First person – Haoting Wang and Hiroko Kobayashi Free

Haoting Wang

How would you explain the main findings of your paper in lay terms?

H.W.: Sperm consist of a head and a tail. The head carries the genetic information (nucleus), while the tail generates movement to help the sperm reach the egg. Many proteins are present in the sperm tail to ensure its mobility. In our study, we focus on one specific protein called MYCBPAP, which is conserved from simple organisms, like flagellates, to humans. In flagellates, MYCBPAP is believed to be important for movement, but its role in mammalian sperm was previously unknown. We created a mouse model that lacks MYCBPAP and found that this protein is essential for the growth of the sperm tail. At a higher resolution, we also discovered that without MYCBPAP, the sperm tail cannot attach to the sperm nucleus. This attachment process is fascinating and mysterious because it only occurs during sperm formation. Our study expands the understanding of how the sperm tail and nucleus connect and provides insight into how the absence of MYCBPAP might affect human sperm formation.

H.K.: I think one of the most important aspects of this research is that a single protein can greatly affect the state of sperm formation. MYCBPAP localizes along the sperm tail, and the lack of this protein results in abnormal tail elongation. It also contributes significantly to normal head formation as well as the docking of the centrosome and nuclear envelope.

Were there any specific challenges associated with this project? If so, how did you overcome them?

H.W.: The backbone of a sperm's tail, known as the flagellum, is composed of a structure called the axoneme. The axoneme consists of two microtubule singlets and nine microtubule doublets. In Chlamydomonas, the ortholog of MYCBPAP associates with the two microtubule singlets, forming what is known as the central apparatus. To determine whether MYCBPAP is located in the central apparatus of mouse sperm, I was advised to use immunogold transmission electron microscopy. However, separating the central apparatus from the nearby structures called radial spokes was challenging due to their proximity. To tackle this, we broke the sperm membrane and added ATP, which caused a portion of the microtubule singlets to separate from the axoneme. We then performed immunocytochemistry. Our results indicate that MYCBPAP is localized to the central apparatus and not the surrounding microtubule doublets. This was confirmed by the absence of MYCBPAP fluorescence on the separated microtubule doublets.

H.K.: The major challenge was determining the localization, role and interacting proteins of MYCBPAP owing to the lack of any good antibodies against it. It took a very long time to generate Mycbpap–FLAG transgenic mice. However, we were happy to eventually identify the binding proteins.

When doing the research, did you have a particular result or ‘eureka’ moment that has stuck with you?

H.K.: It was interesting to identify the proteins that MYCBPAP binds to. We made various plasmids to express flagellar proteins with tags and tested the binding between MYCBPAP with these proteins using a heterologous expression system followed by immunoprecipitation. I was thrilled to observe interactions between MYCBPAP and these proteins, which were later confirmed to likely bind based on the interactome data. Although this finding was not included in the data for this paper, it was a special moment for me.

Why did you choose Journal of Cell Science for your paper?

H.W.: Reading papers from the Journal of Cell Science is always enjoyable. JCS includes many interesting and high-quality studies, including one on CFAP70, the Chlamydomonas ortholog of the MYCBPAP-interacting protein (Hou et al., 2021). I also personally appreciate the website design of JCS.

Hiroko Kobayashi

Have you had any significant mentors who have helped you beyond supervision in the lab? How was their guidance special?

H.W.: I sincerely appreciate the help from one of the corresponding authors, Haruhiko Miyata, whom we refer to as Miyata-san. Interacting with Miyata-san is always a pleasant and enjoyable experience. I especially value the trust he has placed in me, which has allowed me the freedom to conduct my experiments comfortably in the lab.

H.K.: Professor Masahito Ikawa taught me the basics of reproduction, molecular biology and genetic engineering. He always took time to answer my questions in detail. Sometimes he even participated in experiments, including operating the micromanipulator – a memory I will never forget. Additionally, a former lab member, Seiya Oura, taught me all the basics of experimental techniques. Thanks to their guidance, I am now able to advance my research.

What motivated you to pursue a career in science, and what have been the most interesting moments on the path that led you to where you are now?

H.W.: When I was a child, everyone around me dreamed of becoming either a scientist or an astronaut, as these were considered the coolest jobs among kids at the time. I was one of them and have been chasing my dream of becoming a scientist for a long time. It has been a challenging journey, but I am grateful that my current lab accept me.

H.K.: This research was my first as an undergraduate student. In the Ikawa lab, members often generate genome edited mice independently. Through these experiences, along with the phenotypic and functional analyses of the mice, I learned how interesting it is to elucidate biological phenomena using experimental techniques. The fascination of searching for unknown and unexplained phenomena has kept me engaged in the world of science.

Who are your role models in science? Why?

H.W.: My role model in science is Derek Alexander Muller, known as Veritasium on YouTube. I feel that the scientific community can sometimes be exclusive and even isolated from society, and this needs to change. In this sense, Veritasium videos have always been my favorite. Derek has a talent for explaining complex and high-quality content in simpler terms. His videos inspire me and millions of others in the next generation to pursue a path in science.

H.K.: I do not have a specific role model yet as I am just starting my career as a researcher. However, I hope to learn a lot from many researchers as I progress in my career.

What's next for you?

H.W.: I plan to finish my PhD while contributing to uncovering the mechanisms that ensure spermatogenesis. Ultimately, I aim to contribute more to science, in any shape and form in the future.

H.K.: I am currently a first-year PhD student at the Graduate School of Medicine, Kyoto University. While I am currently working on drug discovery using tumor model mice, I am also focusing on basic research in cell biology. I will do my best to contribute to the field of cell biology by making use of what I have learned.

Tell us something interesting about yourself that wouldn't be on your CV

H.W.: I have lived in Japan for 8 years, and this experience has profoundly changed my life. I want to thank the Japan Student Services Organization for providing me with the Monbukagakusho Honors Scholarship during my undergraduate studies, which made all of this possible.

H.K.: I love to travel both within Japan and abroad. Currently, I have to look after experimental mice, making it difficult to take extended trips, but I hope to travel someday.