Mitochondrial genome of the Komodo dragon: efficient sequencing method with reptile-oriented primers and novel gene rearrangements.

The mitochondrial genome of the Komodo dragon (Varanus komodoensis) was nearly completely sequenced, except for two highly repetitive noncoding regions. An efficient sequencing method for squamate mitochondrial genomes was established by combining the long polymerase chain reaction (PCR) technology and a set of reptile-oriented primers designed for nested PCR amplifications. It was found that the mitochondrial genome had novel gene arrangements in which genes from NADH dehydrogenase subunit 6 to proline tRNA were extensively shuffled with duplicate control regions. These control regions had 99% sequence similarity over 700 bp. Although snake mitochondrial genomes are also known to possess duplicate control regions with nearly identical sequences, the location of the second control region suggested independent occurrence of the duplication on lineages leading to snakes and the Komodo dragon. Another feature of the mitochondrial genome of the Komodo dragon was the considerable number of tandem repeats, including sequences with a strong secondary structure, as a possible site for the slipped-strand mispairing in replication. These observations are consistent with hypotheses that tandem duplications via the slipped-strand mispairing may induce mitochondrial gene rearrangements and may serve to maintain similar copies of the control region.     

Nicotine affects mineralized nodule formation by the human osteosarcoma cell line Saos-2

Several in vitro and in vivo studies have indicated that tobacco smoking may be an important risk factor for the development and severity of inflammatory periodontal disease. In the present study, we examined the effect of nicotine on cell proliferation, alkaline phosphatase (ALPase) activity, mineralized nodule formation, and the expression of extracellular matrix proteins in the human osteosarcoma cell line Saos-2. The cells were cultured with Dulbecco's modified Eagle medium containing 10% fetal bovine serum with 0, 10(-4) M, and 10(-3) M nicotine for up to 14 days. Mineralized nodule formation was examined by alizarin red staining, and the calcium content in mineralized nodules was determined using a calcium E-test kit. The expression of extracellular matrix proteins was estimated by determining the levels of their mRNAs using the real-time polymerase chain reaction. Mineralized nodule formation and calcium content in mineralized nodules were remarkably suppressed by nicotine on days 10 and 14 of culture, respectively. ALPase activity as well as type I collagen and osteopontin expression also decreased in the presence of nicotine after 5, 10, and 14 days of culture, respectively. By contrast, the amount of bone sialoprotein increased during 14 days of culture with nicotine. These results suggest that nicotine suppresses osteogenesis through a decrease in ALPase and type I collagen production by osteoblasts.     

Potential for enlarging DNA memory: the validity of experimental operations of scaled-up nested primer molecular memory

DNA is an attractive memory unit because of its immense information density. Here, we describe a memory model made of DNA, called Nested Primer Molecular Memory (NPMM). NPMM consists of many DNA strands, and each DNA strand consists of two areas: a data area and a data address area. When the address of target data is specified, only the target data can be extracted from NPMM. In this paper, we evaluate the validity of the basic operations of NPMM and then discuss the feasibility of scaled-up NPMM through some laboratory experiments. In the latter, we deal with scaled-up NPMM simulated by the Concentration Scaling method.     

Molecular characterization of ENU mouse mutagenesis and archives

The large-scale mouse mutagenesis with ENU has provided forward-genetic resources for functional genomics. The frozen sperm archive of ENU-mutagenized generation-1 (G1) mice could also provide a "mutant mouse library" that allows us to conduct reverse genetics in any particular target genes. We have archived frozen sperm as well as genomic DNA from 9224 G1 mice. By genome-wide screening of 63 target loci covering a sum of 197 Mbp of the mouse genome, a total of 148 ENU-induced mutations have been directly identified. The sites of mutations were primarily identified by temperature gradient capillary electrophoresis method followed by direct sequencing. The molecular characterization revealed that all the identified mutations were point mutations and mostly independent events except a few cases of redundant mutations. The base-substitution spectra in this study were different from those of the phenotype-based mutagenesis. The ENU-based gene-driven mutagenesis in the mouse now becomes feasible and practical.     

Foraminiferal ecological zonation along a Brazilian mangrove transect: Diversity, morphotypes and the influence of subaerial exposure time

Two foraminiferal associations comprising only arenaceous species define two distinct environments in a 340 m-long mangrove transect at Cardoso Island, Trapandé Bay (Cananéia-Iguape estuarine system, SP, Brazil). The “lower muddy flat” (LMF), from the outer mangrove fringe inwards towards land (100 m), is positioned in the lower plain between 0.04 and 0.23 m above the mean sea level (msl), and remains subaerially exposed between 48.5 and 65.6% of the time. This environment is characterized by higher foraminiferal diversity and evenness (McIntosh's D = 0.54 ± 0.21 and Pielou's E = 0.68 ± 0.25, respectively) and is dominated by Arenoparrella mexicana and Trochammina inflata, and to a lesser extent by Ammotium directum and Textularia earlandi. The mangrove plant of this segment is a Rhizophoretum with average height of 8.4 ± 1.2 m. The sediment is characterized by higher concentration of organic matter (93.5 ± 32.3 g dm⁻³) and metals (e.g. V = 53.4 ± 21.8 ppm and Zn = 46.4 ± 21.3 ppm). The “upper sandy flat” (USF), 240 m wide along the transect, is positioned in the upper plain between 0.28 and 0.89 m above the msl, and remains subaerially exposed between 69.7 and 98.5% of the time. This environment is characterized by a lower diversity and evenness (D = 0.33 ± 0.17 and E = 0.49 ± 0.20, respectively). The association is dominated by species T. inflata and Miliammina fusca. The Rhizophoretum exhibits a lower average height of 3.6 ± 0.6 m. The sediment is poorer in organic matter (39.3 ± 15.0 g dm⁻³) and metals (e.g. V = 13.0 ± 6.8 ppm and Zn = 6.9 ± 3.7 ppm). Whereas “elongate” tests (uniserial, biserial and planospiral followed by a uniserial portion) are restricted to the LMF, “spiraled” species dominate the USF. Subaerial exposure time seems to exert a primary influence on species distribution, in addition to salinity and sediment type. Species may be adapted to different exposure times, a factor dependent on their position on the intertidal zone and the tidal regime, which should be taken into account in relative sea level reconstructions based on intertidal foraminifera. These patterns have important implications for studies investigating the ecology and paleoecology of foraminifera and subtle fluctuations in relative sea level during the Quaternary.     

Overexpression of CNP in chondrocytes rescues achondroplasia through a MAPK-dependent pathway

Achondroplasia is the most common genetic form of human dwarfism, for which there is presently no effective therapy. C-type natriuretic peptide (CNP) is a newly identified molecule that regulates endochondral bone growth through GC-B, a subtype of particulate guanylyl cyclase. Here we show that targeted overexpression of CNP in chondrocytes counteracts dwarfism in a mouse model of achondroplasia with activated fibroblast growth factor receptor 3 (FGFR-3) in the cartilage. CNP prevented the shortening of achondroplastic bones by correcting the decreased extracellular matrix synthesis in the growth plate through inhibition of the MAPK pathway of FGF signaling. CNP had no effect on the STAT-1 pathway of FGF signaling that mediates the decreased proliferation and the delayed differentiation of achondroplastic chondrocytes. These results demonstrate that activation of the CNP-GC-B system in endochondral bone formation constitutes a new therapeutic strategy for human achondroplasia.