Reduction of Hox Gene Expression by Histone H1 Depletion

The evolutionarily conserved homeotic (Hox) genes are organized in clusters and expressed collinearly to specify body patterning during embryonic development. Chromatin reorganization and decompaction are intimately connected with Hox gene activation. Linker histone H1 plays a key role in facilitating folding of higher order chromatin structure. Previous studies have shown that deletion of three somatic H1 subtypes together leads to embryonic lethality and that H1c/H1d/H1e triple knockout (TKO) embryonic stem cells (ESCs) display bulk chromatin decompaction. To investigate the potential role of H1 and higher order chromatin folding in the regulation of Hox gene expression, we systematically analyzed the expression of all 39 Hox genes in triple H1 null mouse embryos and ESCs by quantitative RT-PCR. Surprisingly, we find that H1 depletion causes significant reduction in the expression of a broad range of Hox genes in embryos and ESCs. To examine if any of the three H1 subtypes (H1c, H1d and H1e) is responsible for decreased expression of Hox gene in triple-H1 null ESCs, we derived and characterized H1c−/−, H1d−/−, and H1e−/− single-H1 null ESCs. We show that deletion of individual H1 subtypes results in down-regulation of specific Hox genes in ESCs. Finally we demonstrate that, in triple-H1- and single-H1- null ESCs, the levels of H3K4 trimethylation (H3K4me3) and H3K27 trimethylation (H3K27me3) were affected at specific Hox genes with decreased expression. Our data demonstrate that marked reduction in total H1 levels causes significant reduction in both expression and the level of active histone mark H3K4me3 at many Hox genes and that individual H1 subtypes may also contribute to the regulation of specific Hox gene expression. We suggest possible mechanisms for such an unexpected role of histone H1 in Hox gene regulation

Inhibition of DNA methyltransferases and histone deacetylases induces astrocytic differentiation of neural progenitors

AbstractUnderstanding how to specify rapid differentiation of human neural progenitor towards enriched non-transformed human astrocyte progenitors will provide a critical cell source to further our understanding of how astrocytes play a pivotal role in neural function and development. Human neural progenitors derived from pluripotent embryonic stem cells and propagated in adherent serum-free cultures provide a fate restricted renewable source for quick production of neural cells; however, such cells are highly refractive to astrocytogenesis and show a strong neurogenic bias, similar to neural progenitors from the early embryonic central nervous system (CNS). We found that several astrocytic genes are hypermethylated in such progenitors potentially preventing generation of astrocytes and leading to the proneuronal fate of these progenitors. However, epigenetic modification by Azacytidine (Aza-C) and Trichostatin A (TSA), with concomitant signaling from BMP2 and LIF in neural progenitor cultures shifts this bias, leading to expression of astrocytic markers as early as 5days of differentiation, with near complete suppression of neuronal differentiation. The resultant cells express major astrocytic markers, are amenable to co-culture with neurons, can be propagated as astrocyte progenitors and are cryopreservable. Although previous reports have generated astrocytes from pluripotent cells, the differentiation required extensive culture or selection based on cell surface antigens. The development of a label free and rapid differentiation process will expedite future derivation of astrocytes from various sources pluripotent cells including, but not limited to, human astrocytes associated with various neurological diseases

NLRP6 Serves as a Negative Regulator of Neutrophil Recruitment and Function During Streptococcus pneumoniae Infection

Streptococcus pneumoniae is an invasive pathogen with high morbidity and mortality in the immunocompromised children and elderly. NOD-like receptor family pyrin domain containing 6 (NLRP6) plays an important role in the host innate immune response against pathogen infections. Our previous studies have shown that NLRP6 plays a negative regulatory role in host defense against S. pneumoniae, but the underlying mechanism is still unclear. The further negative regulatory role of NLRP6 in the host was investigated in this study. Our results showed that NLRP6(-/-) mice in the lung had lower bacterial burdens after S. pneumoniae infection and expressed higher level of tight junction (TJ) protein occludin compared to WT mice, indicating the detrimental role of NLRP6 in the host defense against S. pneumoniae infection. Transcriptome analysis showed that genes related to leukocytes migration and recruitment were differentially expressed between wild-type (WT) and NLRP6 knockout (NLRP6(-/-)) mice during S. pneumoniae infection. Also, NLRP6(-/-) mice showed higher expression of chemokines including C-X-C motif chemokine ligand 1 (CXCL1) and 2 (CXCL2) and lower gene expression of complement C3a receptor 1 (C3aR1) and P-selectin glycoprotein ligand-1 (PSGL-1) which are the factors that inhibit the recruitment of neutrophils. Furthermore, NLRP6(-/-) neutrophils showed increased intracellular bactericidal ability and the formation of neutrophil extracellular traps (NETs) during S. pneumoniae infection. Taken together, our study suggests that NLRP6 is a negative regulator of neutrophil recruitment and function during S. pneumoniae infection. Our study provides a new insight to develop novel strategies to treat invasive pneumococcal infection

Colorimetric Assay for Determination of Lead (II) Based on Its Incorporation into Gold Nanoparticles during Their Synthesis

In this report, we present a new method for visual detection of Pb2+. Gold nanoparticles (Au-NPs) were synthesized in one step at room temperature, using gallic acid (GA) as reducer and stabilizer. Pb2+ is added during the gold nanoparticle formation. Analysis of Pb2+ is conducted by a dual strategy, namely, colorimetry and spectrometry. During Au-NPs synthesis, addition of Pb2+ would lead to formation of Pb-GA complex, which can induce the aggregation of newly-formed small unstable gold nanoclusters. Consequently, colorimetric detection of trace Pb2+ can be realized. As the Pb2+ concentration increases, the color turns from red-wine to purple, and finally blue. This method offers a sensitive linear correlation between the shift of the absorption band (Δλ) and logarithm of Pb2+ concentration ranging from 5.0 × 10−8 to 1.0 × 10−6 M with a linear fit coefficient of 0.998, and a high selectivity for Pb2+ detection with a low detection limit down to 2.5 × 10−8 M

Histone H1 Depletion Impairs Embryonic Stem Cell Differentiation

Pluripotent embryonic stem cells (ESCs) are known to possess a relatively open chromatin structure; yet, despite efforts to characterize the chromatin signatures of ESCs, the role of chromatin compaction in stem cell fate and function remains elusive. Linker histone H1 is important for higher-order chromatin folding and is essential for mammalian embryogenesis. To investigate the role of H1 and chromatin compaction in stem cell pluripotency and differentiation, we examine the differentiation of embryonic stem cells that are depleted of multiple H1 subtypes. H1c/H1d/H1e triple null ESCs are more resistant to spontaneous differentiation in adherent monolayer culture upon removal of leukemia inhibitory factor. Similarly, the majority of the triple-H1 null embryoid bodies (EBs) lack morphological structures representing the three germ layers and retain gene expression signatures characteristic of undifferentiated ESCs. Furthermore, upon neural differentiation of EBs, triple-H1 null cell cultures are deficient in neurite outgrowth and lack efficient activation of neural markers. Finally, we discover that triple-H1 null embryos and EBs fail to fully repress the expression of the pluripotency genes in comparison with wild-type controls and that H1 depletion impairs DNA methylation and changes of histone marks at promoter regions necessary for efficiently silencing pluripotency gene Oct4 during stem cell differentiation and embryogenesis. In summary, we demonstrate that H1 plays a critical role in pluripotent stem cell differentiation, and our results suggest that H1 and chromatin compaction may mediate pluripotent stem cell differentiation through epigenetic repression of the pluripotency genes

Aggregation of an anionic porphyrin with chiral metal complexes and the competitive binding influences of a surfactant and a polyelectrolyte

The non-covalent interactions of chiral metal complexes with the achiral 5,10,15,20-tetrakis (4-sulonatophenyl) porphyrin (H(4)TTPS(2-)) have been investigated by UV-vis and circular dichroism (CD) spectra. The results show that under acidic environments, only the chiral complex cations ([CoBr(NH(3))(en)(2)](+), [Co(en)(3)](3+), [Ru(phen)(3)](2+)) could interact with H(4)TTPS(2-) to form chiral aggregates, accompanied with the metal-centered chirality information transferred to the formed J-aggregates. However, the chiral complex anion ([Co(edta)](-)) does not cause the self-assembly process. The competitive binding interactions between an achiral water-soluble cationic surfactant (N-hexadecyltrimethyl ammonium chloride, CTAC) and a cationic polyelectrolyte (polyallylamine, PAA) with the chiral metal complex H(4)TTPS(2-) J-aggregates, respectively, were also investigated. It was found that chiral-symmetry-breaking phenomena occur in the cationic surfactant induced event. In the case of a cationic polyelectrolyte, it could change the conformational flexibility of the H(4)TTPS(2-) aggregates. These results may lead us to understand the possible mechanism of the supramolecular self-assembly process by the non-covalent interactions.National Natural Science Foundation of China[20973136, 20877099, 20972183]; State Key Laboratory of Natural and Biomimetic Drugs[20080208]; Hunan High Education Research Fund[06C068]; GUCAS (A B); Ministry of Science and Technology of China[2008AA100801]; Guangdong Province[2010B090300031]; CAS[2010B090300031

Genome-wide profiling of H1 linker histone variants in mouse embryonic stem cells

H1 linker histone facilitates the formation of higher order chromatin structure and is essential for mammalian development. Mice have 11 H1 variants which are differentially regulated and conserved in human. Previous research indicates that H1 regulates the expression of specific genes in mouse embryonic stem cells (ESCs). However, whether individual variants have distinct functions and how H1 participates in gene regulation remain elusive. An investigation of the precise localization of individual H1 variants in vivo would facilitate the elucidation of mechanisms underlying chromatin compaction regulated gene expression, while it has been extremely difficult due to the lacking of specific antibodies toward H1 variants. In this dissertation, I have generated a knock-in system in ESCs and shown that the N-terminally tagged H1 proteins are functionally interchangeable to their endogenous counterparts in vivo. H1d and H1c are depleted from GC- and gene-rich regions and active promoters, inversely correlated with H3K4me3, but positively correlated with H3K9me3 and associated with characteristic sequence features. Surprisingly, both H1d and H1c are significantly enriched at major satellites, which display increased nucleosome spacing compared with bulk chromatin. While also depleted at active promoters and enriched at major satellites, overexpressed H10 displays differential binding patterns in specific repetitive sequences compared with H1d and H1c. Depletion of H1c, H1d ,and H1e causes pericentric chromocenter clustering and de-repression of major satellites. Collectively, these results integrate the localization of an understudied type of chromatin proteins, namely the H1 variants, into the epigenome map of mouse ESCs, and demonstrate significant changes at pericentric heterochromatin upon depletion of this epigenetic mark.Ph.D

Salumycin, a New Pyrazolequinone from a Streptomyces albus J1074 Mutant Strain

Heterocyclic natural products with various bioactivities play significant roles in pharmaceuticals. Here, we isolated a heterocyclic compound salumycin (1) from a Streptomyces albus J1074 mutant strain. The structure of (1) was elucidated via single-crystal X-ray diffraction, mass spectrometry (MS), fourier transform infrared spectrometer (FTIR), and nuclear magnetic resonance (NMR) data analysis. Salumycin (1) contained a novel pyrazolequinone ring, which had never been previously reported in a natural product. Salumycin (1) exhibited moderate 2,2′-diphenyl-1-picrylhydrazyl (DPPH)-radical scavenging activity (EC50 = 46.3 ± 2.2 μM) compared with tert-butylhydroquinone (EC50 = 4.7 ± 0.3 μM). This study provides a new example of discovering novel natural products from bacteria

Study on Material Properties of Magnesium Oxide Carbonized Prestressed Pipe Piles

Traditional PHC pipe pile in foundation engineering consumes high energy and has insufficient durability. A magnesium oxide carbonization test block is a new type of environmental protection block which bases on activated magnesium oxide cementation technology. The use of CO2 carbonation technology allows reactive magnesia to react to form basic magnesium carbonate to increase the compressive strength and durability of the block. Three kinds of different magnesium oxide powders were subjected to pressure test and determined the key technical parameters, such as optimal raw materials, sample preparation methods, carbonization environment and technology, and optimized design of pipe pile concrete material system