Zobrazení zdroje stránky Mooneycrawford8606

The elongated antennae decorating eukaryotic glycans are built from polylactosamine repeats. Polylactosamine forms a lectin recognition site and also acts as a platform for presenting diverse additional modifications (e.g., terminal cell-surface antigens); it therefore plays important roles in cell adherence, development, and immunity. Two new papers present a detailed structural and mechanistic investigation of β1-3-N-acetylgucosaminyltransferase 2, a key enzyme in antennae synthesis. The resulting insights will also help decipher other members of GT31, the single largest human glycosyltransferase family.Androglobin (ADGB) represents the latest addition to the globin superfamily in metazoans. The chimeric protein comprises a calpain domain and a unique circularly permutated globin domain. ADGB expression levels are most abundant in mammalian testis, but its cell-type-specific expression, regulation, and function have remained unexplored. Analyzing bulk and single-cell mRNA-Seq data from mammalian tissues, we found that-in addition to the testes-ADGB is prominently expressed in the female reproductive tract, lungs, and brain, specifically being associated with cell types forming motile cilia. Correlation analysis suggested coregulation of ADGB with FOXJ1, a crucial transcription factor of ciliogenesis. Investigating the transcriptional regulation of the ADGB gene, we characterized its promoter using epigenomic datasets, exogenous promoter-dependent luciferase assays, and CRISPR/dCas9-VPR-mediated activation approaches. Reporter gene assays revealed that FOXJ1 indeed substantially enhanced luciferase activity driven by the ADGB promoter. ChIP assays confirmed binding of FOXJ1 to the endogenous ADGB promoter region. We dissected the minimal sequence required for FOXJ1-dependent regulation and fine mapped the FOXJ1 binding site to two evolutionarily conserved regions within the ADGB promoter. FOXJ1 overexpression significantly increased endogenous ADGB mRNA levels in HEK293 and MCF-7 cells. Similar results were observed upon RFX2 overexpression, another key transcription factor in ciliogenesis. The complex transcriptional regulation of the ADGB locus was illustrated by identifying a distal enhancer, responsible for synergistic regulation by RFX2 and FOXJ1. Finally, cell culture studies indicated an ADGB-dependent increase in the number of ciliated cells upon overexpression of the full-length protein, confirming a ciliogenesis-associated role of ADGB in mammals.Alzheimer's disease (AD) is characterized by accumulation of tau and amyloid-beta in the brain, and recent evidence suggests a correlation between associated protein aggregates and trace elements, such as copper, iron, and zinc. In AD, a distorted brain redox homeostasis and complexation by amyloid-beta and hyperphosphorylated tau may alter the isotopic composition of essential mineral elements. Therefore, high-precision isotopic analysis may reveal changes in the homeostasis of these elements. We used inductively coupled plasma-mass spectrometry (ICP-MS)-based techniques to determine the total Cu, Fe, and Zn contents in the brain, as well as their isotopic compositions in both mouse brain and serum. Results for male transgenic tau (Line 66, L66) and amyloid/presenilin (5xFAD) mice were compared with those for the corresponding age- and sex-matched wild-type control mice (WT). Our data show that L66 brains showed significantly higher Fe levels than did those from the corresponding WT. Significantly less Cu, but more Zn was found in 5xFAD brains. We observed significantly lighter isotopic compositions of Fe (enrichment in the lighter isotopes) in the brain and serum of L66 mice compared with WT. For 5xFAD mice, Zn exhibited a trend toward a lighter isotopic composition in the brain and a heavier isotopic composition in serum compared with WT. Neither mouse model yielded differences in the isotopic composition of Cu. Our findings indicate significant pathology-specific alterations of Fe and Zn brain homeostasis in mouse models of AD. The associated changes in isotopic composition may serve as a marker for proteinopathies underlying AD and other types of dementia.Rho/Ras family small GTPases are known to regulate numerous cellular processes, including cytoskeletal reorganization, cell proliferation, and cell differentiation. These processes are also controlled by Ca2+, and consequently, cross talk between these signals is considered likely. However, systematic quantitative evaluation has not yet been reported. To fill this gap, we constructed optogenetic tools to control the activity of small GTPases (RhoA, Rac1, Cdc42, Ras, Rap, and Ral) using an improved light-inducible dimer system (iLID). We characterized these optogenetic tools with genetically encoded red fluorescence intensity-based small GTPase biosensors and confirmed these optogenetic tools' specificities. Using these optogenetic tools, we investigated calcium mobilization immediately after small GTPase activation. Unexpectedly, we found that a transient intracellular calcium elevation was specifically induced by RhoA activation in RPE1 and HeLa cells. RhoA activation also induced transient intracellular calcium elevation in MDCK and HEK293T cells, suggesting that generally RhoA induces calcium signaling. Interestingly, the molecular mechanisms linking RhoA activation to calcium increases were shown to be different among the different cell types In RPE1 and HeLa cells, RhoA activated phospholipase C epsilon (PLCε) at the plasma membrane, which in turn induced Ca2+ release from the endoplasmic reticulum (ER). The RhoA-PLCε axis induced calcium-dependent nuclear factor of activated T cells nuclear translocation, suggesting that it does activate intracellular calcium signaling. Conversely, in MDCK and HEK293T cells, RhoA-ROCK-myosin II axis induced the calcium transients. These data suggest universal coordination of RhoA and calcium signaling in cellular processes, such as cellular contraction and gene expression.Acoustic trauma damages inner ear neural structures including cochlear hair cells which result in hearing loss and neurotransmitter imbalances within the synapses of the central auditory pathway. Disruption of GABA/glutamate levels underlies, tinnitus, a phantom perception of sound that persists post-exposure to blast noise which may manifest in tandem with acute/chronic loss of hearing. Many putative theories explain tinnitus physiology based on indirect and direct assays in animal models and humans, although there is no comprehensive evidence to explain the phenomenon. Here, GABA/glutamate levels were imaged and quantified in a blast overpressure model of chinchillas using Fourier transform ion cyclotron resonance mass spectrometry imaging. The direct measurement from whole-brain sections identified the relative levels of GABA/glutamate in the central auditory neuraxis centers including the cochlear nucleus, inferior colliculus, and auditory cortex. These preliminary results provide insight on the homeostasis of GABA/glutamate within whole-brain sections of chinchilla for investigation of the pathomechanism of blast-induced tinnitus.Retinal vein occlusion (RVO) is a vascular disease that represents characteristic retinal hemorrhage and dilated retinal veins. Despite its clinical importance, its pathogenesis remains largely unknown because of limited opportunities to acquire human retinal samples. Therefore, an animal model that reproduces the clinical features of RVO patients is required for further investigation. In this study, we established a pigmented murine RVO model that reproduced characteristic fundus appearances similar to human RVO findings. [https://www.selleckchem.com/products/hexamethonium-bromide.html click here] Retinal edema in this model was observed in both optical coherence tomography and histological analysis, which is a clinically important outcome. With quantitative real-time PCR analysis on retinal samples, we revealed that the mRNA level of vascular endothelial growth factor (VEGF) increased in the retina induced RVO. Moreover, this retinal edema was reduced by intravitreal injection of anti-VEGF antibody. These results were consistent with human clinical knowledge and suggested that this model could be a useful tool for research into new therapeutic approaches.Hydrogen sulfide (H2S) is an important gasotransmitter expressed in various tissues of the organism, including the eye. It is known that H2S is localized especially in the retina and corneal layers in bovine eye. link2 The enzymes that mediate H2S synthesis are 3-mercaptopyruvate sulfurtransferase (3-MST), cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE). link3 Herein, we aimed to investigate the concentration levels and distribution profiles of these enzymes in bovine retina and retinal artery. Enzyme levels were measured by ELISA and distribution were determined by immunofluorescence microscopic analysis. Much higher concentrations of CBS and CSE have been detected in the retinal artery compared to the retina. In both tissues, particulary 3-MST was found at the lowest level while, CSE was determined to be the most abundant enzyme among the others. CBS distribution was shown in both endothelial and smooth muscle layers, while CSE was seen especially in the endothelial layer of the retinal artery. In the retina, CBS and CSE were expressed in cone-basil cells and retinal ganglion cells, while CSE was also present in bipolar cells. Our results indicated that H2S is synthesized endogenously in ocular tissues. The widespread expression of H2S synthesizing enzymes in the retina and retinal artery of the bovine eye, which has anatomical similarities with the human eye, may suggest a protective role for H2S against retinal vascular diseases as well as a regulatory role in the retinal vascular tone.Retinoblastoma (Rb) is the most common intraocular malignancy in children that accounts for approximately 4% of all pediatric malignancies. Since chemotherapy is a widely practiced treatment for Rb, there is a growing interest in developing new and effective drugs to overcome systemic and local side effects of chemotherapy to improve the quality of life and increase the chances of survival. This study sought to fabricate thiolated chitosan nanoparticles containing topotecan (TPH-TCs-NPs) with a view of enhancing drug loading and release control. This research was also designed to assess the ability of TPH-TCs-NPs to improve cell association, increase treatment efficacy in retinoblastoma cells and xenograft-rat-model of retinoblastoma, and overcome current topotecan hydrochloride (TPH) intravitreal administration challenges, including stability loss and poor cellular uptake. Modified ionic gelation method was optimized to fabricate TPH-TCs-NPs and TPH-TMC-NPs (N-trimethyl chitosan nanoparticles containing TPH)between the tumor control and TPH-TCs-NPs treated group in xenograft-rat-model ( Range of P-value 0.026 to 0.035) was shown by Bonferroni post hoc test. The current investigation demonstrated enhanced efficacy and association of TPH-TCs-NPs relative to free TPH in retinoblastoma cells and tumor in vitro and in vivo.Multistability and natural biological variability can result in significant heterogeneity within a cell population, leading to challenges in understanding and modulating cell behavior. Energy landscapes can offer qualitatively intuitive visualizations of cell phenotype and facilitate a more quantitative understanding of cellular dynamics, but current methods for landscape generation are mathematically involved and often require specific system properties (e.g., ergodicity or independent gene/protein probability distributions) that do not always hold. Here, we present a simple kinetic Monte Carlo-based method for landscape generation from a system of ordinary differential equations using only simulation trajectories initialized throughout the phase space of interest. The resulting landscape produces three quantitative features relevant to understanding cell behavior stability (reflected by the depth or potential of landscape valleys), velocity (representing average directional movement on the landscape), and variance in velocity (indicative of landscape positions with heterogeneous movements).