Mitochondrial morphology dynamics and ROS regulate apical polarity and differentiation in Drosophila follicle cells.

Mitochondrial morphology dynamics regulate signaling pathways during epithelial cell formation and differentiation. The mitochondrial fission protein Drp1 affects the appropriate activation of EGFR and Notch signaling-driven differentiation of posterior follicle cells in Drosophila oogenesis. The mechanisms by which Drp1 regulates epithelial polarity during differentiation are not known. In this study, we show that Drp1-depleted follicle cells are constricted in early stages and present in multiple layers at later stages with decreased levels of apical polarity protein aPKC. These defects are suppressed by additional depletion of mitochondrial fusion protein Opa1. Opa1 depletion leads to mitochondrial fragmentation and increased reactive oxygen species (ROS) in follicle cells. We find that increasing ROS by depleting the ROS scavengers, mitochondrial SOD2 and catalase also leads to mitochondrial fragmentation. Further, the loss of Opa1, SOD2 and catalase partially restores the defects in epithelial polarity and aPKC, along with EGFR and Notch signaling in Drp1-depleted follicle cells. Our results show a crucial interaction between mitochondrial morphology, ROS generation and epithelial cell polarity formation during the differentiation of follicle epithelial cells in Drosophila oogenesis.

A new mechanism of fibronectin fibril assembly revealed by live imaging and super-resolution microscopy.

Fibronectin (Fn1) fibrils have long been viewed as continuous fibers composed of extended, periodically aligned Fn1 molecules. However, our live-imaging and single-molecule localization microscopy data are inconsistent with this traditional view and show that Fn1 fibrils are composed of roughly spherical nanodomains containing six to eleven Fn1 dimers. As they move toward the cell center, Fn1 nanodomains become organized into linear arrays, in which nanodomains are spaced with an average periodicity of 105±17 nm. Periodical Fn1 nanodomain arrays can be visualized between cells in culture and within tissues; they are resistant to deoxycholate treatment and retain nanodomain periodicity in the absence of cells. The nanodomain periodicity in fibrils remained constant when probed with antibodies recognizing distinct Fn1 epitopes or combinations of antibodies recognizing epitopes spanning the length of Fn1. Treatment with FUD, a peptide that binds the Fn1 N-terminus and disrupts Fn1 fibrillogenesis, blocked the organization of Fn1 nanodomains into periodical arrays. These studies establish a new paradigm of Fn1 fibrillogenesis. This article has an associated First Person interview with the first author of the paper.

ERK regulates mitochondrial membrane potential in fission deficient Drosophila follicle cells during differentiation.

Mitochondrial morphology regulatory proteins interact with signaling pathways involved in differentiation. In Drosophila oogenesis, EGFR signaling regulates mitochondrial fragmentation in posterior follicle cells (PFCs). EGFR driven oocyte patterning and Notch signaling mediated differentiation are abrogated when PFCs are deficient for the mitochondrial fission protein Drp1. It is not known whether fused mitochondrial morphology in drp1 mutant PFCs exerts its effects on these signaling pathways through a change in mitochondrial electron transport chain (ETC) activity. In this study we show that aggregated mitochondria in drp1 mutant PFCs have increased mitochondrial membrane potential. We perform experiments to assess the signaling pathway regulating mitochondrial membrane potential and how this impacts follicle cell differentiation. We find that drp1 mutant PFCs show increase in phosphorylated ERK (dpERK) formed downstream of EGFR signaling. ERK regulates high mitochondrial membrane potential in drp1 mutant PFCs. PFCs depleted of ERK and drp1 are able to undergo Notch mediated differentiation. Notably mitochondrial membrane potential decrease via ETC inhibition activates Notch signaling at an earlier stage in wild type and suppresses the Notch signaling defect in drp1 mutant PFCs. Thus, this study shows that the EGFR pathway maintains mitochondrial morphology and mitochondrial membrane potential in follicle cells for its functioning and decrease in mitochondrial membrane potential is needed for Notch mediated differentiation.

Mitochondrial morphology and activity regulate furrow ingression and contractile ring dynamics in Drosophila cellularization.

Mitochondria are maternally inherited in many organisms. Mitochondrial morphology and activity regulation is essential for cell survival, differentiation, and migration. An analysis of mitochondrial dynamics and function in morphogenetic events in early metazoan embryogenesis has not been carried out. In our study we find a crucial role of mitochondrial morphology regulation in cell formation in Drosophila embryogenesis. We find that mitochondria are small and fragmented and translocate apically on microtubules and distribute progressively along the cell length during cellularization. Embryos mutant for the mitochondrial fission protein, Drp1 (dynamin-related protein 1), die in embryogenesis and show an accumulation of clustered mitochondria on the basal side in cellularization. Additionally, Drp1 mutant embryos contain lower levels of reactive oxygen species (ROS). ROS depletion was previously shown to decrease myosin II activity. Drp1 loss also leads to myosin II depletion at the membrane furrow, thereby resulting in decreased cell height and larger contractile ring area in cellularization similar to that in myosin II mutants. The mitochondrial morphology and cellularization defects in Drp1 mutants are suppressed by reducing mitochondrial fusion and increasing cytoplasmic ROS in superoxide dismutase mutants. Our data show a key role for mitochondrial morphology and activity in supporting the morphogenetic events that drive cellularization in Drosophila embryos.

Analysis of mitochondrial organization and function in the Drosophila blastoderm embryo.

Mitochondria are inherited maternally as globular and immature organelles in metazoan embryos. We have used the Drosophila blastoderm embryo to characterize their morphology, distribution and functions in embryogenesis. We find that mitochondria are relatively small, dispersed and distinctly distributed along the apico-basal axis in proximity to microtubules by motor protein transport. Live imaging, photobleaching and photoactivation analyses of mitochondrially targeted GFP show that they are mobile in the apico-basal axis along microtubules and are immobile in the lateral plane thereby associating with one syncytial cell. Photoactivated mitochondria distribute equally to daughter cells across the division cycles. ATP depletion by pharmacological and genetic inhibition of the mitochondrial electron transport chain (ETC) activates AMPK and decreases syncytial metaphase furrow extension. In summary, we show that small and dispersed mitochondria of the Drosophila blastoderm embryo localize by microtubule transport and provide ATP locally for the fast syncytial division cycles. Our study opens the possibility of use of Drosophila embryogenesis as a model system to study the impact of maternal mutations in mitochondrial morphology and metabolism on embryo patterning and differentiation.