IU Indianapolis ScholarWorks :: Browsing by Subject "Signaling"

Browsing by Subject "Signaling"

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Autophagy induced by calcium phosphate precipitates targets damaged endosomes
(ASBMB, 2014-04-18) Chen, Xi; Khambu, Bilon; Zhang, Hao; Gao, Wentao; Li, Min; Chen, Xiaoyun; Yoshimori, Tamotsu; Yin, Xiao-Ming; Department of Pathology & Laboratory Medicine, IU School of Medicine
Calcium phosphate precipitates (CPPs) form complexes with DNA, which enter cells via endocytosis. Under this condition CPPs induce autophagy via the canonic autophagy machinery. Here we showed that CPP-induced autophagy was also dependent on endocytosis as the process was significantly inhibited by methyl-β-cyclodextrin and dynasore, which suppress clathrin-dependent endocytosis. Consistently, CPP treatment triggered the formation of filipin-positive intracellular vesicles whose membranes are rich in cholesterol. Unexpectedly, these vesicles were also positive for galectin 3, suggesting that they were damaged and the membrane glycans became accessible to galectins to bind. Endosome damage was caused by endocytosis of CPPs and was reversed by calcium chelators or by endocytosis inhibitors. Notably, CPP-induced LC3-positive autophagosomes were colocalized with galectin 3, ubiquitin, and p62/SQSTM1. Inhibition of galectin 3 reduced p62 puncta and autophagosome formation. Knockdown of p62 additionally inhibited the colocalization of autophagosomes with galectins. Furthermore, most of the galectin 3-positive vesicles were colocalized with Rab7 or LAMP1. Agents that affect endosome/lysosome maturation and function, such as bafilomycin A1, also significantly affected CPP-induced tubulovesicular autophagosome formation. These findings thus indicate that endocytosed CPPs caused endosome damage and recruitment of galectins, particularly at the later endosome stage, which led to the interaction of the autophagosomal membranes with the damaged endosome in the presence of p62.
Cell-Cell Communication Breakdown and Endothelial Dysfunction
(Elsevier, 2020-04) Lee, Daniel D.; Schwarz, Margaret A.; Medicine, School of Medicine
Guided by organ-specific signals in both development and disease response, the heterogeneous endothelial cell population is a dynamic member of the vasculature. Functioning as the gatekeeper to fluid, inflammatory cells, oxygen, and nutrients, endothelial cell communication with its local environment is critical. Impairment of endothelial cell-cell communication not only disrupts this signaling process, but also contributes to pathologic disease progression. Expanding our understanding of those processes that mediate endothelial cell-cell communication is an important step in the approach to treatment of disease processes.
Collagen in Wound Healing
(MDPI, 2021-05-11) Mathew-Steiner, Shomita S.; Roy, Sashwati; Sen, Chandan K.; Medicine, School of Medicine
Normal wound healing progresses through inflammatory, proliferative and remodeling phases in response to tissue injury. Collagen, a key component of the extracellular matrix, plays critical roles in the regulation of the phases of wound healing either in its native, fibrillar conformation or as soluble components in the wound milieu. Impairments in any of these phases stall the wound in a chronic, non-healing state that typically requires some form of intervention to guide the process back to completion. Key factors in the hostile environment of a chronic wound are persistent inflammation, increased destruction of ECM components caused by elevated metalloproteinases and other enzymes and improper activation of soluble mediators of the wound healing process. Collagen, being central in the regulation of several of these processes, has been utilized as an adjunct wound therapy to promote healing. In this work the significance of collagen in different biological processes relevant to wound healing are reviewed and a summary of the current literature on the use of collagen-based products in wound care is provided.
Human GPR17 missense variants identified in metabolic disease patients have distinct downstream signaling profiles
(Elsevier, 2021-07) Conley, Jason M.; Sun, Hongmao; Ayers, Kristin L.; Zhu, Hu; Chen, Rong; Shen, Min; Hall, Matthew D.; Ren, Hongxia; Pediatrics, School of Medicine
GPR17 is a G-protein-coupled receptor (GPCR) implicated in the regulation of glucose metabolism and energy homeostasis. Such evidence is primarily drawn from mouse knockout studies and suggests GPR17 as a potential novel therapeutic target for the treatment of metabolic diseases. However, links between human GPR17 genetic variants, downstream cellular signaling, and metabolic diseases have yet to be reported. Here, we analyzed GPR17 coding sequences from control and disease cohorts consisting of individuals with adverse clinical metabolic deficits including severe insulin resistance, hypercholesterolemia, and obesity. We identified 18 nonsynonymous GPR17 variants, including eight variants that were exclusive to the disease cohort. We characterized the protein expression levels, membrane localization, and downstream signaling profiles of nine GPR17 variants (F43L, V96M, V103M, D105N, A131T, G136S, R248Q, R301H, and G354V). These nine GPR17 variants had similar protein expression and subcellular localization as wild-type GPR17; however, they showed diverse downstream signaling profiles. GPR17-G136S lost the capacity for agonist-mediated cAMP, Ca2+, and β-arrestin signaling. GPR17-V96M retained cAMP inhibition similar to GPR17-WT, but showed impaired Ca2+ and β-arrestin signaling. GPR17-D105N displayed impaired cAMP and Ca2+ signaling, but unaffected agonist-stimulated β-arrestin recruitment. The identification and functional profiling of naturally occurring human GPR17 variants from individuals with metabolic diseases revealed receptor variants with diverse signaling profiles, including differential signaling perturbations that resulted in GPCR signaling bias. Our findings provide a framework for structure-function relationship studies of GPR17 signaling and metabolic disease.
Mechanical Regulation of Wnt/β-catenin Signaling in Bone Cells
(Office of the Vice Chancellor for Research, 2010-04-09) Hum, J.M.; Young, S.R.L.; Gerard-O’Riley, R.; Pavalko, F.M.
The Wnt/β-catenin signaling pathway is an important regulatory pathway in development and maintenance of various tissues, including bone. Active Wnt interacts with the frizzled/LRP receptor activating dishevelled, which in turn inactivates the GSK-3β complex and allows βcatenin to accumulate in the cytoplasm. β-catenin translocates to the nucleus where it activates a wide number of developmental target genes. Wnt can be sequestered by soluble frizzled related protein causing the inactivation of dishevelled, allowing for activation of the GSK-3β complex. This activated complex binds β-catenin and targets it for degradation. In addition to its other major role as a linker between cadherins and the actin cytoskeleton, β-catenin accumulation in the cytoplasm and subsequent translocation to the nucleus is a key step in the wnt/β-catenin signaling pathway. In bone, wnt/β-catenin signaling regulates skeletal formation, limb development and osteoblast maturation. Both active and inactive wnt/β-catenin signaling regulate bone cell development, active wnt/β-catenin signaling promotes osteoblast formation, while inactive wnt/β-catenin signaling inhibits osteoclast differentiation. Mechanical regulation of bone cells occurs through a process known as mechanotransduction which can be induced by fluid shear stress that occurs across the surfaces of osteoblasts and osteocytes, the effector cells of mechanotransduction. We hypothesize that knocking down β-catenin expression in mouse osteoblasts and osteoprogenitors will change the way these cells respond to fluid shear stress and regulate expression of relevant bone target genes. The future aims of this project are to assess the role of β-catenin during fluid shear stress induced osteoprogenitor cell differentiation by examining the expression of important osteoblast differentiation markers including: runx2, COX2, osteopontin, and osteocalcin and evaluate the significance of β-catenin during differentiation of bone marrow stromal cells.
Mechanisms Regulating the Association of Protein Phosphatase 1 with Spinophilin and Neurabin
(American Chemical Society, 2018-11-21) Edler, Michael C.; Salek, Asma B.; Watkins, Darryl S.; Kaur, Harjot; Morris, Cameron W.; Yamamoto, Bryan K.; Baucum, Anthony J., II; Biology, School of Science
Protein phosphorylation is a key mediator of signal transduction, allowing for dynamic regulation of substrate activity. Whereas protein kinases obtain substrate specificity by targeting specific amino acid sequences, serine/threonine phosphatase catalytic subunits are much more promiscuous in their ability to dephosphorylate substrates. To obtain substrate specificity, serine/threonine phosphatases utilize targeting proteins to regulate phosphatase subcellular localization and catalytic activity. Spinophilin and its homologue neurabin are two of the most abundant dendritic spine-localized protein phosphatase 1 (PP1) targeting proteins. The association between spinophilin and PP1 is increased in the striatum of animal models of Parkinson's disease (PD). However, mechanisms that regulate the association of spinophilin and neurabin with PP1 are unclear. Here, we report that the association between spinophilin and PP1α or PP1γ1 was increased by CDK5 expression and activation in a heterologous cell system. This increased association is at least partially due to phosphorylation of PP1. Conversely, CDK5 expression and activation decreased the association of PP1 with neurabin. As with dopamine depletion, methamphetamine (METH) abuse causes persistent alterations in dopamine signaling which influence striatal medium spiny neuron function and biochemistry. Moreover, both METH toxicity and dopamine depletion are associated with deficits in motor control and motor learning. Pathologically, we observed a decreased association of spinophilin with PP1 in rat striatum evaluated one month following a binge METH paradigm. Behaviorally, we found that loss of spinophilin recapitulates rotarod pathology previously observed in dopamine-depleted and METH-treated animals. Together, these data have implications in multiple disease states associated with altered dopamine signaling such as PD and psychostimulant drug abuse and delineate a novel mechanism by which PP1 interactions with spinophilin and neurabin may be differentially regulated.
Predictions on and Analysis of Viral Proteins Encoded by Overlapping Genes
(2011-08-19) Khosravi, Mahvash; Dunker, A. Keith
Overlapping genes are adjacent genes that share a portion of their coding sequence. Such genes are often observed in the compact genomes of viruses, prokaryotes,and mitochondria. Overlapping genes are also seen in human and other mammalian genomes. Gene overlapping is a phenomenon to minimize genomic size and maximize encoding capacity. Overlapping genes produce different proteins. A major task in the post genomic era is the large-scale study of the structures and functions of proteins. Proteins play crucial roles in virtually all biological processes. In general it is assumed that 3-D structure determines the function of proteins, but many proteins or region of proteins may function in the absence of 3-D structure. The term disordered is used to describe these proteins. A large number of studies has shown that biological functions depend on both ordered and disordered proteins. Natively disordered regions are common and play essential roles in many proteins, especially, with regard to activities involved in signaling and regulation. The goal of this research was the analysis of the ordered and disordered tendencies of viral proteins encoded by overlapping genes. Our hypothesis is that, in a pair of proteins or protein regions encoded by overlapping genes, at least one of the pair is disordered (or unstructured). Our hypothesis is based on the observation that structural proteins require highly specific amino acid sequences, while unstructured (disordered) sequences are essentially unconstrained. Thus, given a structural protein and its associated mRNA sequence, any sequence derived from an overlapping reading frame seems highly unlikely to have a sequence pattern commensurate with a structural protein; on the other hand, a sequence pattern consistent with a disordered protein seems much more likely. We performed studies on the protein products of overlapping gene sequences, tested the hypothesis and addressed the following two questions: First do the proteins encoded by overlapping genes have opposite order-disorder content, that is, does the ordered part of one of the overlapping proteins correspond to a disordered part in the other overlapping protein? Second, does the encoded protein in the overlapping regions have more disordered amino acids than the non-overlapping regions? Using our database of overlapping viral genes and the protein predictor PONDR VL3, we predicted the order-disorder of amino acids in the sequence of 97 viral protein samples. An analysis of the results supported our hypothesis and indicated that the ordered amino acids are mostly associated with non-overlapping regions while disordered amino acids are more prevalent in overlapping regions. In the overlapping regions for 52 protein pairs, we showed that most of the amino acid pairs facing each other on the protein sequences had at least one disorder for most cases. Out of 52 pairs, there were 3 protein pairs where there were no disordered amino acids and 22 protein pairs where there were no ordered amino acids on either sequence. The fraction of ordered pairs in the pool of overlapping regions of 52 protein pairs was 0.28. The non-overlapping region of 97 proteins had predominantly ordered proteins. The fraction of ordered amino acids in the pool of non-overlapping regions was determined to be 0.77.
Proteomic Analysis of Postsynaptic Protein Complexes Underlying Neuronal Plasticity
(American Chemical Society, 2017-04-19) Baucum, Anthony J., II; Biology, School of Science
Normal neuronal communication and synaptic plasticity at glutamatergic synapses requires dynamic regulation of postsynaptic molecules. Protein expression and protein post-translational modifications regulate protein interactions that underlie this organization. In this Review, we highlight data obtained over the last 20 years that have used qualitative and quantitative proteomics-based approaches to identify postsynaptic protein complexes. Herein, we describe how these proteomics studies have helped lay the foundation for understanding synaptic physiology and perturbations in synaptic signaling observed in different pathologies. We also describe emerging technologies that can be useful in these analyses. We focus on protein complexes associated with the highly abundant and functionally critical proteins: calcium/calmodulin-dependent protein kinase II, the N-methyl-d-aspartate, and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid glutamate receptors, and postsynaptic density protein of 95 kDa.
Signal Transduction and Molecular Regulation in Fatty Liver Disease
(Mary Ann Liebert, 2021) Dong, Xiaocheng Charlie; Chowdhury, Kushan; Huang, Menghao; Kim, Hyeong Geug; Biochemistry and Molecular Biology, School of Medicine
Significance: Fatty liver disease is a major liver disorder in the modern societies. Comprehensive understanding of the pathophysiology and molecular mechanisms is essential for the prevention and treatment of the disease. Recent Advances: Remarkable progress has been made in the recent years in basic and translational research in the field of fatty liver disease. Multiple signaling pathways have been implicated in the development of fatty liver disease, including AMP-activated protein kinase, mechanistic target of rapamycin kinase, endoplasmic reticulum stress, oxidative stress, inflammation, transforming growth factor β, and yes1-associated transcriptional regulator/transcriptional coactivator with PDZ-binding motif (YAP/TAZ). In addition, critical molecular regulations at the transcriptional and epigenetic levels have been linked to the pathogenesis of fatty liver disease. Critical Issues: Some critical issues remain to be solved so that research findings can be translated into clinical applications. Robust and reliable biomarkers are needed for diagnosis of different stages of the fatty liver disease. Effective and safe molecular targets remain to be identified and validated. Prevention strategies require solid scientific evidence and population-wide feasibility. Future Directions: As more data are generated with time, integrative approaches are needed to comprehensively understand the disease pathophysiology and mechanisms at multiple levels from population, organismal system, organ/tissue, to cell. The interactions between genes and environmental factors require deeper investigation for the purposes of prevention and personalized treatment of fatty liver disease.
Spinophilin Limits Metabotropic Glutamate Receptor 5 Scaffolding to the Postsynaptic Density and Cell Type Specifically Mediates Excessive Grooming
(Elsevier, 2023) Morris, Cameron W.; Watkins, Darryl S.; Shah, Nikhil R.; Pennington, Taylor; Hens, Basant; Qi, Guihong; Doud, Emma H.; Mosley, Amber L.; Atwood, Brady K.; Baucum, Anthony J., II; Pharmacology and Toxicology, School of Medicine
Background: Grooming dysfunction is a hallmark of the obsessive-compulsive spectrum disorder trichotillomania. Numerous preclinical studies have utilized SAPAP3-deficient mice for understanding the neurobiology of repetitive grooming, suggesting that excessive grooming is caused by increased metabotropic glutamate receptor 5 (mGluR5) activity in striatal direct- and indirect-pathway medium spiny neurons (MSNs). However, the MSN subtype-specific signaling mechanisms that mediate mGluR5-dependent adaptations underlying excessive grooming are not fully understood. Here, we investigated the MSN subtype-specific roles of the striatal signaling hub protein spinophilin in mediating repetitive motor dysfunction associated with mGluR5 function. Methods: Quantitative proteomics and immunoblotting were utilized to identify how spinophilin impacts mGluR5 phosphorylation and protein interaction changes. Plasticity and repetitive motor dysfunction associated with mGluR5 action were measured using our novel conditional spinophilin mouse model in which spinophilin was knocked out from striatal direct-pathway MSNs and/or indirect-pathway MSNs. Results: Loss of spinophilin only in indirect-pathway MSNs decreased performance of a novel motor repertoire, but loss of spinophilin in either MSN subtype abrogated striatal plasticity associated with mGluR5 function and prevented excessive grooming caused by SAPAP3 knockout mice or treatment with the mGluR5-specific positive allosteric modulator VU0360172 without impacting locomotion-relevant behavior. Biochemically, we determined that the spinophilin-mGluR5 interaction correlates with grooming behavior and that loss of spinophilin shifts mGluR5 interactions from lipid raft-associated proteins toward postsynaptic density proteins implicated in psychiatric disorders. Conclusions: These results identify spinophilin as a novel striatal signaling hub molecule in MSNs that cell subtype specifically mediates behavioral, functional, and molecular adaptations associated with repetitive motor dysfunction in psychiatric disorders.

Browsing by Subject "Signaling"

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