and metabolic activities of notochord cells during tube formation.


  2. Secretory pathways (nature of secretory vesicles and their directional trafficking) for extracellular matrix production and apical/luminal membrane biogenesis.


  3. Molecular mechanisms for dynamic remodeling of cell adhesion complexes, including adherens junctions and tight junctions, during the rearrangement of notochord cells (Fig. 4) as they form a tube


  4. Molecular and mechanistic basis of the force generation underlying cell shape changes and cell movements





Fig. 1: 3D reconstruction of single notochord cell as it undergoes dramatic cell shape
changes and produces large amounts of extracellular lumen at both anterior and posterior ends.
Click image to see video.




Fig. 2: Small ion transporter SLC26A2 staining (red) of notochord cells reveals apical/luminal membrane
biogenesis concurrent with extracellular matrix production. Notochord cells are counter-stained green for F-actin.
Click image to see video.




Fig. 3: Complex bi-directional movements of individual notochord cells as they take on an endothelial cell shape.
Mosaic expression of actin-GFP reveals the motile cell edges.
Click image to see video.




Fig. 4: Illustration of cell junction rearrangement at the later stage of notochord tube formation.
Directional cell movements (green arrows) result into the loss of cell-cell junctions between orange and
purple cells, and between orange and green cells; and new cell-cell junction between purple and green cells is established.
Click image to see video.

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