Regulation of spermiogenesis by combover and the E3 ligase mindbomb 2

The National Institute of Child Health and Human Development awarded Albert Einstein College of Medicine $230,388 on January 1, 2026, under the Child Health and Human Development Extramural Research program (CFDA 93.865) to conduct hypothesis-driven research on the regulation of spermiogenesis by the COMBOVER protein and the E3 ubiquitin ligase mindbomb 2.

The research employs Drosophila as a model organism to understand the mechanisms by which COMBOVER, an intrinsically disordered protein, and mindbomb 2 regulate male fertility and the individualization of spermatids. The study will investigate how these proteins interact with conserved membrane raft and axonemal components to coordinate the transition between axoneme elongation and spermatid maturation, with findings applicable to understanding human male reproduction and fertility pathways.

Performance occurs in Bronx, New York, with an award period ending August 31, 2028.

Abstract

The ability of an organism to reproduce is one of the essential characteristics that ensure its survival. Any disruption of the gamete formation results in fertility issues and, in the worst cases, sterility. Due to the conservation of spermatogenesis across species, drosophila has been a valuable model for understanding reproductive biology and fertility. In mammals and drosophila, a plasma membrane tightly embeds mature spermatids upon the differentiation and individualization of syncytial spermatids. Elongated interconnected spermatids begin to separate in a process where specialized actin cones move along the sperm tails removing inter-spermatid bridges and excess cytoplasm. This individualization process is required for the maturation of individual and functional spermatids. Previously, IT was shown that CMB, an intrinsically disordered protein, regulates fertility and the individualization of spermatids. To understand the mechanisms behind CMB's regulation, we will study proteins that CMB might interact with. From our preliminary data, we know that CMB interacts with members of a conserved protein network that are expressed in human spermatids, including the membrane raft proteins flotillin 1/2 and E3 ubiquitin ligase mindbomb 2 (MIB2), as well as with the radial spoke protein 3 (RSP3), an axonemal component required for individualization. Interestingly, CMB colocalizes with spectrin, a cytoskeletal protein, in the elongation complexes (ECS) at the distal tip of the growing axonemes. Similarly, MIB2 is also found at the ECS but not simultaneously with CMB, which might indicate a spatiotemporal regulation between MIB2 and CMB. We hypothesize that MIB2 targets CMB for degradation to coordinate the transition between axoneme elongation and spermatid individualization. Here, i will determine how and at which stage MIB2 regulates spermatogenesis and male fertility using immunohistochemistry and a rescue-based structure-function analysis. To complement this, i will perform live imaging of endogenously fluorescently tagged CMB and MIB2 to address their dynamic interplay in localization. Understanding CMB and MIB2 regulation will be instrumental in advancing our knowledge of the complex dynamics during spermiogenesis, an unknown mechanism in all organisms.