Biology subjects

Ness, K. M.

Publications and source records attributed to Ness, K. M..

3 recordsLinked to original sources

bioRxiv2023

GAGA Factor Overcomes 1D Diffusion Barrier by 3D Diffusion in Search of Nucleosomal Targets

To elucidate how eukaryotic sequence-specific transcription factors (TFs) search for gene targets on chromatin, we used multi-color smFRET and single-particle imaging to track the diffusion of purified GAGA-Associated Factor (GAF) on DNA and nucleosomes. Monomeric GAF DNA-binding domain (DBD) bearing one zinc finger finds its cognate site by 1D or 3D diffusion on bare DNA and rapidly slides back-and-forth between naturally clustered motifs for seconds before escape. Multimeric, full-length GAF also finds clustered motifs on DNA by 1D-3D diffusion, but remains locked on target for longer periods. Nucleosome architecture effectively blocks GAF-DBD 1D-sliding into the histone core but favors retention of GAF-DBD when targeting solvent-exposed sites by 3D-diffusion. Despite the occlusive power of nucleosomes, 1D-3D facilitated diffusion enables GAF to effectively search for clustered cognate motifs in chromatin, providing a mechanism for navigation to nucleosome and nucleosome-free sites by a member of the largest TF family.

biophysics↗

bioRxiv2022

Microglial inflammatory activation paradoxically improves glucose tolerance during diet-induced obesity

Hypothalamic gliosis associated with high fat diet (HFD) feeding increases susceptibility to hyperphagia and weight gain, and is therefore presumed to promote obesity-associated consequences such as glucose intolerance as well. Nevertheless, the body weight-independent contribution of microglial activation to glucose regulation has not been determined. Here we show that reducing microglial NF-{kappa}B signaling via cell-specific IKK{beta} deletion exacerbates HFD-induced glucose intolerance and insulin resistance despite reducing body weight and adiposity. This effect was associated with reduced activity of hypothalamic glucose sensing neurons. Conversely, a genetic approach to increase microglial inflammatory activity improved glucose tolerance independently of diet in lean rodents. To avoid confounding effects due to chronic alterations to microglial signaling pathways from dietary or genetic interventions, we developed an inducible model of microglial activation using DREADD-based chemogenetics. Gq-coupled GPCR activation rapidly increased microglial calcium levels, cytokine gene expression, and morphological hallmarks of inflammatory activation. In both lean and obese rodents, chemogenetic microglial activation caused a marked improvement in glucose tolerance along with increased activation of hypothalamic glucose sensing neurons, effects abrogated by central blockade of TNF signaling. Thus, while diet-induced microglial activation promotes weight gain, it may also serve an adaptive function--to prevent the deterioration of glucose tolerance associated with obesity, an important consideration for immune-modulating metabolic therapies.

neuroscience↗

bioRxiv2021

Prostaglandin PGE2 receptor EP4 regulates microglial phagocytosis and increases susceptibility to diet-induced obesity

BackgroundIn rodents, susceptibility to diet-induced obesity requires microglial activation, but the molecular components of this pathway remain incompletely defined. Prostaglandin E2 (PGE2) levels increase in the mediobasal hypothalamus during high fat diet (HFD) feeding, and the PGE2 receptor EP4 regulates microglial activation state and phagocytic activity, suggesting a potential role for microglial EP4 signaling in obesity pathogenesis. MethodMetabolic phenotyping, as assessed by body weight, energy expenditure, glucose, and insulin tolerance, was performed in microglia-specific EP4 knockout (MG-EP4 KO) mice and littermate controls on HFD. Morphological and gene expression analysis of microglia, and a histological survey of microglia-neuron interactions in the arcuate nucleus was performed. Phagocytosis was assessed using in vivo and in vitro pharmacological techniques. ResultsMicroglial EP4 deletion markedly reduced weight gain and food intake in response to HFD feeding. In correspondence with this lean phenotype, insulin sensitivity was also improved in the HFD-fed MG-EP4 KO mice though glucose tolerance remained surprisingly unaffected. Mechanistically, EP4-deficient microglia showed an attenuated phagocytic state marked by reduced CD68 expression and fewer contacts with POMC neuron soma and processes. These cellular changes observed in the microglial EP4 knockout mice corresponded with an increased density of POMC neurites extending into the paraventricular nucleus. ConclusionThese findings reveal that microglial EP4 signaling promotes body weight gain and insulin resistance during HFD feeding. Furthermore, the data suggest that curbing microglial phagocytic function may preserve POMC cytoarchitecture and PVN input to limit overconsumption during diet-induced obesity.

physiology↗