Interaction and Regulation of Dynein by Meiosis-specific Protein KASH5 in Chromosome Motility
Creators: name: Agrawal, Ritvija
Publisher: name: My University
Year: 2023
Subjects: subject: Meiosis
subject: Dynein activating adaptor
subject: Linker of Nucleoskeleton and Cytoskeleton
subject: homologous pairing
subject: telomeres
subject: Nucleus
subject: Molecular, Cellular and Developmental Biology
subject: Health Sciences

description: Meiosis generates haploid gametes essential for sexual reproduction. A hallmark of meiosis is homologous pairing of chromosomes during meiotic prophase I which is essential to ensure correct segregation of chromosomes and promote genetic diversity. Errors in this process result in various germline defects, miscarriages, and infertility. Pairing of homologous chromosomes during meiotic prophase I is facilitated by rapid chromosome movements via telomeres, the ends of linear chromosomes. During meiotic prophase I, telomeres tether to the inner nuclear membrane linking the chromosomes to cytoplasmic dynein motors outside the nucleus. Cytoplasmic dynein forces are transmitted across the nuclear envelope to drive chromosomal movements. Dynein tethers chromosomes to the nuclear envelope via Linker of Nucleoskeleton and Cytoskeleton (LINC), a dual-protein complex. This complex consists of SUN1 on the inner nuclear membrane and KASH5 on the outer nuclear membrane. Cytosolic dynein forms a complex with KASH5 during immunoprecipitation. KASH5 is a KASH protein that is expressed exclusively during prophase I of meiosis and is crucial for mouse fertility. While SUN1 tethers telomeres to inner nuclear membrane, KASH5 completes the linkage between dynein and the chromosomal cargo. Despite being a process essential to meiosis progression and fertility, how dynein drives meiotic chromosomes across the nuclear envelope is not well understood. For long distance transport of cargoes, dynein displays processive motility, capable of taking multiple steps along their microtubule track without detaching. However, mammalian dynein is not an active/processive motor on its own. Dynein requires binding of two regulatory proteins, dynactin and one of a class of proteins called activating adaptors, to form the activated dynein complex that moves processively on microtubules. Although the processive motility of dynein necessitates binding to an activating adaptor, the specific identity of an activating adaptor required for dynein to move meiotic chromosomes remains unidentified. My research uncovered that the meiosis-specific nuclear-envelope protein, KASH5, acts as a dynein activating adaptor. It binds to dynein using a mechanism that's conserved among activating adaptors and transforms dynein into a processive motor. Using biochemical assays, I showed that EF-hand containing N-terminal domain of KASH5 binds the light intermediate chain (LIC) of dynein through its conserved helix. My studies demonstrated that KASH5 binds LIC with a unique stoichiometry of 2:1 where a dimer of KASH5 binds one LIC. Even though KASH5 is an activating adaptor that contains an EF-hand, it does not bind to calcium and does not require calcium for interaction with LIC. Using cultured cells and mouse spermatocytes, we further demonstrated that KASH5 recruits dynein to meiotic telomeres at nuclear envelope in a LIC-dependent manner. Moreover, my studies revealed that the KASH5-LIC interaction is essential for converting dynein into a fast processive motor. Finally, my research helped identify the dynein-binding surface of KASH5, pinpointing mutations that inhibit dynein binding in vitro. These mutations also interfere with the recruitment of the dynein machinery to the nuclear envelope in cultured cells as well as mouse spermatocytes in vivo. These findings establish KASH5 as a new EF-hand pair-type activating adaptor, the first transmembrane dynein activating adaptor, and a meiosis specific dynein activator. In conclusion, my research emphasizes the significance of the KASH5-dynein interaction in linking telomeres to the cytoskeletal machinery across the nuclear envelope which drives chromosome motility in meiotic prophase I- a prerequisite for meiotic progression and mammalian fertility.