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Background: MicroRNAs (miRNAs) are 21~25-nucleotides (nt) long and interact with mRNAs to trigger either translational repression or RNA cleavage through RNA interference (RNAi), depending on the degree of complementarity with the target mRNAs. Our recent study has shown that HIV-1 nef dsRNA from AIDS patients who are long-term non-progressors (LTNPs) inhibited the transcription of HIV-1. Results: Here, we show the possibility that nef-derived miRNAs are produced in HIV-1 persistently infected cells. Furthermore, nef short hairpin RNA (shRNA) that corresponded to a predicted nef miRNA (~25 nt, miR-N367) can block HIV-1 Nef expression in vitro and the suppression by shRNA/miR-N367 would be related with low viremia in an LTNP (15-2-2). In the 15-2-2 model mice, the weight loss, which may be rendered by nef was also inhibited by shRNA/miR-N367 corresponding to suppression of nef expression in vivo. Conclusions: These data suggest that nef/U3 miRNAs produced in HIV-1-infected cells may suppress both Nef function and HIV-1 virulence through the RNAi pathway. [ABSTRACT FROM AUTHOR]

Background: The efficiency of cellulolytic enzymes is important in industrial biorefinery processes, including biofuel production. Chemical methods, such as alkali pretreatment, have been extensively studied and demonstrated as effective for breaking recalcitrant lignocellulose structures. However, these methods have a detrimental effect on the environment. In addition, utilization of these chemicals requires alkali- or acid-resistant equipment and a neutralization step., Results: Here, a radical generator based on non-thermal atmospheric pressure plasma technology was developed and tested to determine whether oxygen-radical pretreatment enhances cellulolytic activity. Our results showed that the viscosity of carboxymethyl cellulose (CMC) solutions was reduced in a time-dependent manner by oxygen-radical pretreatment using the radical generator. Compared with non-pretreated CMC, oxygen-radical pretreatment of CMC significantly increased the production of reducing sugars in culture supernatant containing various cellulases from Phanerochaete chrysosporium . The production of reducing sugar from oxygen-radical-pretreated CMC by commercially available cellobiohydrolases I and II was 1.7- and 1.6-fold higher, respectively, than those from non-pretreated and oxygen-gas-pretreated CMC. Moreover, the amount of reducing sugar from oxygen-radical-pretreated wheat straw was 1.8-fold larger than those from non-pretreated and oxygen-gas-pretreated wheat straw., Conclusions: Oxygen-radical pretreatment of CMC and wheat straw enhanced the degradation of cellulose by reducing- and non-reducing-end cellulases in the supernatant of a culture of the white-rot fungus P. chrysosporium . These findings indicated that oxygen-radical pretreatment of plant biomass offers great promise for improvements in lignocellulose-deconstruction processes.

Background: Exosomes or extracellular vesicles have the potential as a diagnostic marker for various diseases including cancer. In order to identify novel exosomal markers for prostate cancer (PC), we performed proteomic analysis of exosomes isolated from PC cell lines and examined the usefulness of the marker in patients., Methods: Exosomes isolated by differential centrifugation from the culture medium of androgen-dependent LNCaP prostate cancer cell line and its sublines of partially androgen-independent C4, androgen-independent C4-2 and bone metastatic C4-2B were subjected to iTRAQ-based proteomic analysis. Exosomes were also isolated by immunocapture and separated by size exclusion chromatography and density gradient centrifugation. Protein expression was determined by Western blot analysis. GGT activity was measured using a fluorescent probe, γ-glutamyl hydroxymethyl rhodamine green (gGlu-HMRG). Immunohistochemical analysis of tissues was performed using anti-GGT1 antibody., Results: Among proteins upregulated in C4-2 and C4-2B cells than in LNCaP cells, we focused on gamma-glutamyltransferase 1 (GGT1), a cell-surface enzyme that regulates the catabolism of extracellular glutathione. The levels of both GGT1 large and small subunits were elevated in exosomes isolated from C4-2 and C4-2B cells by differential centrifugation and by immunocapture with anti-CD9 or -prostate-specific membrane antigen (PSMA) antibody. In cell lysates and exosomes, GGT1 expression correlated with GGT activity. Size exclusion chromatography of human serum demonstrated the presence of GGT activity and GGT1 subunits in fractions positive for CD9. Density gradient centrifugation revealed the co-presence of GGT1 subunits with CD9 in exosomes isolated by differential centrifugation from human serum. Since GGT activity correlated with GGT1 expression in serum exosomes isolated by differential centrifugation, we measured serum exosomal GGT activity in patients. Unexpectedly, we found that serum exosomal GGT activity was significantly higher in PC patients than in benign prostatic hyperplasia (BPH) patients. In support of this finding, immunohistochemical analysis showed increased GGT1 expression in PC tissues compared with BPH tissues., Conclusions: Our results suggest that serum exosomal GGT activity could be a useful biomarker for PC.

Background: Senescence-accelerated mice (SAM) are a series of mouse strains originally derived from unexpected crosses between AKR/J and unknown mice, from which phenotypically distinct senescence-prone (SAMP) and -resistant (SAMR) inbred strains were subsequently established. Although SAMP strains have been widely used for aging research focusing on their short life spans and various age-related phenotypes, such as immune dysfunction, osteoporosis, and brain atrophy, the responsible gene mutations have not yet been fully elucidated., Results: To identify mutations specific to SAMP strains, we performed whole exome sequencing of 6 SAMP and 3 SAMR strains. This analysis revealed 32,019 to 38,925 single-nucleotide variants in the coding region of each SAM strain. We detected Ogg1 p.R304W and Mbd4 p.D129N deleterious mutations in all 6 of the SAMP strains but not in the SAMR or AKR/J strains. Moreover, we extracted 31 SAMP-specific novel deleterious mutations. In all SAMP strains except SAMP8, we detected a p.R473W missense mutation in the Ldb3 gene, which has been associated with myofibrillar myopathy. In 3 SAMP strains (SAMP3, SAMP10, and SAMP11), we identified a p.R167C missense mutation in the Prx gene, in which mutations causing hereditary motor and sensory neuropathy (Dejerine-Sottas syndrome) have been identified. In SAMP6 we detected a p.S540fs frame-shift mutation in the Il4ra gene, a mutation potentially causative of ulcerative colitis and osteoporosis., Conclusions: Our data indicate that different combinations of mutations in disease-causing genes may be responsible for the various phenotypes of SAMP strains.