Rab8, Rab11, and Rab35 coordinate lumen and cilia formation during Zebrafish Left-Right Organizer development

Abstract

An essential process for cilia formation during epithelialization is the movement of the centrosome to dock with the cell′s nascent apical membrane. Our study examined centrosome positioning during the development of Danio rerio′s left-right organizer (Kupffer′s Vesicle, KV). We found that when KV mesenchymal-like cells transition into epithelial cells that are organizing into a rosette-like structure, KV cells move their centrosomes from random intracellular positions to the forming apical membrane in a Rab11 and Rab35 dependent manner. During this process, centrosomes construct cilia intracellularly that associated with Myo-Va while the centrosomes repositioned towards the rosette center. Once the centrosomes with associated cilia reach the rosette center, the intracellular cilia recruit Arl13b until they extend into the forming lumen. This process begins when the lumen reaches an area of approximately 300 μm2. Using optogenetic and depletion strategies, we identified that the small GTPases, Rab11 and Rab35, regulate not only cilia formation, but lumenogenesis, whereas Rab8 was primarily involved in regulating cilia length. These studies substantiate both conserved and unique roles for Rab11, Rab35, and Rab8 function in cilia formation during lumenogenesis.

Hepatoblast iterative apicobasal polarization is regulated by extracellular matrix remodeling

“…This means that the polarization process is repeated, leading to multiaxial polarity independently of cell division. The population of AREs at the initial polarization site was retained, presumably by the MTOC, as it was previously shown that the MTOC is involved in the sub-apical retention of AREs (Le Droguen, 2015 9 ; Aljiboury, 2022 60 ). The association of movement of pAREs with centrosome relocation is in line with previously reported polarization events in intestinal cells (Feldman, 2012 61 ).…”

Temporal and anteriorly positioned mitotic zones drive asymmetric microtubule patterns needed for Left-Right Organizer development

“…Recent studies of LRO formation have elucidated the fundamental role of LRO progenitors in forming the rosette-like structure with epithelial-like characteristics before the initiation of lumen formation 8,15,30 . However, the steps preceding lumen formation is not clear, especially in the context of a living embryo.…”

“…MT-bundled structures that aren’t assembled by a bonafied centrosome have been reported at the base cof C. elegans sensory cilia 42 and in the early mouse embryo 43 . In the KV, one potential possibility for these bundled MTs is that they are potentially used to help reposition the centrosome and intracellular forming cilia to the apical membrane that occurs during this time 15,44 . As KV cells progress from a mesenchymal to epithelial-like state, the centrosome transforms into a basal body responsible for cilium assembly in a subset of KV progenitor cells before rosette formation 15 .…”

“…In the KV, one potential possibility for these bundled MTs is that they are potentially used to help reposition the centrosome and intracellular forming cilia to the apical membrane that occurs during this time 15,44 . As KV cells progress from a mesenchymal to epithelial-like state, the centrosome transforms into a basal body responsible for cilium assembly in a subset of KV progenitor cells before rosette formation 15 . The centrosome has been documented to decrease its MTOC function when required to reposition where it can transport along pre-existing MTs to get to its future site of function 45,46 .…”

“…To understand how a functional LRO is formed in a living organism—from a small group of precursor cells to a fluid-filled sac lined with both motile and non-motile ciliated cells 15,16 , we studied the development of the Kupffer’s Vesicle (KV), the zebrafish LRO, at single-cell resolution using a series of novel transgenic zebrafish lines and cell biological techniques, including high-resolution live microscopy, lineage tracing, laser ablation, and pharmacological treatments. Our study shows that FGF signaling-mediated early divisions of KV precursor cells are pivotal for proper KV development as the progeny of these early division events constitute a proportion of cells in the mature KV.…”