Investigating a simplified method for noninvasive genetic sampling in East African mammals using silica dried scat swabs

Swabbing scat has proved to be an effective noninvasive method to collect DNA from mammals in the field. Previously, this method has relied on preservative liquids or freezing to preserve the DNA collected on swabs. In this study, we determine the effectiveness of using silica to simply dry the swab in field as an alternative way to prevent DNA degredation. Four species were included in the study; reticulated giraffe, impala, fringe‐eared oryx, and lion. Swabs were taken at multiple time points for giraffe and impala scat samples, with the lion and oryx sampled opportunistically. Mitochondrial DNA was successfully amplified and sequenced from scat swabs from all species; however, effectiveness varied between species, with 81.8% amplification success rate from swabs taken from impala scat compared to 25% amplification success rate in giraffe. This variation in success rate was overcome by taking multiple swabs, thus increasing the probability of a successful amplification. The true merit of this method is in its simplicity and cheapness; no preservative liquids were required to be brought into the field, at no stage in the 2 weeks of field sampling were samples frozen, and no commercial kits were used for DNA extraction.     

Arabidopsis HAP2 (GCS1) is a sperm-specific gene required for pollen tube guidance and fertilization

In flowering plants, sperm cells develop in the pollen cytoplasm and are transported through floral tissues to an ovule by a pollen tube, a highly polarized cellular extension. After targeting an ovule, the pollen tube bursts, releasing two sperm that fertilize an egg and a central cell. Here, we identified the gene encoding Arabidopsis HAP2, demonstrating that it is allelic to GCS1. HAP2 is expressed only in the haploid sperm and is required for efficient pollen tube guidance to ovules. We identified an insertion (hap2-1) that disrupts the C-terminal portion of the protein and tags mutant pollen grains with the beta-glucuronidase reporter. By monitoring reporter expression, we showed that hap2-1 does not diminish pollen tube length in vitro or in the pistil, but it reduces ovule targeting by twofold. In addition, we show that the hap2 sperm that are delivered to ovules fail to initiate fertilization. HAP2 is predicted to encode a protein with an N-terminal secretion signal, a single transmembrane domain and a C-terminal histidine-rich domain. These results point to a dual role for HAP2, functioning in both pollen tube guidance and in fertilization. Moreover, our findings suggest that sperm, long considered to be passive cargo, are involved in directing the pollen tube to its target.     

Phosphorylation of EZH2 by CDK1 and CDK2

Histone H3 lysine 27 trimethylation (H3K27me3) catalyzed by the enzymatic subunit EZH2 in the Polycomb repressive complex 2 (PRC2) is essential for cells to ‘memorize’ gene expression patterns through cell divisions and plays an important role in establishing and maintaining cell identity during development. However, how the epigenetic mark is inherited through cell generations remains poorly understood. Recently, we and others demonstrate that CDK1 and CDK2 phosphorylate EZH2 at threonine 350 (T350) and that T350 phosphorylation is important for the binding of EZH2 to PRC2 recruiters, such as noncoding RNAs (ncRNAs) HOTAIR and XIST, and for the effective recruitment of PRC2 to EZH2 target loci in cells. These findings imply that phosphorylation of EZH2 by CDK1 and CDK2 may provide cells a mechanism that enhances EZH2 function during S and G2 phases of the cell cycle, thereby ensuring K27me3 on de novo synthesized H3 incorporated in nascent nucleosomes before sister chromosomes are divided into two daughter cells. Additionally, a potential role of T350 phosphorylation of EZH2 in differing EZH2 from its homolog EZH1 in catalyzing H3K27me3 as well as the interplay between phosphorylation at T350 and other residues (e.g. phosphorylation by p38 at threonine 372 (T372)) in governing EZH2 activity in proliferating versus non-dividing cells are also discussed. Together, CDK phosphorylation of EZH2 at T350 may represent a key regulatory mechanism of EZH2 function that is essential for the maintenance of H3K27me3 marks through cell divisions.     

Vitamin A cycle byproducts impede dark adaptation

Impaired dark adaptation (DA), a defect in the ability to adjust to dimly lit settings, is a universal hallmark of aging. However, the mechanisms responsible for impaired DA are poorly understood. Vitamin A byproducts, such as vitamin A dimers, are small molecules that form in the retina during the vitamin A cycle. We show that later in life, in the human eye, these byproducts reach levels commensurate with those of vitamin A. In mice, selectively inhibiting the formation of these byproducts, with the investigational drug C20D₃-vitamin A, results in faster DA. In contrast, acutely increasing these ocular byproducts through exogenous delivery leads to slower DA, with otherwise preserved retinal function and morphology. Our findings reveal that vitamin A cycle byproducts alone are sufficient to cause delays in DA and suggest that they may contribute to universal age-related DA impairment. Our data further indicate that the age-related decline in DA may be tractable to pharmacological intervention by C20D₃-vitamin A.     

Identification and characterization of an essential gene in Listeria monocytogenes using an inducible gene expression system

The Listeria monocytogenes gene lmo1594 is a homolog of the Bacillus subtilis cell division gene ezrA. EzrA is a negative regulator of FtsZ ring formation, which is required for efficient cell division as it regulates the frequency and position of Z-rings in the cell and prevents aberrant polar cell division. Previously identified as a putative high pressure (HP) resistance mechanism; conferring enhanced barotolerance when heterologously expressed against an Escherichia coli background; the aim of the current study was to investigate whether lmo1594 plays a role in listerial barotolerance. When the creation of a deletion mutant proved unsuccessful, the role of lmo1594 was addressed by creating a conditional knockout mutant which demonstrated that the gene is in fact essential for cell survival and growth in L. monocytogenes. In order to investigate the effect of lmo1594 on barotolerance, the gene was over-expressed. The over-expression of lmo1594 increased survival levels in L. monocytogenes treated at 300 MPa, but survival levels similar to those of the wild-type strain were observed when treated at a higher pressure (≥400 MPa). In conclusion, this study reveals for the first time that lmo1594 is absolutely essential for listerial cell growth and survival, and also plays an important role in listerial barotolerance.     

Synaptonemal complex analysis of hybrid cattle. I. Pachytene substaging and the normal full bloods

Meiotic chromosome pairing abnormalities in full-blood and hybrid Bos taurus and B. indicus cattle have been surveyed by electron microscopy of pachytene synaptonemal complex spreads. In this paper the full-blood spreads are described in detail, including the use of XY and nucleolar morphology and other measured parameters for pachytene substaging. Pairing abnormalities were observed in up to 9% of the full-blood spreads. Most of these pairing abnormalities (83%) occurred in early-pachytene spreads, suggesting that the mechanism of synaptic adjustment may operate in cattle.     

HIV-infected macrophages resist efficient NK cell-mediated killing while preserving inflammatory cytokine responses

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