Localized mitochondrial metabolic activity in Xenopus mesendoderm cells undergoing collective cell migration

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PROJECT SUMMARY/ABSTRACT [30 LINES MAX] Biomedical advances over the past twenty years have reduced the infant mortality rate in the US throughout the 21st century. However, congenital malformations that arise from dysregulated morphogenesis remain the leading cause of US infant mortality. An especially critical period of morphogenesis is gastrulation, a period of dynamic cellular rearrangements that establish the three tissue germ layers of ectoderm, mesoderm, and endoderm in the embryo. One notable cellular rearrangement in gastrulation is the collective migration of the mesendoderm along the blastocoel roof. Our laboratory identified α5ß1 integrin adhesion to fibronectin as a critical regulator of mesendoderm motility in gastrulation. During collective mesendoderm migration, the front row of migrating cells (leader cells) establishes substantial traction stress on the fibronectin extracellular matrix through α5ß1 integrin that allows for cells in rows behind the leader cells (follower cells) to be pulled forward. While the morphogenetic movements of gastrulation have been characterized, the metabolic processes that support these energetically expensive movements remain poorly understood. The experiments proposed herein will aim to unveil the mechanisms that integrate mechanical and metabolic signals in collective mesendoderm motility. Preliminary live imaging of mitochondrial activity and localization in collectively migrating mesendoderm explants supports the hypothesis that α5ß1 integrin mechanotransduction is a potent inducer of mitochondrial activity and recruitment to adhesions. In Specific Aim 1, we will test the role of α5ß1 integrin in organizing mitochondrial activity and localization, the spatial distribution of metabolites, and rates of glycolysis and mitochondrial respiration. We will induce differential activation of α5ß1 integrin in mesendoderm explants and dissociated primary cells via various fibronectin fusion proteins and varied mechanical environments. We will also modulate α5ß1 integrin activation in the mesendoderm of intact embryos via integrin-targeted monoclonal antibodies. In Specific Aim 2, we will explore the effects of mitochondrial and metabolic dynamic regulators on collective mesendoderm migration using live, high spatiotemporal resolution imaging of mitochondrial activity, mitochondrial localization, the spatial distribution of metabolites, and the function of metabolic regulatory proteins within the context of α5ß1 integrin-driven collective mesendoderm migration. In summary, the experiments proposed will reveal how α5ß1 integrin adhesion and signaling impact the metabolic processes that fuel energetically expensive gastrulation movements. By expanding our knowledge of the mechanisms that engage and fuel tissue rearrangements in gastrulation, we will form a better understanding of processes that contribute to congenital malformations in humans. This proposed fellowship will be performed in a collaborative and supportive environment to develop the trainee’s scientific independence and prepare him for a career as an independent physician-scientist in pediatric pathology. The applicant will be mentored by a supportive dissertation mentor (sponsor) and expert dissertation committee. 1