ABSTRACT
Chromatin, which consists of DNA and associated proteins, contains genetic information and is a mechanical component of the nucleus. Heterochromatic histone methylation controls nucleus and chromosome stiffness, but the contribution of heterochromatin protein HP1α (CBX5) is unknown. We used a novel HP1α auxin-inducible degron human cell line to rapidly degrade HP1α. Degradation did not alter transcription, local chromatin compaction, or histone methylation, but did decrease chromatin stiffness. Single-nucleus micromanipulation reveals that HP1α is essential to chromatin-based mechanics and maintains nuclear morphology, separate from histone methylation. Further experiments with dimerization-deficient HP1αI165E indicate that chromatin crosslinking via HP1α dimerization is critical, while polymer simulations demonstrate the importance of chromatin-chromatin crosslinkers in mechanics. In mitotic chromosomes, HP1α similarly bolsters stiffness while aiding in mitotic alignment and faithful segregation. HP1α is therefore a critical chromatin-crosslinking protein that provides mechanical strength to chromosomes and the nucleus throughout the cell cycle and supports cellular functions.
Subject(s)
Cell Nucleus/metabolism , Chromatin , Chromosomal Proteins, Non-Histone , Chromosomes , Mitosis/physiology , Cell Line , Cell Nucleus/chemistry , Chromatin/chemistry , Chromatin/metabolism , Chromobox Protein Homolog 5 , Chromosomal Proteins, Non-Histone/chemistry , Chromosomal Proteins, Non-Histone/metabolism , Chromosomes/chemistry , Chromosomes/metabolism , Humans , MethylationABSTRACT
During human brain development, multiple signaling pathways generate diverse cell types with varied regional identities. Here, we integrate single-cell RNA sequencing and clonal analyses to reveal lineage trees and molecular signals underlying early forebrain and mid/hindbrain cell differentiation from human embryonic stem cells (hESCs). Clustering single-cell transcriptomic data identified 41 distinct populations of progenitor, neuronal, and non-neural cells across our differentiation time course. Comparisons with primary mouse and human gene expression data demonstrated rostral and caudal progenitor and neuronal identities from early brain development. Bayesian analyses inferred a unified cell-type lineage tree that bifurcates between cortical and mid/hindbrain cell types. Two methods of clonal analyses confirmed these findings and further revealed the importance of Wnt/ß-catenin signaling in controlling this lineage decision. Together, these findings provide a rich transcriptome-based lineage map for studying human brain development and modeling developmental disorders.
Subject(s)
Brain/embryology , Cell Lineage , Embryonic Development , Human Embryonic Stem Cells/cytology , Single-Cell Analysis/methods , Animals , Brain/metabolism , Cell Line , Cell Lineage/genetics , Clone Cells , Embryonic Development/genetics , Humans , Mice , Models, Biological , Neurons/cytology , Neurons/metabolism , Reproducibility of Results , Sequence Analysis, RNA , Transcription Factors/metabolism , Transcriptome/genetics , Wnt Signaling Pathway/geneticsABSTRACT
Spectral sensitivity of humans and rhesus monkeys was compared using identical displays and similar procedures. Detection thresholds were measured for the following: 1) 15-Hz modulation of a blue and a green cathode-ray tube phosphor; 2) 15-Hz modulation of all three phosphors together; and 3) slow (<1 Hz) modulations of a blue and a green phosphor under scotopic conditions. Monkeys had lower blue-to-green threshold ratios than humans at all eccentricities tested (0.5 to 7°), consistent with a lower lens optical density in monkeys. In addition to apparently having a lower lens density than humans, monkeys were more sensitive to 15-Hz red-green isoluminant modulations than humans, an effect that cannot be explained by optical factors.
Subject(s)
Color Perception , Contrast Sensitivity , Psychophysics , Signal Detection, Psychological , Adult , Animals , Female , Humans , Macaca mulatta , Male , Neurophysiology , Photic Stimulation , Visual Fields , Young AdultABSTRACT
Quercetin is a common flavonoid polyphenol which has been shown to exert neuroprotective actions in vitro and in vivo. Though quercetin has antioxidant properties, it has been suggested that neuroprotection may be ascribed to its ability of inducing the cell's own defense mechanisms. The present study investigated whether quercetin could increase the levels of paraoxonase 2 (PON2), a mitochondrial enzyme expressed in brain cells, which has been shown to have potent antioxidant properties. PON2 protein, mRNA, and lactonase activity were highest in mouse striatal astrocytes. Quercetin increased PON2 levels, possibly by activating the JNK/AP-1 pathway. The increased PON2 levels induced by quercetin resulted in decreased oxidative stress and ensuing toxicity induced by two oxidants. The neuroprotective effect of quercetin was significantly diminished in cells from PON2 knockout mice. These findings suggest that induction of PON2 by quercetin represents an important mechanism by which this polyphenol may exert its neuroprotective action.
