A molecular corset for spermatids: Insights into the regulation and function of the manchette during spermiogenesis

Sperm cells are highly specialized and polarized cells that deliver the paternal genome to the female oocyte. Spermiogenesis is the crucial process of sperm shaping. It involves acrosome formation, chromatin condensation, nuclear shaping, tail formation, and cytoplasm reduction; failures during these processes inevitably lead to morphology defects and infertility. During spermiogenesis, a temporal organelle called the manchette assembles and facilitates the high polarization of round spermatids to sperm cells. The manchette primarily comprises microtubules, highly dynamic, hollow biopolymers involved in many fundamental cellular processes, including cell division and cellular motility. The manchette is relevant for our understanding of microtubule regulation and stabilization, as it maintains its structural integrity for several weeks and is used as a scaffold for protein cargo. The work presented here provides fundamental new insights into the molecular mechanisms involved in manchette stabilization and function. Chapter I provides fundamentals of sperm cell development and the molecular role of the manchette during spermiogenesis. Chapter II overviews the current state-of-the-art of cryo-ET and its applications in cell biology. Chapter III explores the stabilization of manchette microtubules, showcasing the involvement of multiple molecular players that work together to maintain the manchette integrity over a long period. The study unveils a protein (mMIP) regularly binding to the inside of the microtubule. Its binding site suggests that it is involved in the stabilization of the seam of microtubules. The seam is the most unstable part of the microtubule, and its stabilization via the mMIP likely contributes to the prolonged integrity of the manchette. Chapter IV focuses on the utilization of the manchette as a bidirectional transport scaffold and its involvement in the polarization of the spermatid. Two diverging mechanisms of cellular cargo transport are revealed. Soluble proteins are transported along the manchette microtubules via dyneins, while vesicles are transported via other transport machinery. The study further revealed actin filaments as a second biopolymer integral to the manchette. Actin filaments appear as bundles and singlets, indicating two different roles in the manchette. Chapter V presents a workflow that allows the investigation of whole spermatids with cryo-EM. The workflow enables visualization of multiple spermiogenesis-specific structures like the manchette and the acroplaxome within spermatids. Finally, Chapter VI discusses the results in the context of published data and provides a scientific outlook.