Shizue Ohsawa

Imaginal discs: a unique evolutionary innovation

Leg development in Drosophila

Fig 1. Leg development in Drosophila

The life cycle of Drosophila includes four stages: embryo, larva, pupa, and adult (or “imago”). Development and growth does not simply occur by increasing the size of existing structures: instead it involves a dramatic series of transformations that will ultimately give rise to the adult fly. During larval stages, special populations of epithelial cells, known as imaginal discs are set aside as sac-like patches that are the precursors of adult structures such as antennae, eyes, wings, halteres or legs. Prof. Ohsawa and her team are focusing on the leg imaginal disc, using live imaging, ex vivo organ culturing and state of the art molecular and genetic techniques.

In response to a pulse of the molting hormone ecdysone, each leg disc everts and elongates - the central portion of the disc becoming the distal part of the appendage - and through a series of tightly orchestrated events transforms itself into an adult leg. Surprisingly this process occurs in the absence of cell proliferation. One could say that at the end of the pupal stage, the adult leg is already “hidden” in the imaginal disc, waiting to be unfolded - like a reverse origami - at the appropriate developmental moment. Through studying the mechanisms that govern the transformation of a leg disc into an adult leg, Prof. Ohsawa is trying to answer universal questions concerning the folding and unfolding of an epithelial tissue in 3-dimensional space.

Growing a leg in a Petri dish

Relying upon previous publications, Prof. Ohsawa and her co-workers could demonstrate that the serine protease trypsin can promote the eversion of leg discs (upon removal of the peripodial membrane) in an ex vivo culture. Now why should a protein hydrolyzing molecule be necessary for the correct spatio-temporal unfolding of an epithelial tissue? In trying to answer that question - convinced it could possibly lead to an understanding of the mechanism involved - a search was launched for trypsin target proteins, which could be causally linked to leg disc elongation.

Fly leg and beetle horn: a universal folding principle?

A new chapter in unraveling the mechanism of 3D tissue (un) folding has recently been written, when it became clear that leg development in the fly shows remarkable similarities to horn development in beetles:

* It has recently been demonstrated that the folded structure of a beetle horn imaginal disc closely resembles that of a fly wing disc, manipulated and programmed to overexpress Yorkie (engrailed Gal4; UAS-Yki). The transcriptional activator Yorkie has previously been characterized as the major downstream effector of the Hippo signaling pathway: an evolutionary conserved gene network that controls organ size in animals.
* Moreover, mutations in some key components of the Hippo pathway exhibit leg abnormalities in the fly. For example, disrupting the function of genes, encoding:

A fly imaginal disc over-expressing Yokie resembles a beetle horn imaginal disc

Fig 2. A fly imaginal disc over-expressing Yokie resembles a beetle horn imaginal disc

- Fat: an integral membrane protein of the cadherin family,
- Dachs: an unconventional myosin,
- Four-jointed: a kinase that phosphorylates a subset of cadherin domains, leads to shorter and thicker legs in the adult fly.
Surprisingly, when some of the corresponding genes in the beetle are knocked-down by RNAi interference, similar phenotypes are observed: shorter and thicker horns.

As such, these data strongly suggest that an evolutionary conserved mechanism controls the complex repertoire of transcriptional activity, cell movements and cytoskeletal changes that are associated with a 3D folding program. Recently, a collaborative research with Prof. Shigeru Kondo (Osaka University) has been initiated in an attempt to understand the role of the key players in both the fly and the beetle.

A look into the Future
Science Fiction
Science or Fiction?
Prof. Ohsawa, in which directions are your research projects developing?
Prof. Ohsawa:
One of our research programs deals with answering questions, like “What is the relationship between folding of an epithelial tissue and the 3D structure that eventually emerges?” One project deals with understanding how an adult fly leg originates, so to speak, from the unfolding of an imaginal disc. We want to understand the 3D folding and unfolding rules and characterize the key players in this process. Presently we are focusing on the role of Type IV collagen.
Any experiments you want to perform in the near future?
Prof. Ohsawa:
We plan to knock down Type IV collagen expression in the leg disc, using RNAi interference. As such, we hope to find an interesting phenotype, which would allow us to establish a causal link between downregulation of a gene and leg disc elongation defects in vivo. That would identify Type IV collagen as a possible determinant of imaginal disc folding in Drosophila.
Apart from characterizing molecular key players in imaginal disc folding, are you searching for other types of determinants?
Prof. Ohsawa:
Definitely. I expect multiple factors to be involved in imaginal disc folding and unfolding, both in the fly and in beetles: on the one hand, cellular and molecular key players - gene networks and signaling pathways controlling cellular dynamics, such as cytoskeletal changes - and on the other hand, biomechanical determinants. Recently, we have started collaborations with Prof. Yasuhiro Inoue and Prof. Masakazu Akiyama, who are developing mathematical models that simulate the unfolding of an imaginal disc in 3D space. In this way, we hope to identify and characterize mechanical forces that guide this process.
How would you describe your scientific dreams?
Prof. Ohsawa:
Our research work ultimately aims at understanding two fundamental biological processes. First, during morphogenesis, animals are often affected by various damaging stimuli, such as environmental stress or genetic damage. Nevertheless, in most cases, a normal adult organism comes into existence, meaning that developmental circuits must be buffered somehow. I want to understand the mechanisms that secure such robust development and protect it against noise. Second, I want to understand how the 3D shape of a biological body is created - and maybe I should add: despite all these developmental and genetic biases that threaten it.
Do you believe that the mechanisms controlling biological shape are universal?
Prof. Ohsawa:
I can certainly imagine that behind the different 3D patterning mechanisms we observe, a few basic principles are hidden. Slight deviations from a basic developmental strategy - changing the biological and/or the physical parameter settings, so to speak - could give rise to entirely different 3D shapes and could explain, for example, the amazing diversity among beetle horns. So, I believe there is indeed a common logic behind the 3D shape of a biological body, waiting to be discovered.
  1. #Nakamura M, #Ohsawa S (# equal contribution), *Igaki T. Mitochondrial defects trigger proliferation of neighboring cells via senescence-associated secretory phenotype in Drosophila. Nat Commun 5, 5264, 2014.
  2. #Ohsawa S, Sato Y,Enomoto M, Nakamura M, Betsumiya A, *Igaki T. Mitochondrial defect drives non-autonomous tumor progression through Hippo signalling in Drosophila. Nature 490, 547-551, 2012
  3. #Ohsawa S, Sugimura K, Takino K, Xu T, Miyawaki A, *Igaki T. Elimination of oncogenic neighbors by JNK-mediated engulfment in Drosophila. Dev Cell 20, 315-328, 2011.
Who is Who?

Prof. Ohsawa

Prof. Shizue Ohsawa

Prof. Shizue Ohsawa was born and raised near Tokyo and studied Chemistry and Biology at Tohoku University. Thereafter, she did her doctoral work on the non-apoptotic role of caspases during mouse neural development, under supervision of Prof. Masayuki Miura at the Department of Genetics, Graduate School of Pharmaceutical Sciences, University of Tokyo. After her PhD thesis, Prof. Ohsawa moved to Kobe University Graduate School of Medicine, where she studied the mechanisms of cell competition in the fruit fly. At present she is working at the Graduate School of Biostudies at Kyoto University (Laboratory of Genetics) under supervision of Prof. Tatsushi Igaki. Her topics of interest are 3D morphogenesis and epithelial cell homeostasis through cell-cell communication, using Drosophila as a model system.