Raw_data_supplementary_Regeneration in calcareous sponge relies on ‘purse-string’ mechanism and the rearrangements of actin cytoskeleton

Raw images from manuscript's supplemenraty files. Fixation method (if used) is stated as well as applied immunostaining.

The crucial step in any regeneration process is epithelization, i.e. the restoration of epithelium structural and functional integrity. Epithelialization 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: formation of a temporary structure, 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 depend 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 microvilli 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 are based 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.

原始图像来自手稿的补充文件。若使用了固定方法，则亦将予以说明，并描述了所应用的免疫染色技术。任何再生过程中的关键步骤均为上皮化，即上皮结构的恢复及功能完整性的重建。上皮化过程需要细胞骨架的重排，主要是肌动蛋白丝和微管。海绵动物门（Porifera）是早期分支的脊索动物，具有显著的再生能力。钙质海绵在其再生过程中具有独特的步骤：形成临时结构，即再生膜，最初覆盖伤口。此结构因外皮层和腔上表皮样层的形态发生重排而形成。本研究通过时间推移成像、免疫细胞化学和共聚焦激光扫描显微镜，定量评估了在泡状钙质海绵 Leucosolenia variabilis 中再生膜形成过程中的形态学变化，并描述了其下肌动蛋白细胞骨架的重排。再生膜的形成具有非线性的随机动力学，存在诸多波动。再生膜前沿的针状细胞形成收缩的肌动蛋白肌丝缆。再生膜的形成依赖于其收缩或通过其进行协调。细胞形态在再生膜形成过程中发生显著变化。外针状细胞变平，面积增大，而圆形度降低。海绵细胞发生转分化为内针状细胞，失去微绒毛颈和鞭毛。它们的面积增大，圆形度降低。随后的内针状细胞重新分化为海绵细胞，伴随着细胞形态的相反变化。所有这些转变均基于类似于两侧对称动物的特征性肌动蛋白丝重排。总之，我们在此提供了关于钙质海绵在修复上皮形态发生过程中细胞转变的定性和定量描述。