Summary The giant, single-celled organism Stentor coeruleus has a long history as a model system for studying pattern formation and regeneration in single cells. Stentor (Figure 1A,B [1,2]) is a heterotrichous ciliate distantly related to familiar ciliate models such as Tetrahymena or Paramecium. The primary distinguishing feature of Stentor is its incredible size: a single cell is 1 millimeter long. Early developmental biologists, including T.H. Morgan[3], were attracted to the system because of its regenerative abilities -- if large portions of a cell are surgically removed, the remnant reorganizes into a normal-looking but smaller cell with correct proportionality [2,3]. These biologists were also drawn to Stentor because it exhibits a rich repertoire of behaviors, including light avoidance, mechanosensitive contraction, food selection, and even the ability to habituate to touch, a simple form of learning usually seen in higher organisms [4]. While early microsurgical approaches demonstrated a startling array of regenerative and morphogenetic processes in this single-celled organism, Stentor was never developed as a molecular model system. We report the sequencing of the Stentor coeruleus macronuclear genome and reveal key features of the genome: First, we find that Stentor uses the standard genetic code, suggesting that ciliate specific genetic codes arose after Stentor branched from other ciliates. We also discover that ploidy correlates with Stentor’s cell size. Finally, in the Stentor genome, we discover the smallest spliceosomal introns reported for any species. The sequenced genome opens the door to molecular analysis of single-cell regeneration in Stentor.

Here we demonstrate that RNAi can be used in molecular studies of the giant single-celled ciliate Stentor coeruleus, revealing morphogenetic functions of Mob1 and highlighting the potential of this classical model for studies of morphogenesis and regeneration.

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Although wound-healing is often addressed at the level of whole tissues, in many cases individual cells are able to heal wounds within themselves, repairing broken cell membrane before the cellular contents leak out. The giant unicellular organism Stentor coeruleus, in which cells can be more than one millimeter in size, have been a classical model organism for studying wound healing in single cells. Stentor cells can be cut in half without loss of viability, and can even be cut and grafted together. But this high tolerance to cutting raises the question of why the cytoplasm does not simply flow out from the size of the cut. Here we present a method for cutting Stentor cells while simultaneously imaging the movement of cytoplasm in the vicinity of the cut at high spatial and temporal resolution. The key to our method is to use a "double decker" microscope configuration in which the surgery is performed under a dissecting microscope focused on a chamber that is simultaneously viewed from below at high resolution using an inverted microscope with a high NA lens. This setup allows a high level of control over the surgical procedure while still permitting high resolution tracking of cytoplasm.