Methylation of cytosines in the dinucleotide CpG has been shown to suppress transcription of a number of tissue-specific genes, yet the precise mechanism is not fully understood. The vertebrate globin genes were among the first examples in which an inverse correlation was shown between CpG methylation and transcription. We studied the methylation pattern of the 235-bp -globin gene promoter in genomic DNA from primary chicken erythroid cells using the sodium bisulfite conversion technique and found all CpGs in the promoter to be methylated in erythroid cells from adult chickens in which the -globin gene is silent but unmethylated in 5-day (primitive) embryonic red cells in which the gene is transcribed. To elucidate further the mechanism of methylation-induced silencing, an expression construct consisting of 235 bp of 5 promoter sequence of the -globin gene along with a strong 5 erythroid enhancer driving a chloramphenicol acetyltransferase reporter gene, -CAT, was transfected into primary avian erythroid cells derived from 5-day embryos. Methylation of just the 235-bp -globin gene promoter fragment at every CpG resulted in a 20-to 30-fold inhibition of transcription, and this effect was not overridden by the presence of potent erythroid-specific enhancers. The ability of the 235-bp -globin gene promoter to bind to a DNA Methyl Cytosine binding Protein Complex (MeCPC) was tested in electrophoretic mobility shift assays utilizing primary avian erythroid cell nuclear extract. The results were that fully methylated but not unmethylated 235-bp -globin gene promoter fragment could compete efficiently for MeCPC binding. These results are a direct demonstration that site-specific methylation of a globin gene promoter at the exact CpGs that are methylated in vivo can silence transcription in homologous primary erythroid cells. Further, these data implicate binding of MeCPC to the promoter in the mechanism of silencing.Methylation of cytosine residues in the dinucleotide CpG is the most common postsynthetic eukaryotic DNA modification. Since the reports of an inverse correlation between DNA methylation and expression of vertebrate ␤-type globin genes (1-3), a large body of evidence relating DNA methylation to gene expression has accumulated (4, 5). At the same time, the absence of detectable DNA methylation in some eukaryotes such as Drosophila (6) and Saccharomyces cerevisiae (7) has raised doubts about its role in normal development and tissue-specific gene expression. However a study by Li et al. (8) showing abnormal development and embryonic lethality in transgenic mice expressing decreased but not completely absent DNA methyl transferase activity following knockout of the DNA methyl transferase gene lends strong support to a critical role for DNA methylation in developmental gene regulation. Recently, a similar critical function of DNA methylation in plant development has been demonstrated by Ronemus et al. (9). However, a direct demonstration of the role of DNA methylation in suppressing transcription of a...

The transcriptional regulatory network governing the establishment of retinal neuron diversity is not well delineated. We report experimental results suggesting proneural gene neurogenin3 (ngn3) participating in this regulatory network. Retinal expression of chick ngn3 was confined to early neurogenesis. Overexpression of ngn3 in chick retina reduced cell proliferation and expanded the population of ganglion cells into the territory normally occupied by amacrine cells. Ngn3 overexpression altered the expression of a number of regulatory genes, including ash1, ath3, ath5, chx10, neuroD, ngn1, ngn2, and NSCL1. Early gene ngn1 was induced, but ash1, ngn2, ath3, and chx10, whose expressions persist through later phases of neurogenesis, were down-regulated. Expression of ath5 was up-regulated at the locale corresponding to young ganglion cells, but was down-regulated at the locale corresponding to progenitor cells. These results suggest that ngn3 regulates retinal neurogenesis by inducing regulatory genes for early-born neurons and repressing those for later-born cells.

These markers will be useful to study population ecology and population differentiation among M. charantia species and its related species.

The diverse types of neurons and Müller glia in the vertebrate retina are believed to arise from common progenitor cells. To better understand how neural diversity is achieved during retinal neurogenesis, we examined the function of ash1, a proneural bHLH gene expressed in progenitor cells throughout retinal neurogenesis. Published studies using retinal explant culture derived from knockout mice concluded that ash1 is required for the production of late-born neurons, including bipolar cells. In this study, gain-of-function experiments were carried out in ovo in embryonic chick retina. In the developing chick retina, expression of ash1 temporally overlapped with, but spatially differed from, the expression of ngn2, also a proneural gene expressed in progenitor cells throughout retinal neurogenesis. Retrovirus-driven overexpression of ash1 in the developing chick retina decreased the progenitor population (BrdU+ or expressing ngn2), expanded the amacrine population (AP2α+ or Pax6+), and reduced bipolar (chx10 mRNA+) and Müller glial (vimentin+) populations. Photoreceptor deficiency occurred after the completion of neurogenesis. The number of ganglion cells, which are born first during retinal neurogenesis, remained unchanged. Similar overexpression of ngn2 did not produce discernible changes in retinal neurogenesis, nor in ash1 expression. These results suggest that ash1 promotes the production of amacrine cells and thus may participate in a regulatory network governing neural diversity in the chick retina.

ZnS:F quantum dots (QDs) capping with and without L-cys were synthesized by a solid-state method at low temperature, and the influence of L-cys on the properties of ZnS:F QDs were investigated. The crystal structure, surface morphology and luminescent properties of the samples were analyzed by X-ray diffractometer (XRD), transmission electron microscope (TEM), fourier transform infrared (FTIR), photoluminescence spectrometer (PL) and ultraviolet-visible spectrometer (UV-Vis). The results showed that all samples had a zinc blende structure with particle size in the range of 2-6 nm. The emission intensity was significantly enhanced after capping with L-cys, and the strongest luminescence was obtained when the ratio of L-cys/ZnS:F was 0.8:1, and was about 2.5 times of that of ZnS:F QDs. The capping of L-cys increased the grain size of ZnS:F QDs and their water solubility.