Intercellular bridges are a conserved feature of spermatogenesis in mammalian germ cells and derive from arresting cell abscission at the final stage of cytokinesis. However, it remains to be fully understood how germ cell abscission is arrested in the presence of general cytokinesis components. The
TEX14 (testis-expressed gene 14) protein is recruited to the midbody and plays a key role in the inactivation of germ cell abscission. To gain insights into the structural organization of
TEX14 at the midbody, we have determined the crystal structures of the EABR [endosomal sorting complex required for transport (ESCRT) and ALIX-binding region] of CEP55 bound to the
TEX14 peptide (or its chimeric peptides) and performed functional characterization of the CEP55–
TEX14 interaction by multiexperiment analyses. We show that
TEX14 interacts with CEP55-EABR via its AxGPPx
3Y (Ala793, Gly795, Pro796, Pro797, and Tyr801) and PP (Pro803 and Pro804) sequences, which together form the AxGPPx
3YxPP motif.
TEX14 competitively binds to CEP55-EABR to prevent the recruitment of ALIX, which is a component of the ESCRT machinery with the AxGPPx
3Y motif. We also demonstrate that a high affinity and a low dissociation rate of
TEX14 to CEP55, and an increase in the local concentration of
TEX14, cooperatively prevent ALIX from recruiting ESCRT complexes to the midbody. The action mechanism of
TEX14 suggests a scheme of how to inactivate the abscission of abnormal cells, including cancer cells.Intercellular bridges are a distinct feature of spermatogenesis in mammalian germ cells. Although observations of intercellular bridges were reported more than 100 y ago, their molecular function is largely unknown and we have only recently begun to learn how they form at the molecular level. Interestingly, stable bridges have recently been recognized as providing a unique means of intercellular communication, because cytoplasmic molecules can pass through them (
1). The loss of germ cell intercellular bridges disrupts spermatogenesis and causes sterility (
2).The most direct method of cell-to-cell communication is to connect the separate cytosols of cells using a tunnel that allows macromolecules to pass from one cell to another. Various organisms achieve this type of direct intercellular transfer using tunneling nanotubes (
3), intercellular bridges (also called ring canals) (
1), and bacterial intercellular nanotubes (
4). Somatic ring canals have also been found to equilibrate the levels of some proteins between connected cells in invertebrates such as
Drosophila (
5). Among these mechanisms, it has been shown that intercellular bridges having channels that are 0.5–3 μm in diameter are formed by the arrest of cell abscission at the final stage of cytokinesis in the germ cells of vertebrates (
1).Whether the process of cell abscission is completed or not depends on the cell type. In the somatic cells of vertebrates, cell abscission occurs at the midbody (
6), a structure that tethers two daughter cells. The midbody protein CEP55 plays a key role in recruiting the ALIX–endosomal sorting complex required for transport (ESCRT) I complex to the midbody (
7,
8). After this event, ESCRT-III subunits, which have a membrane scission activity, are recruited (
9–
13). Alternatively, to inactivate cell abscission,
TEX14, a testis-expressed gene and germ cell-specific component, is recruited to the midbody. It is essential for intercellular bridges and fertility in male mice (
2), and has recently been identified as one of the susceptibility genes for testicular germ cell tumors (
14).In germ cells, intercellular bridges are formed throughout spermatogenesis and the arrest of cell abscission is controlled precisely by a sophisticated interplay among the proteins
TEX14, ALIX, TSG101 (expressed by tumor susceptibility gene 101;
TSG101), and CEP55. Therefore, it is important to investigate how
TEX14 safeguards intercellular bridges from the potentially damaging membrane scissor in germ cells. To understand the molecular mechanisms involved in this process, we have performed both structural and functional analyses of the CEP55–
TEX14 interaction.
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