Wnt signaling is a well-known mode of cell-to-cell communication in multicellular organisms. It involves small Wnt glycoproteins secreted by signaling cells, binding to receptor proteins in receiving cells, leading to growth, division, or differentiation.
This mode of communication plays a crucial role in normal and altered cellular development, such as cancer and wound healing. It has been a subject of research for over four decades, with open questions about the complexity of the pathway and how specific members achieve different outputs.
A team of scientists, led by Professor Antónia Monteiro from NUS, used butterfly wings as a model system to uncover some of this complexity. The findings, published in Science Advances on 26 July 2023, revealed the presence and function of various Wnt pathway members in the genome of Bicyclus anynana butterflies. They also explored how these elements interact during wing development to pattern the exquisite color patterns on the wings.
The researchers used advanced in-situ localization technologies to decipher the expression patterns of all eight Wnt glycoproteins and four membrane receptor proteins (Frizzleds). They also studied the dynamics of an intra-cellular protein (Armadillo) during wing development and tested the function of several pathway members using CRISPR-Cas9 genome editing.
Dynamic role of Wnt signaling in butterfly wing development
Dr. Tirtha Das Banerjee, a Research Fellow at NUS Department of Biological Sciences, made significant discoveries about the dynamic nature of Wnt pathway members. Using immunostaining and gene localizing technologies, the study revealed the presence of distinct and dynamic bands as well as circular blotches of multiple proteins from the Wnt pathway during wing development.
One notable finding was the behavior of the Armadillo protein. Initially, it was uniformly distributed across all cells of early wings. However, as the wings developed, it became progressively localized to specific cells, particularly in the eyespot centers and wing margin. This localization played a crucial role in color differentiation, adding to the intricate patterning of butterfly wings. The research sheds new light on the complexity and regulation of the Wnt signaling pathway during the development of butterfly wings, providing valuable insights into the fascinating process of color pattern formation.
Intriguingly, during the study conducted by Dr. Heidi Connahs, a Research Fellow at NUS Department of Biological Sciences, the gene responsible for WntA, another Wnt pathway member, was found to exhibit a distinctive expression pattern. It formed a thick band running along the center of the developing butterfly wing. To delve deeper into its function, Dr. Connahs knocked out this gene, resulting in noticeable disruptions not only in the color but also in the width of the central band pattern. This striking observation demonstrated the dual role of WntA in both color and width regulation during wing development.
Further investigations by Dr. Suriya Murugesan, another Research Fellow at NUS Department of Biological Sciences, focused on Frizzled4, a member of the Frizzled gene family. The study revealed that Frizzled4 also serves a dual purpose in butterfly wing development. Firstly, it plays a vital role in the differentiation of the eyespot centers, contributing to the formation of distinct patterns. Secondly, Frizzled4 is involved in the orientation of scale cells within the eyespot domain. Interestingly, this gene family, known as Frizzleds, shares a similar role in the orientation of bristles in flies and hairs in mammals, underlining the conserved nature of these mechanisms across different species. These findings provide valuable insights into the multifaceted functions of specific Wnt pathway members in butterfly wing patterning, advancing our understanding of this intricate process.
Interaction between Wnt signaling pathways defines patterns on butterfly wings
The study's crucial revelation lies in the intricate spatial and temporal regulation of various Wnt signaling pathways that shape the wings of butterflies. The cells expressing different Wnt pathway members seem to interact and control each other's expression, creating specific signals for pattern formation.
One fascinating example is the protein frizzled2, which exhibits a complex pattern of expression. Cells without this receptor are found in the central (wntA), eyespot (frizzled4), and wing margin (frizzled9) domains. Together, these domains cover the entire wing cell field of the butterfly.
According to Professor Monteiro, this implies that Wnt signaling is active throughout the wing, but distinct pathway members are responsible for controlling various colors and patterns in different regions.
Dr. Banerjee emphasized that there is much more to explore regarding Wnt signaling in the butterfly system. Understanding how this fundamental pathway, present in all multicellular organisms, has evolved into such a sophisticated communication system over millions of years of evolution remains a compelling endeavor.
The implications of these studies extend beyond just understanding color patterns in insects. As Wnt signaling is conserved in various animals, the research could offer profound insights into other species' complex cell-cell communication systems. This journey of exploration holds the potential to shed light on the diverse and evolved nature of this fundamental signaling pathway, enriching our comprehension of biological diversity and evolution.
Source: National University of Singapore