Abstract

Membrane Dynamics at the Nuclear Exchange Junction during Early Mating (One to Four Hours) in the Ciliate Tetrahymena thermophila Eric S. Cole a , Thomas H. Giddings Jr. b , Courtney Ozzello b , Mark Winey b , Eileen O'Toole c , Judy Orias d , Eileen Hamilton d , Sabrice Guerrier e , Anna Ballard a and Tyler Aronstein a a Biology Department, St. Olaf College, Northfield, Minnesota, USA b Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder, Colorado, USA c The Boulder Laboratory for 3D Electron Microscopy of Cells, Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder, Colorado, USA d Department of Molecular, Cellular and Developmental Biology, University of California, Santa Barbara, California, USA e Biology Department, Millsaps College, Jackson, Mississippi, USA ABSTRACT Using serial-section transmission electron microscopy and three-dimensional (3D) electron tomography, we characterized membrane dynamics that accompany the construction of a nuclear exchange junction between mating cells in the ciliate Tetrahymena thermophila . Our methods revealed a number of previously unknown features. (i) Membrane fusion is initiated by the extension of hundreds of 50-nm-diameter protrusions from the plasma membrane. These protrusions extend from both mating cells across the intercellular space to fuse with membrane of the mating partner. (ii) During this process, small membrane-bound vesicles or tubules are shed from the plasma membrane and into the extracellular space within the junction. The resultant vesicle-filled pockets within the extracellular space are referred to as junction lumens. (iii) As junction lumens fill with extracellular microvesicles and swell, the plasma membrane limiting these swellings undergoes another deformation, pinching off vesicle-filled vacuoles into the cytoplasm (reclamation). (iv) These structures (resembling multivesicular bodies) seem to associate with autophagosomes abundant near the exchange junction. We propose a model characterizing the membrane-remodeling events that establish cytoplasmic continuity between mating Tetrahymena cells. We also discuss the possible role of nonvesicular lipid transport in conditioning the exchange junction lipid environment. Finally, we raise the possibility of an intercellular signaling mechanism involving microvesicle shedding and uptake.