During Drosophila oogenesis, the organization of microtubules (MTs) drives polarized transport and cytoplasmic flows. Although Kinesin-1 plays a central role in these processes, its molecular and spatial regulation remains poorly understood. Here, we show that Ensconsin, an activator of the Kinesin heavy chain (Khc), is enriched in the oocyte through MT binding, Dynein-mediated transport and by the MT-anchoring protein Ninein. Strikingly, loss of Ensconsin enrichment reduces cortical non-centrosomal microtubule-organizing centers (ncMTOCs), weakens MT nucleation, impairs MT organization and MT-dependent cytoplasmic flows. Additionally, a specific Khc variant unable to bind Ensconsin, is constitutively active and is able to partly rescue the loss of Ensconsin in oocyte and neural stem cells, thereby providing in vivo support for the relief-of-auto-inhibition model. Overall, we uncover a novel function for the Khc–Ensconsin complex in the recruitment of ncMTOCs in oocytes for MT organization. Our findings are in line with a model in which Ensconsin enrichment acts as a rheostat to modulate Khc activation, ensuring precise spatial remodeling of the microtubule cytoskeleton in the oocyte.
Despite extensive research on the olfactory epithelium during embryonic and postnatal stages, little is known about peripartum adaptations. Here we describe the profound changes that occur between E18 and P8 - shortly after birth and exposure to air. In the lamina propria, we observed a significant expansion of extracellular space and axon bundles. Olfactory ensheathing cells, the glia cells of the lamina propria, stopped proliferating and upregulate SOX10 after birth. In addition, only after birth did olfactory ensheathing cells express CLDN5 and PECAM1, both of which tighten blood vessels, as well as gap junction communication and potassium transport proteins. Functional Bowman glands emerged only after birth, similar as the layer of horizontal basal cells. Since horizontal basal cells produce extracellular matrix proteins, laminin localization changed after these cells populated the basal epithelium. Interestingly, embryonic mice showed multiple invaginations of blood vessels throughout the olfactory epithelium, which disappear immediately after birth. Our findings reveal that the lamina propria of the olfactory mucosa undergoes profound postnatal maturation, leading to the establishment of a specialized adult tissue architecture.
ABSTRACT JRAB (also known as MICAL-L2), an effector protein for the small G proteins Rab8A and Rab13, interacts with actin filaments (F-actin), actinin-1, actinin-4 and filaminA, and plays an important role in the spatiotemporal actin cytoskeletal reorganization underlying cell migration and cell–cell adhesion. JRAB forms droplets that include Rab8A by liquid–liquid phase separation, leading to deformation of the endosomal membrane. In this study, we showed that JRAB functions in actin dynamics as a droplet, leading to the development of cell–cell adhesions. In vitro reconstitution assays revealed that JRAB droplets promoted the assembly of thick F-actin bundles, a process that was distinct from that observed with JRAB in its soluble form. Moreover, we identified Rab13 as a modulator of JRAB droplet function. JRAB droplets containing Rab13 inhibited the formation of F-actin bundles. The difference in mechanical properties among those droplets might be correlated with the ability of JRAB to regulate F-actin bundle morphology. Taken together, our findings provide new insight into a previously unrecognized mechanism through which droplets regulate actin dynamics by changing their components.
Epithelial cells primarily segregate transmembrane proteins to apical or basal surfaces, establishing apical-basal polarity. In 3D tissues, apical proteins face inwards. Recent work by our group has shown that increased RhoA activation causes epithelial spheroids to invert apical-basal polarity via a collective rearrangement of cells, a process we and others have termed eversion. In this work, we determined that alpha 6(34-integrin-laminin interactions are required for spheroids to evert to apical-out polarity. Additionally, we show that increased cell proliferation and anchorage independence are required to sustain apical-out polarity. We also observed that apical-out spheroids can 'revert' to apical-in polarity through apoptotic cavitation of cells located in the center of the spheroids. This study provides new mechanistic insights into the biochemical and biophysical mechanisms that drive eversion and maintain apical-out polarity, and supports the concept that apical-basal polarity orientation might drive phenotypic switching of epithelia.
