A new orphan member of the nuclear hormone receptor superfamily that interacts with a subset of retinoic acid response elements.

We have identified and characterized a new orphan member of the nuclear hormone receptor superfamily, called MB67, which is predominantly expressed in liver. MB67 binds and transactivates the retinoic acid response elements that control expression of the retinoic acid receptor beta 2 and alcohol dehydrogenase 3 genes, both of which consist of a direct repeat hexamers related to the consensus AGGTCA, separated by 5 bp. MB67 binds these elements as a heterodimer with the 9-cis-retinoic acid receptor, RXR. However, MB67 does not bind or activate other retinoic acid response elements with alternative hexamer arrangements or any of several other wild-type and synthetic hormone response elements examined. The transactivation of retinoic acid response elements by MB67 is weaker than that conferred by the retinoic acid receptors but does not require the presence of all-trans retinoic acid, 9-cis-retinoic acid, or any exogenously added ligand. We propose that MB67 plays an important role in the complex network of proteins that govern response to retinoic acid and its metabolites.     

Cyclin-dependent kinase 4 signaling acts as a molecular switch between syngenic differentiation and neural transdifferentiation in human mesenchymal stem cells

Multipotent mesenchymal stem/stromal cells (MSCs) are capable of differentiating into a variety of cell types from different germ layers. However, the molecular and biochemical mechanisms underlying the transdifferentiation of MSCs into specific cell types still need to be elucidated. In this study, we unexpectedly found that treatment of human adipose- and bone marrow-derived MSCs with cyclin-dependent kinase (CDK) inhibitor, in particular CDK4 inhibitor, selectively led to transdifferentiation into neural cells with a high frequency. Specifically, targeted inhibition of CDK4 expression using recombinant adenovial shRNA induced the neural transdifferentiation of human MSCs. However, the inhibition of CDK4 activity attenuated the syngenic differentiation of human adipose-derived MSCs. Importantly, the forced regulation of CDK4 activity showed reciprocal reversibility between neural differentiation and dedifferentiation of human MSCs. Together, these results provide novel molecular evidence underlying the neural transdifferentiation of human MSCs; in addition, CDK4 signaling appears to act as a molecular switch from syngenic differentiation to neural transdifferentiation of human MSCs.     

Effects of Dietary Linseed Oil and Propionate Precursors on Ruminal Microbial Community,Composition, and Diversity in Yanbian Yellow Cattle

The rumen microbial ecosystem is a complex system where rumen fermentation processes involve interactions among microorganisms. There are important relationships between diet and the ruminal bacterial composition. Thus, we investigated the ruminal fermentation characteristics and compared ruminal bacterial communities using tag amplicon pyrosequencing analysis in Yanbian yellow steers, which were fed linseed oil (LO) and propionate precursors. We used eight ruminally cannulated Yanbian yellow steers (510 ± 5.8 kg) in a replicated 4 × 4 Latin square design with four dietary treatments. Steers were fed a basal diet that comprised 80% concentrate and 20% rice straw (DM basis, CON). The CON diet was supplemented with LO at 4%. The LO diet was also supplemented with 2% dl-malate or 2% fumarate as ruminal precursors of propionate. Dietary supplementation with LO and propionate precursors increased ruminal pH, total volatile fatty acid concentrations, and the molar proportion of propionate. The most abundant bacterial operational taxonomic units in the rumen were related to dietary treatments. Bacteroidetes dominated the ruminal bacterial community and the genus Prevotella was highly represented when steers were fed LO plus propionate precursors. However, with the CON and LO diet plus malate or fumarate, Firmicutes was the most abundant phylum and the genus Ruminococcus was predominant. In summary, supplementing the diets of ruminants with a moderate level of LO plus propionate precursors modified the ruminal fermentation pattern. The most positive responses to LO and propionate precursors supplementation were in the phyla Bacteriodetes and Firmicutes, and in the genus Ruminococcus and Prevotella. Thus, diets containing LO plus malate or fumarate have significant effects on the composition of the rumen microbial community.     

An Integrated Approach for Analysis of the DNA Damage Response in Mammalian Cells: NUCLEOTIDE EXCISION REPAIR,DNA DAMAGE CHECKPOINT,AND APOPTOSIS*

DNA damage by UV and UV-mimetic agents elicits a set of inter-related responses in mammalian cells, including DNA repair, DNA damage checkpoints, and apoptosis. Conventionally, these responses are analyzed separately using different methodologies. Here we describe a unified approach that is capable of quantifying all three responses in parallel using lysates from the same population of cells. We show that a highly sensitive in vivo excision repair assay is capable of detecting nucleotide excision repair of a wide spectrum of DNA lesions (UV damage, chemical carcinogens, and chemotherapeutic drugs) within minutes of damage induction. This method therefore allows for a real-time measure of nucleotide excision repair activity that can be monitored in conjunction with other components of the DNA damage response, including DNA damage checkpoint and apoptotic signaling. This approach therefore provides a convenient and reliable platform for simultaneously examining multiple aspects of the DNA damage response in a single population of cells that can be applied for a diverse array of carcinogenic and chemotherapeutic agents.     

Intestinal Helminthic Infections Diagnosed by Colonoscopy in a Regional Hospital during 2001-2008

The present study investigated characteristics of 24 parasite infection cases detected during colonoscopy in a regional hospital from January 2001 to December 2008. Sixteen patients were confirmed with Trichuris trichiura infection, 6 patients were with Ascaris lumbricoides infection, 1 patient with Enterobius vermicularis infection, and 1 patient with Anisakis infection. Among them, 7 patients (43.8%) were asymptomatic. Colonoscopy findings were normal in 18 patients (75.0%). Among the patients with T. trichiura infection, colonoscopy showed several erosions in 2 patients (8.3%) and non-specific inflammation of the affected segment of the colon in 3 patients (12.5%). In 1 patient with anisakiasis, colonoscopy revealed a markedly swollen colonic wall. Stool examinations were performed before treatment in 7 patients (29.2%) and were all negative for parasite eggs or worms. These results suggest that colonoscopy is a useful diagnostic approach for parasitic infections even for asymptomatic patients and for patients with negative stool examinations.     

Suprafenacine,an Indazole-Hydrazide Agent,Targets Cancer Cells Through Microtubule Destabilization

Microtubules are a highly validated target in cancer therapy. However, the clinical development of tubulin binding agents (TBA) has been hampered by toxicity and chemoresistance issues and has necessitated the search for new TBAs. Here, we report the identification of a novel cell permeable, tubulin-destabilizing molecule - 4,5,6,7-tetrahydro-1H-indazole-3-carboxylic acid [1p-tolyl-meth-(E)-ylidene]-hydrazide (termed as Suprafenacine, SRF). SRF, identified by in silico screening of annotated chemical libraries, was shown to bind microtubules at the colchicine-binding site and inhibit polymerization. This led to G₂/M cell cycle arrest and cell death via a mitochondria-mediated apoptotic pathway. Cell death was preceded by loss of mitochondrial membrane potential, JNK - mediated phosphorylation of Bcl-2 and Bad, and activation of caspase-3. Intriguingly, SRF was found to selectively inhibit cancer cell proliferation and was effective against drug-resistant cancer cells by virtue of its ability to bypass the multidrug resistance transporter P-glycoprotein. Taken together, our results suggest that SRF has potential as a chemotherapeutic agent for cancer treatment and provides an alternate scaffold for the development of improved anti-cancer agents.     

Cooperative activation of the ATR checkpoint kinase by TopBP1 and damaged DNA

TopBP1, acting in concert with DNA containing bulky base lesions, stimulates ATR kinase activity under physiologically relevant reaction conditions. Here, we analyze the roles of the three components in ATR activation: DNA, base damage and TopBP1. We show that base adducts caused by a potent carcinogen, benzo[a]pyrene diol epoxide (BPDE), constitute a strong signal for TopBP1-dependent ATR kinase activity on Chk1 and p53. We find that the C-terminus of TopBP1 binds preferentially to damaged DNA and is sufficient to mediate damaged DNA-dependent ATR activation in a manner similar to full-length TopBP1. Significantly, we find that stimulation of ATR by BPDE-damaged DNA exhibits strong dependence on the length of DNA, with essentially no stimulation with fragments of 0.2 kb and reaching maximum stimulation with 2 kb fragments. Moreover, TopBP1 shows preferential binding to longer DNA fragments and, in contrast to previous biochemical studies, TopBP1 binding is completely independent of DNA ends. We find that TopBP1 binds to circular and linear DNAs with comparable affinities and that these DNA forms elicit the same level of TopBP1-dependent ATR activation. Taken together, these findings suggest a cooperative activation mechanism for the ATR checkpoint kinase by TopBP1 and damaged DNA.     

Spot-fixed fin digging behavior in foraging of the benthophagous maiden goby, Pterogobius virgo (Perciformes: Gobiidae)

 Several patterns of feeding behaviors have been documented in benthophagous fishes. The foraging behavior of the maiden goby, Pterogobius virgo, was studied at Kurahashi Island in the Seto Inland Sea, Japan. Pterogobius virgo foraged mostly on polychaetes by volume from among several available prey items by digging in the sandy bottom. The digging behavior comprised swing of only pectoral fins or of both pectoral fins and body. Pectoral fin swing exposed the cryptic prey within the bottom, and fins and body swing exposed the prey and washed the sediment away. The swings were repeatedly and continuously conducted at a site during the daytime, making a pit several centimeters deep in which the fish was located. After the prey was exposed, the fish immediately and rapidly picked up the prey. Polychaetes were abundant prey in the sediment, occurring in the layer 3–5 cm deep from the bottom surface in the study area. In this goby, spot-fixed fin digging, the first documentation of feeding habits in gobies, may be effective for feeding on the most valuable prey, i.e., polychaetes, which may be otherwise unavailable for this fish. Received: April 24, 2001 / Revised: April 26, 2002 / Accepted: May 7, 2002