Preliminary genetic characterization of two lineages of black rats (Rattus rattus sensu lato) in Japan, with evidence for introgression at several localities

We conducted a pilot survey of genetic diversity among 37 karyotyped individuals of the black rat Rattus rattus (sensu lato) from six localities on the Japanese Islands, using complete gene sequences of mitochondrial cytochrome b (cyt b) and nuclear interphotoreceptor retinoid binding protein (IRBP). Our sampling included two previously documented karyotypic groups: 'Oceanian' with 2n = 38 and 'Asian' with 2n = 42. Cyt b sequences for most individuals clustered according to their karyotypic groups, with an average between-group divergence of 3.8%. One exception was that individuals from Kagoshima (Kyushu Island) showed 'Asian' karyotypes combined with a cyt b haplotype that differed by a single nucleotide substitution from the haplotype of the 'Oceanian' karyotypic group. Six IRBP haplotypes were identified. They belonged to three distinct IRBP lineages (I-III), with an average inter-lineage divergence of 1%. Among homozygous individuals, these lineages showed good association with the karyotypic groups: IRBP lineage I occurred only with 'Oceanian' karyotypes, while IRBP lineages II and III both occurred with 'Asian' karyotypes. Individuals from Kagoshima all possessed IRBP of 'Asian' lineages, despite the presence of an 'Oceanian' mitochondrial type. The Chichijima population (Ogasawara Islands) featured exclusively 'Asian' karyotypes and cyt b sequences, but various combinations of all three IRBP lineages. The Kagoshima and Chichijima populations thus provide strong evidence of viable hybridization and genetic introgression between the two karyotypic groups, but with variable genetic outcomes. Our results demonstrate the potential of combined analysis of karyotypes and mitochondrial and nuclear gene sequences to elucidate the complex dispersal and population history of the black rat.     

Distribution of branched-chain fatty acid in the skin surface lipid of laboratory animals

1. The distribution of the branched-chain fatty acid (BCFA) was studied in skin surface lipids of laboratory animals (rat, mouse, hamster and rabbit) and the experimental animal for the study of the metabolic fate of BCFA was chosen. 2. The monoester fraction resistant to microbial degradation was the index of which fatty acids were identified and the compositions were analyzed by capillary gas chromatography and mass spectrometry (GC-MS). 3. The contents of monoester fractions in rat, mouse, hamster and rabbit were 78.9, 15.9, 30.4 and 45.6% of the total skin surface lipid, respectively. 4. BCFAs were exclusively identified to be either iso- or anteiso-series type by means of mass spectrometry. 5. The sum of iso- and anteiso-acid was the highest in hamster comprising 53% in male and 38% in female of the monoester acid. 6. Sex related differences of BCFA concentration were not so evident in the other three species. 7. BCFA contents in the monoester fractions were as follows: rat 32%, mouse 25% and rabbit 3%. 8. Concentrations of iso-series fatty acid were consistently higher than that of anteiso-series type in all animals studied. 9. Abundance of 2-hydroxy fatty acid in the rabbit monoester fraction was noted for the first time. 10. Suitability of these laboratory animals for the study of BCFA metabolism was discussed.     

S100A10 protein expression is associated with oxaliplatin sensitivity in human colorectal cancer cells

Background

C-reactive protein (CRP) is a predictor of cardiovascular risk. It circulates as a pentameric protein in plasma. Recently, a potential dissociation mechanism from the disc-shaped pentameric CRP (pCRP) into single monomers (monomeric or mCRP) has been described. It has been shown that mCRP has strong pro-inflammatory effects on monocytes. To further define the role of mCRP in determining monocyte phenotype, the effects of CRP isoforms on THP-1 protein expression profiles were determined. The hypothesis to be tested was that mCRP induces specific changes in the protein expression profile of THP-1 cells that differ from that of pCRP.

Methods

Protein cell lysates from control and mCRP, pCRP or LPS-treated THP-1 cells were displayed using 2-dimensional SDS PAGE and compared. Differentially expressed proteins were identified by MALDI-TOF MS and confirmed by Western blotting.

Results

mCRP significantly up-regulates ubiquitin-activating enzyme E1, a member of the ubiquitin-proteasome system in THP-1 monocytes. Furthermore, HSP 70, alpha-actinin-4 (ACTN4) and alpha-enolase/enolase 1 were upregulated. The proteomic profile of LPS and pCRP treated monocytes differ significantly from that of mCRP.

Conclusion

The data obtained in this study support the hypothesis that isoform-specific effects of CRP may differentially regulate the phenotype of monocytes.     

Role of Rho-kinase signaling and endothelial dysfunction in modulating blood flow distribution in pulmonary hypertension

Rho-kinase-mediated vasoconstriction and endothelial dysfunction are considered two primary instigators of pulmonary arterial hypertension (PAH). However, their contribution to the adverse changes in pulmonary blood flow distribution associated with PAH has not been addressed. This study utilizes synchrotron radiation microangiography to assess the specific role, and contribution of, Rho-kinase-mediated vasoconstriction and endothelial dysfunction in PAH. Male adult Sprague-Dawley rats were injected with saline (Cont-rats) or monocrotaline (MCT-rats) 3 wk before microangiography was performed on the left lung. We assessed dynamic changes in vessel internal diameter (ID) in response to 1) the Rho-kinase inhibitor fasudil (10 mg/kg iv); or 2) ACh (3 μg · kg?1 · min?1), sodium nitroprusside (SNP, 5 μg · kg?1 · min?1), and N(ω)-nitro-l-arginine methyl ester (l-NAME, 50 mg/kg iv). We observed that MCT-rats had fewer vessels of the microcirculation compared with Cont-rats. The fundamental result of this study is that fasudil improved pulmonary blood flow distribution and reduced pulmonary pressure in PAH rats, not only by dilating already-perfused vessels (ID > 100 μm), but also by restoring blood flow to vessels that had previously been constricted closed (ID < 100 μm). Endothelium-dependent vasodilation was impaired in MCT-rats primarily in vessels with an ID < 200 μm. Moreover the vasoconstrictor response to l-NAME was accentuated in MCT-rats, but only in the 200- to 300-μm vessels. These results highlight the importance of Rho-kinase-mediated control and endothelial control of pulmonary vascular tone in PAH. Indeed, an effective therapeutic strategy for treating PAH should target both the smooth muscle Rho-kinase and endothelial pathways.     

Cytohesin-2/ARNO,through Its Interaction with Focal Adhesion Adaptor Protein Paxillin,Regulates Preadipocyte Migration via the Downstream Activation of Arf6

The formation of primitive adipose tissue is the initial process in adipose tissue development followed by the migration of preadipocytes into adipocyte clusters. Comparatively little is known about the molecular mechanism controlling preadipocyte migration. Here, we show that cytohesin-2, the guanine-nucleotide exchange factor for the Arf family GTP-binding proteins, regulates migration of mouse preadipocyte 3T3-L1 cells through Arf6. SecinH3, a specific inhibitor of the cytohesin family, markedly inhibits migration of 3T3-L1 cells. 3T3-L1 cells express cytohesin-2 and cytohesin-3, and knockdown of cytohesin-2 with its small interfering RNA effectively decreases cell migration. Cytohesin-2 preferentially acts upstream of Arf6 in this signaling pathway. Furthermore, we find that the focal adhesion protein paxillin forms a complex with cytohesin-2. Paxillin colocalizes with cytohesin-2 at the leading edges of migrating cells. This interaction is mediated by the LIM2 domain of paxillin and the isolated polybasic region of cytohesin-2. Importantly, migration is inhibited by expression of the constructs containing these regions. These results suggest that cytohesin-2, through a previously unexplored complex formation with paxillin, regulates preadipocyte migration and that paxillin plays a previously unknown role as a scaffold protein of Arf guanine-nucleotide exchange factor.     

Community structures of actively growing bacteria shift along a north-south transect in the western North Pacific

Bacterial community structures and their activities in the ocean are tightly coupled with organic matter fluxes and thus control ocean biogeochemical cycles. Bromodeoxyuridine (BrdU), halogenated nucleoside and thymidine analogue, has been recently used to monitor actively growing bacteria (AGB) in natural environments. We labelled DNA of proliferating cells in seawater bacterial assemblages with BrdU and determined community structures of the bacteria that were possible key species in mediating biochemical reactions in the ocean. Surface seawater samples were collected along a north-south transect in the North Pacific in October 2003 and subjected to BrdU magnetic beads immunocapture and PCR-DGGE (BUMP-DGGE) analysis. Change of BrdU-incorporated community structures reflected the change of water masses along a north-south transect from subarctic to subtropical gyres in the North Pacific. We identified 25 bands referred to AGB as BrdU-incorporated phylotypes, belonging to Alphaproteobacteria (5 bands), Betaproteobacteria (1 band), Gammaproteobacteria (4 bands), Cytophaga-Flavobacterium-Bacteroides (CFB) group bacteria (5 bands), Gram-positive bacteria (6 bands), and Cyanobacteria (4 bands). BrdU-incorporated phylotypes belonging to Vibrionales , Alteromonadales and Gram-positive bacteria appeared only at sampling stations in a subtropical gyre, while those belonging to Roseobacter -related bacteria and CFB group bacteria appeared at the stations in both subarctic and subtropical gyres. Our result revealed phylogenetic affiliation of AGB and their dynamic change along with north-south environmental gradients in open oceans. Different species of AGB utilize different amount and kinds of substrates, which can affect the change of organic matter fluxes along transect.     

Gene expression profiles of necrosis and apoptosis induced by 5-fluoro-2'-deoxyuridine

5-Fluoro-2'-deoxyuridine (FUdR), a potent anticancer agent, exerts its effects by inhibiting thymidylate synthase, an essential machinery for DNA synthesis in cell proliferation. Also, cell death is caused by FUdR, primarily due to an imbalance in the nucleotide pool resulting from this enzyme inhibition. We have investigated the cancer cell death induced by FUdR, focusing on its molecular mechanisms. Using mouse mammary tumor FM3A cell lines, the original clone F28-7 and its variant F28-7-A cells, we previously reported an interesting observation that FUdR induces a necrotic morphology in F28-7, but induces, in contrast, an apoptotic morphology in F28-7-A cells. In the present study, to understand the molecular mechanisms underlying these differential cell deaths, i.e., necrosis and apoptosis, we investigated the gene expression changes occurring in these processes. Using the cDNA microarray technology, we found 215 genes being expressed differentially in the necrosis and apoptosis. Further analysis revealed differences between these cell lines in terms of the expressions of both a cluster of heat shock protein (HSP)-related genes and a cluster of apoptosis-related genes. Notably, inhibition of HSP90 in F28-7 cells caused a shift from the FUdR-induced necrosis into apoptosis. These findings are expected to lead to a better understanding of this anticancer drug FUdR for its molecular mechanisms and also of the general biological issue, necrosis and apoptosis.     

The genetics of domestication of yardlong bean, Vigna unguiculata (L.) Walp. ssp. unguiculata cv.-gr. sesquipedalis

Background and Aims

The genetics of domestication of yardlong bean [Vigna unguiculata (L.) Walp. ssp. unguiculata cv.-gr. sesquipedalis] is of particular interest because the genome of this legume has experienced divergent domestication. Initially, cowpea was domesticated from wild cowpea in Africa; in Asia a vegetable form of cowpea, yardlong bean, subsequently evolved from cowpea. Information on the genetics of domestication-related traits would be useful for yardlong bean and cowpea breeding programmes, as well as comparative genome study among members of the genus Vigna. The objectives of this study were to identify quantitative trait loci (QTLs) for domestication-related traits in yardlong bean and compare them with previously reported QTLs in closely related Vigna.

Methods

Two linkage maps were developed from BC₁F₁ and F₂ populations from the cross between yardlong bean (V. unguiculata ssp. unguiculata cv.-gr. sesquipedalis) accession JP81610 and wild cowpea (V. unguiculata ssp. unguiculata var. spontanea) accession TVnu457. Using these linkage maps, QTLs for 24 domestication-related traits were analysed and mapped. QTLs were detected for traits related to seed, pod, stem and leaf.

Key Results

Most traits were controlled by between one and 11 QTLs. QTLs for domestication-related traits show co-location on several narrow genomic regions on almost all linkage groups (LGs), but especially on LGs 3, 7, 8 and 11. Major QTLs for sizes of seed, pod, stem and leaf were principally located on LG7. Pleiotropy or close linkage of genes for the traits is suggested in these chromosome regions.

Conclusions

This is the first report of QTLs for domestication-related traits in yardlong bean. The results provide a foundation for marker-assisted selection of domestication-related QTLs in yardlong bean and enhance understanding of domestication in the genus Vigna.