Regeneration in calcareous sponge relies on 'purse-string' mechanism and the rearrangements of actin cytoskeleton.

The crucial step in any regeneration process is epithelization, i.e. the restoration of an epithelium structural and functional integrity. Epithelization requires cytoskeletal rearrangements, primarily of actin filaments and microtubules. Sponges (phylum Porifera) are early branching metazoans with pronounced regenerative abilities. Calcareous sponges have a unique step during regeneration: the formation of a temporary structure, called regenerative membrane which initially covers a wound. It forms due to the morphallactic rearrangements of exopinaco- and choanoderm epithelial-like layers. The current study quantitatively evaluates morphological changes and characterises underlying actin cytoskeleton rearrangements during regenerative membrane formation in asconoid calcareous sponge Leucosolenia variabilis through a combination of time-lapse imaging, immunocytochemistry, and confocal laser scanning microscopy. Regenerative membrane formation has non-linear stochastic dynamics with numerous fluctuations. The pinacocytes at the leading edge of regenerative membrane form a contractile actomyosin cable. Regenerative membrane formation either depends on its contraction or being coordinated through it. The cell morphology changes significantly during regenerative membrane formation. Exopinacocytes flatten, their area increases, while circularity decreases. Choanocytes transdifferentiate into endopinacocytes, losing microvillar collar and flagellum. Their area increases and circularity decreases. Subsequent redifferentiation of endopinacocytes into choanocytes is accompanied by inverse changes in cell morphology. All transformations rely on actin filament rearrangements similar to those characteristic of bilaterian animals. Altogether, we provide here a qualitative and quantitative description of cell transformations during reparative epithelial morphogenesis in a calcareous sponge.