Hox genes play a key role in determining body plan, but previous research indicated that forewing development occurs independently of Antennapedia, the Hox gene expressed in the thoracic region. Now, a new paper in Development describes an essential role for Antennapedia in wing development of silkworm, Drosophila and Tribolium. We caught up with first author, Chunyan Fang, and corresponding author, Xiaoling Tong, a group leader at the State Key Laboratory of Silkworm Genome Biology at Southwest University in China, to find out more about their research.
Xiaoling Tong (L) and Chunyan Fang (R)
Xiaoling, can you give us your scientific biography and the questions your lab is trying to answer?
XT: I studied genetics and biology and obtained my PhD from Southwest University in China. During my PhD, I focused on the wing development of silkworm. As a postdoctoral fellow, I joined the lab of Professor Antónia Monteiro at Yale University, where I tested the function of the Hox gene Ultrabithorax in regulating eyespot patterns in the butterfly Bicyclus anynana. While doing this research, I became interested in how Hox proteins control morphogenesis and organogenesis, particularly in light of their roles in the evolution of morphology.
In 2012, I moved back to the State Key Laboratory of Silkworm Genome Biology at Southwest University in China. My team primarily focuses on genetics and developmental biology using the silkworm as a model. We are working on how Hox proteins cause morphological diversity at evolutionary levels and dissecting the roles and regulatory mechanisms of the non-coding RNAs within the Hox gene clusters in development and evolution. We also use mutant screens to identify genes and regulatory sequence variations that generate large phenotypic changes to help us understand the genetic basis of phenotype diversity.
Chunyan, how did you come to work in Xiaoling's lab and what drives your research today?
CF: I first met Professor Tong at dinner after my junior internship. Professor Tong's gentle and amiable character moved me very much. More than eight years have passed, but the scene from that time is still vivid. Therefore, Professor Tong's personal charm encouraged me to choose her lab for graduate study and research. After joining the lab, I learned that Professor Tong is a very rigorous scholar in academic research, and I firmly believe I chose a good lab.
I was so lucky to have the chance to take part in this project. Before I took over this project, a senior colleague had conducted some preliminary research and found that the defective appendages (both legs and wings) in the silkworm mutant Wes were caused by a mutation in the Antp gene. Professor Tong told me that we would test the function of Antp in wing development and challenge a long-standing view that insect forewings develop without any Hox input. The interesting project and my own curiosity drive my research.
Can you give us the key results of the paper in a paragraph?
CF & XT: The key result of the paper is that Antp is an essential gene for wing development in Bombyx, Drosophila and Tribolium, and probably across all insects. We show that Antp is expressed both in forewings and in hindwings of Bombyx mori, primarily during the pupal stage. Downregulation of Antp expression by RNA interference or CRISPR/Cas9 resulted in small and malformed adult wings, indicating that Antp plays an essential role in wing growth and development. Furthermore, we found that Antp controls the expression of shade, an enzyme that converts ecdysone to active 20E directly in wing tissues. Antp also regulates the expression of cuticular protein genes (CPH28, CPG24 and CPG9) in both forewings and hindwings. Knockdown of Antp in Tribolium and Drosophila wings during both larval and pupal stages led to crinkled and reduced adult wings, which were previously undescribed phenotypes. We propose that insect wings develop in an Antp-dependent manner, unlike previous reports which suggested that forewing development was independent of Hox genes.
The key result of the paper is that Antp is an essential gene for wing development in Bombyx, Drosophila and Tribolium, and probably across all insects.
What was your first indication that Antennapedia plays a role in wing development, contrary to previously published research?
XT: During my postdoctoral research at Yale University, I found that Antp is expressed in the eyespots and blade of pupal wing by antibody staining. After I moved back to China, I kept working on Hox genes. We found that the heterozygotes of a loss-of-function mutation in the Antp gene produced abnormal wings in silkworm. These results made me firmly believe that the Antp is required for wing development.
When doing the research, did you have any particular result or eureka moment that has stuck with you?
CF: Every experimental result excited me! Of course, the most unforgettable result was when we observed the defect in the adult wing type after downregulating the expression of Antp in Bombyx mori, Drosophila and Tribolium castaneum. It was a heart-stirring moment when I got these results. I couldn't believe it was true.
And what about the flipside: any moments of frustration or despair?
CF: I still remember the molecular cloning of the promoters used in this study, which was about four years ago. I tried many ways to get the full sequences but still failed. These results made me very depressed. I burst into tears alone in my dorm room to release all my pent-up emotions. Frustrating moments still occurred in the later experiments, but this experience was the most memorable one for me. Perhaps it is because of the failure of the experiment for the first time. But, overall, the experience was very rewarding.
How do the genetic tools for the silkworm (Bombyx mori), compare to more traditional model organisms?
CF & XT: In silkworm, we have similar tools to Drosophila, including RNA interference (by injecting dsRNA or siRNA), transgenic techniques using piggyBac transposon-derived vectors, the GAL4/UAS system, site-specific recombination using Cre or FLP recombinases, and genome editing using TALENs or CRISPR-based systems.
Chunyan, what's next for you after this paper?
CF: The research in this paper formed the main part of my PhD studies in Xiaoling's lab. I graduated in July this year and left Xiaoling's lab. At present, I study the development of the male reproductive system at Chengdu University of Traditional Chinese Medicine. I hope to have the opportunity to publish my research findings in Development in the future.
Where will this story take the Tong lab?
XT: I will keep working on the same research line. I will test the role of Hox proteins in determining appendage diversification in silkworm and other insect species during evolution.
Finally, let's move outside the lab – what do you like to do in your spare time?
CF: I enjoy yoga, badminton, food and nature, which make me calm and help me think more clearly.
XT: I love gardening; I like to spend my spare time with my plants.
C.F. & X.T.: State Key Laboratory of Silkworm Genome Biology, Key Laboratory of Sericultural Biology and Genetic Breeding, Ministry of Agriculture and Rural Affairs, Southwest University, Chongqing 400715, China.
C.F.: TCM Regulating Metabolic Diseases Key Laboratory of Sichuan Province, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu 610072, China.
Email: [email protected]
