Immune-related microRNAs are abundant in breast milk exosomes

Breast milk is a complex liquid rich in immunological components that affect the development of the infant's immune system. Exosomes are membranous vesicles of endocytic origin that are found in various body fluids and that can mediate intercellular communication. MicroRNAs (miRNAs), a well-defined group of non-coding small RNAs, are packaged inside exosomes in human breast milk. Here, we identified 602 unique miRNAs originating from 452 miRNA precursors (pre-miRNAs) in human breast milk exosomes using deep sequencing technology. We found that, out of 87 well-characterized immune-related pre-miRNAs, 59 (67.82%) are presented and enriched in breast milk exosomes (P < 10(-16), χ(2) test). In addition, compared with exogenous synthetic miRNAs, these endogenous immune-related miRNAs are more resistant to relatively harsh conditions. It is, therefore, tempting to speculate that these exosomal miRNAs are transferred from the mother's milk to the infant via the digestive tract, and that they play a critical role in the development of the infant immune system.     

Genetic Variability of PRNP in Chinese Indigenous Goats

Polymorphism of the prion protein gene (PRNP) is usually associated with scrapie susceptibility or resistance. To determine the variability of PRNP in Chinese indigenous goat breeds, we isolated genomic DNA from goat blood and amplified and sequenced the coding region of the gene. We identified 10 polymorphic sites that gave rise to 28 haplotypes. Clear frequency differences were found between northern and southern breeds and confirmed by genetic distance analysis, except for the Tangshan dairy goat. Phylogeographic analysis supported the idea that northern and southern breeds might be considered separate clusters, except for the Tangshan dairy goat. The finding of significant differences in allele distribution in northern and southern goats, especially if involved in modulating resistance/susceptibility, needs to be carefully considered for the feasibility of selection plans for resistance to scrapie.     

Cultured gill epithelia as models for the freshwater fish gill

We review recent progress in the development of models for the freshwater teleost gill based on reconstructed flat epithelia grown on permeable filter supports in primary culture. Methods are available for single-seeded insert (SSI) preparations consisting of pavement cells (PVCs) only from trout and tilapia, and double-seeded insert (DSI) preparations from trout, containing both PVCs (85%) and mitochondria-rich cells (MRCs, 15%), as in the intact gill. While there are some quantitative differences, both SSI and DSI epithelia manifest electrical and passive permeability characteristics typical of intact gills and representative of very tight epithelia. Both preparations withstand apical freshwater exposure, exhibiting large increases in transepithelial resistance (TER), negative transepithelial potential (TEP), and low rates of ion loss, but there is only a small active apical-to-basolateral "influx" of Cl(-) (and not of Na(+)). Responses to various hormonal treatments are described (thyroid hormone T3, prolactin, and cortisol). Cortisol has the most marked effects, stimulating Na(+),K(+)-ATPase activity and promoting active Na(+) and Cl(-) influxes in DSI preparations, and raising TER and reducing passive ion effluxes in both epithelia via reductions in paracellular permeability. Experiments using DSI epithelia lacking Na(+) uptake demonstrate that both NH(3) and NH(4)(+) diffusion occur, but are not large enough to account for normal rates of branchial ammonia excretion, suggesting that Na(+)-linked carrier-mediated processes are important for ammonia excretion in vivo. Future research goals are suggested.     

PNAS-4 expression and its relationship to p53 in colorectal cancer

PNAS-4 is a novel pro-apoptotic protein activated during the early response to DNA damage; however, the molecular mechanisms and pathways regulating PNAS-4 expression in tumors are not well understood. We hypothesized that PNAS-4 is a p53 down-stream target gene and designed this study. We searched online for putative p53-binding sites in the entire PNAS-4 gene and did not find any corresponding information. In HCT116 colon cancer cells, after being transfected with small interfering RNA to silence p53, the expressions of PNAS-4 and other known p53 target gene (Apaf1, Bax, Fas and Dr5) were determined by real-time PCR. We found that PNAS-4 was up-regulated while Apaf1, Bax, Fas and Dr5 were down-regulated. We then examined the expression of PNAS-4 and p53 mutation in colorectal cancer patients. PNAS-4 expressed both in colorectal cancers and normal tissues, but compared with paired control, PNAS-4 was up-regulated in cancers (P = 0.018). PNAS-4 overexpression ratios were correlated to the p53 mutant status (P = 0.001). The mean PNAS-4 expression levels of p53 mutant homozygote group and heterozygote group were higher than that of p53 wild type group (P = 0.013). The expression ratios of PNAS-4 (every sample in relative to its paired normal mucosa) were different between negative lymph node metastasis (66% up-regulated, 34% down-regulated) and positive metastasis (42% up-regulated, 58% down-regulated). Taken together, these findings suggested that PNAS-4 was not a p53 target, but overexpression of PNAS-4 was correlated to p53 inactivity in colorectal cancer.     

Identification of a movement protein of the tenuivirus rice stripe virus

Rice stripe virus (RSV) is the type member of the genus Tenuivirus. RSV has four single-stranded RNAs and causes severe disease in rice fields in different parts of China. To date, no reports have described how RSV spreads within host plants or the viral and/or host factor(s) required for tenuivirus movement. We investigated functions of six RSV-encoded proteins using trans-complementation experiments and biolistic bombardment. We demonstrate that NSvc4, encoded by RSV RNA4, supports the intercellular trafficking of a movement-deficient Potato virus X in Nicotiana benthamiana leaves. We also determined that upon biolistic bombardment or agroinfiltration, NSvc4:enhanced green fluorescent protein (eGFP) fusion proteins localize predominantly near or within the walls of onion and tobacco epidermal cells. In addition, the NSvc4:eGFP fusion protein can move from initially bombarded cells to neighboring cells in Nicotiana benthamiana leaves. Immunocytochemistry using tissue sections from RSV-infected rice leaves and an RSV NSvc4-specific antibody showed that the NSvc4 protein accumulated in walls of RSV-infected leaf cells. Gel retardation assays revealed that the NSvc4 protein interacts with single-stranded RNA in vitro, a common feature of many reported plant viral movement proteins (MPs). RSV NSvc4 failed to interact with the RSV nucleocapsid protein using yeast two-hybrid assays. Taken together, our data indicate that RSV NSvc4 is likely an MP of the virus. This is the first report describing a tenuivirus MP.