Developmental Biology Alliance
University of Georgia
This laboratory and lecture course is designed to provide neuroscientists at all levels with a conceptual and practical understanding of several of the most advanced techniques in molecular neuroscience.
G protein-coupled receptors play broad roles in development and stem cell biology, but few roles for G protein-coupled receptor signaling in complex tissue regeneration have been uncovered. Planarian flatworms robustly regenerate all tissues and provide a model with which to explore potential functions for G protein-coupled receptor signaling in somatic regeneration and pluripotent stem cell biology. As a first step toward exploring G protein-coupled receptor function in planarians, we investigated downstream signal transducers that work with G protein-coupled receptors, called heterotrimeric G proteins. Here, we characterized the complete heterotrimeric G protein complement in Schmidtea mediterranea for the first time and found that 7 heterotrimeric G protein subunits promote regeneration. We further characterized 2 subunits critical for regeneration, Gαq1 and Gβ1-4a, finding that they promote the late phase of anterior polarity reestablishment, likely through anterior pole-produced Follistatin. Incidentally, we also found that 5 G protein subunits modulate planarian behavior. We further identified a putative serotonin receptor, gcr052, that we propose works with Gαs2 and Gβx2 in planarian locomotion, demonstrating the utility of our strategy for identifying relevant G protein-coupled receptors. Our work provides foundational insight into roles of heterotrimeric G proteins in planarian biology and serves as a useful springboard toward broadening our understanding of G protein-coupled receptor signaling in adult tissue regeneration.
Sensory neurons (SNs) detect a wide range of information from the body and the environment that is critical for homeostasis. There are three main subtypes of SNs: nociceptors, mechanoreceptors, and proprioceptors, which express different membrane proteins, such as TRKA, TRKB, or TRKC, respectively. Human pluripotent stem cell technology provides an ideal platform to study development and diseases of SNs, however there is not a viable method to isolate individual SN subtype for downstream analysis available. Here, we employ the method immunopanning to isolate each SN subtype. This method is very gentle and allows proper survival after the isolation. We use antibodies against TRKA, TRKB, and TRKC to isolate nociceptors, mechanoreceptors, and proprioceptors, respectively. We show that our cultures are enriched for each subtype and express their respective subtype markers. Furthermore, we show that the immunopanned SNs are electrically active and respond to specific stimuli. Thus, our method can be used to purify viable neuronal subtypes using respective membrane proteins for downstream studies.
O-GlcNAcylation is a post-translational modification (PTM) that regulates a wide range of cellular functions and has been associated with multiple metabolic diseases in various organs. The sympathetic nervous system (SNS) is the efferent portion of the autonomic nervous system that regulates metabolism of almost all organs in the body. How much the development and functionality of the SNS are influenced by O-GlcNAcylation, as well as how such regulation could contribute to sympathetic neuron (symN)-related neuropathy in diseased states, remains unknown. Here, we assessed the level of protein O-GlcNAcylation at various stages of symN development, using a human pluripotent stem cell (hPSC)-based symN differentiation paradigm. We found that pharmacological disruption of O-GlcNAcylation impaired both the growth and survival of hPSC-derived symNs. In the high glucose condition that mimics hyperglycemia, hPSC-derived symNs were hyperactive, and their regenerative capacity was impaired, which resembled typical neuronal defects in patients and animal models of diabetes mellitus. Using this model of sympathetic neuropathy, we discovered that O-GlcNAcylation increased in symNs under high glucose, which lead to hyperactivity. Pharmacological inhibition of O-GlcNAcylation rescued high glucose-induced symN hyperactivity and cell stress. This framework provides the first insight into the roles of O-GlcNAcylation in both healthy and diseased human symNs and may be used as a platform for therapeutic studies.
Snakes and mice might not look much alike, but much of what we know about skin coloration and patterning in vertebrates, including snakes, is based on research involving lab mice. There are limits, however, to what mice can tell us about other vertebrates because they don’t share all the same types of color-producing cells, known as chromatophores. For example, snakes have a type of chromatophore called iridophores that can generate iridescent colors by reflecting light.
Familial dysautonomia (FD), a rare neurodevelopmental and neurodegenerative disorder affects the sympathetic and sensory nervous system. Although almost all patients harbor a mutation in ELP1, it remains unresolved exactly how function of sympathetic neurons (symNs) is affected; knowledge critical for understanding debilitating disease hallmarks, including cardiovascular instability or dysautonomic crises, that result from dysregulated sympathetic activity. Here, we employ the human pluripotent stem cell (hPSC) system to understand symN disease mechanisms and test candidate drugs. FD symNs are intrinsically hyperactive in vitro, in cardiomyocyte co-cultures, and in animal models. We report reduced norepinephrine transporter expression, decreased intracellular norepinephrine (NE), decreased NE re-uptake, and excessive extracellular NE in FD symNs. SymN hyperactivity is not a direct ELP1 mutation result, but may connect to NET via RAB proteins. We found that candidate drugs lowered hyperactivity independent of ELP1 modulation. Our findings may have implications for other symN disorders and may allow future drug testing and discovery.
University of Georgia researcher Pengpeng Bi received a pair of National Institutes of Health grants in September: a Maximizing Investigators’ Research Award (MIRA, 2022–2027) and an Exploratory/Developmental Research Grant Award (R21, 2022–2024). The $2.3 million awards will support efforts to uncover the molecular mechanism of human muscle development and homeostasis.
University of Georgia faculty member Tania Rozario has received a $2 million grant from the National Institutes of Health Director’s New Innovator Award Program, which supports early-career investigators of exceptional creativity who propose high-risk, high-reward research projects.
Chromatin modifications function as critical regulators of gene expression and cellular identity, especially in the regulation and maintenance of the pluripotent state. However, many studies of chromatin modification in stem cells—and pluripotent stem cells in particular—are performed in mammalian stem cell culture, an in vitro condition mimicking a very transient state during mammalian development. Thus, new models for studying pluripotent stem cells in vivo could be helpful for understanding the roles of chromatin modification, for confirming prior in vitro studies, and for exploring evolution of the pluripotent state.
The most important organ for your body’s immune system isn’t working well anymore. The thymus is an organ most people have never heard of, but it serves a vital purpose. The thymus produces the body’s T cells, which serve as the immune system’s front line against disease.
