Cytoskeletal reorganization and plasma membrane fusion in conjugating Tetrahymena

Abstract

The conjugation junction of Tetrahymena is the specialized site where plasma membrane fusion occurs between two cells of complementary mating types. The junction is constructed through a series of cooperative interactions and morphogenetic steps. A contact-mediated interaction between free-swimming, sexually mature and mating-competent cells of two complementary mating types induces a morphological transformation of the anterior tips. Cells then join in pairs aligned by the apposition of their modified tips. Thin sections show that the plasma membranes of the tips are separated by approximately 500 A of extracellular space, in which some strands of matrix material can be identified. The cytoplasmic face of the membrane is in contact with a junction-specific thick layer of electron-dense material. At hundreds of independent sites in this junction plasma membranes fuse in a limited manner, thereby establishing hundreds of separate membrane-ensheathed cytoplasmic channels that connect the two cells. At the same locations the thick submembrane layer is interrupted. Consequently, the junction appears to be a structure that is perforated with hundreds of pores. This study poses the question of whether the junction's submembrane layer is, or includes, a skeletal element. Cells were extracted with the non-ionic detergent Triton X-100 under conditions that yield cytoskeletal frameworks (CFs) that maintain the morphological integrity of the cells. The CFs include chromatin and also cortical structures such as microtubule bands, basal bodies, ciliary axonemes, kinetodesmal fibres and fibrillar epiplasm. CFs of conjugant pairs are also paired, indicating that the junction contains a skeletal element that is responsible for integrating the individual CFs into a higher-order complex. At the ultrastructural level the skeletal structure of the junction includes membrane lamina and a submembrane scaffold, residues of the plasma membrane and thick submembrane layer, respectively, both of which are interrupted at the pores. However, the two separate scaffolds are joined at the rims of the pores. This provides a means by which the separate CFs become integrated. On the basis of images of junctional CFs, which show interruptions of the scaffold without concomitant membrane fusion, but where laminae are pressed close together, a specific model of membrane fusion is proposed. According to this model, the submembrane skeletal scaffold regulates membrane fusion by limiting its occurrence, and the extent of its occurrence.