Jiang Group - Ascidian notochord morphogenesis
I am 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 convergence/extension (C/E), a four by ten sheet of differentiated notochord cells are organized into a single column of 40-stacked cells. Following C/E, notochord development continues as individual cells elongate and vacuolize along the anterior/posterior (A/P) axis. Finally, these vacuoles fuse to form a tube that runs the length of the larval tail.
In addition to this simple and stereotypic mode of development and morphogenesis, ascidian offers several other advantages for developmental genetic research. 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 technique has made gene manipulation and generation of transgenic animals not only feasible but easy. Fourth, large scale culture of Ciona and their being self-fertilizing hermaphrodites have made ascidians attractive animal models for classic genetic study.
My previous work includes an analysis of a natural mutant (aimless) that carries a mutation in ascidian planar cell polarity (PCP) gene prickle, and demonstration of an essential role for prickle in C/E movement. This work has lead to the discovery of an A/P polarity in individual notochord cells, evidenced by the anterior localization of prickle and strabismus (also in the PCP pathway - see Figure 1). With the use of aim mutant, I was able to demonstrate a requirement of prickle for the A/P polarity of notochord cells. These findings reveal that the PCP pathway not only operates along the medial/lateral (M/L) axis during C/E, but also regulates notochord morphogenesis along the A/P axis at the elongation phase. The simple morphogenesis of the ascidian embryos, particularly of the notochord, has allowed us to advance our understanding of the role of the PCP pathway in chordate development to a degree that has not been achieved using vertebrate model organisms.
Fig. 1: Anterior/posterior polarity in ascidian notochord cells.
My group will focus on the following areas in ascidian developmental genetics:
- Develop more efficient methods to generate mutant and transgenic lines
in Ciona intestinalis.
We will explore different transposon vectors for generating stable mutant and reporter lines for notochord and other tissues in ascidian embryos that undergo major morphogenetic transformation. Other method of mutagenesis, including forward genetic approaches, will be investigated as well. - Cell biology of ascidian notochord morphogenesis.
Elongation of the tail after notochord cell convergence is achieved by cell shape changes, cell rearrangement, extracellular matrix formation, and tubulization. Several basic cell biology events are taking place: cell growth, possibly polarized secretion of matrix, cell neighbor exchanges. Bio-mechanic forces reshape and move cells in an organized manner; cell-cell and cell-matrix adhesions are also remodeled to accommodate these processes. Confocal imaging techniques allow us to visualize these events at high resolution. I have begun to use these approaches to examine the localization of regulatory proteins, cytoskeletons, secretory pathways, and adhesion apparatus in notochord (see Figure 2), in an attempt to reveal the dynamic changes in notochord elongation. - Wnt signaling pathway in convergence/extension of ascidian notochord
The non-canonical pathway downstream of several wnt proteins has been demonstrated to be important in the establishment of PCP and chordate C/E movement. However, we don’t know the source of wnt in any system used to study PCP and C/E, and we know very little about the outcome of wnt/wnt receptor interaction with respect to the behavior of its downstream signaling proteins in C/E. What is the source of wnt and how does it regulate C/E in ascidian notochord? One of my findings from the analysis of wild type embryos is the exclusion of disheveled, a crucial protein in the PCP pathway, from the notochord/muscle boundary (see Figure 3). Ascidian wnt5, which encodes a wnt protein known to signal through the non-canonical pathway, is expressed predominantly in the muscle lateral to the notochord. Our current hypothesis is that wnt5 signals to the border notochord cells, and disheveled is consequently excluded from the notochord cell membrane at the notochord/muscle boundary. An asymmetry is therefore established in that the lateral side of border notochord cells is disheveled negative, and the medial side of border notochord cell is disheveled positive. This initial asymmetry is subsequently perpetuated throughout the entire notochord, and M/L intercalation commences. Our lab will test this hypothesis by experimentally manipulate the level of wnt signaling received by notochord cells and follow their morphogenesis.
Fig. 2: green: F-actin; red: α-tubulin
Cytoskeleton architecture of notochord cells
Fig. 3: Wnt signaling in convergence/extension of ascidian notochord
