Analysis of the APX, PGD1 and R1G1B constitutive gene promoters in various organs over three homozygous generations of transgenic rice plants

We have previously characterized the constitutively active promoters of the APX, PGD1 and R1G1B genes in rice (Park et al. 2010 in J Exp Bot 61:2459–2467). To have potential crop biotechnology applications, gene promoters must be stably active over many generations. In our current study, we report our further detailed analysis of the APX, PGD1 and R1G1B gene promoters in various organs and tissues of transgenic rice plants for three (T_3–5) homozygous generations. The copy numbers in 37 transgenic lines that harbor promoter:gfp constructs were determined and promoter activities were measured by real-time qPCR. Analysis of the 37 lines revealed that 15 contained a single copy of one of the three promoter:gfp chimeric constructs. The promoter activity levels were generally higher in multi-copy lines, whereas variations in these levels over the T_3–5 generations studied were observed to be smaller in single-copy than in multi-copy lines. The three promoters were further found to be highly active in the whole plant body at both the vegetative and reproductive stages of plant growth, with the exception of the APX in the ovary and R1G1B in the pistil and filaments where zero or very low levels of activity were detected. Of note, the spatial activities of the PGD1 promoter were found to be strikingly similar to those of the ZmUbi1, a widely used constitutive promoter. Our comparison of promoter activities between T₃, T₄ and T₅ plants revealed that the APX, PGD1 and R1G1B promoters maintained their activities at comparable levels in leaves and roots over three homozygous generations and are therefore potentially viable alternative promoters for crop biotechnology applications.     

Fossil human remains from Bolomor Cave (Valencia, Spain)

Systematic excavations carried out since 1989 at Bolomor Cave have led to the recovery of four Pleistocene human fossil remains, consisting of a fibular fragment, two isolated teeth, and a nearly complete adult parietal bone. All of these specimens date to the late Middle and early Late Pleistocene (MIS 7-5e). The fibular fragment shows thick cortical bone, an archaic feature found in non-modern (i.e. non-Homo sapiens) members of the genus Homo. Among the dental remains, the lack of a midtrigonid crest in the M₁ represents a departure from the morphology reported for the majority of Neandertal specimens, while the large dimensions and pronounced shoveling of the marginal ridges in the C¹ are similar to other European Middle and late Pleistocene fossils. The parietal bone is very thick, with dimensions that generally fall above Neandertal fossils and resemble more closely the Middle Pleistocene Atapuerca (SH) adult specimens. Based on the presence of archaic features, all the fossils from Bolomor are attributed to the Neandertal evolutionary lineage.     

2-Styrylindolium based fluorescent probes visualize neurofibrillary tangles in Alzheimer’s disease

We evaluated 2-styrylindolium derivatives (6–11) as novel and selective probes for neurofibrillary tangles (NFTs) on brain sections of AD patients. The staining experiments indicated that these compounds may bind selectively to NFTs in the presence of ß-amyloid (Aß) plaques. Cell free binding assays confirmed that 2-[2-[4-(1-pyrrolidinyl)phenyl]ethenyl]-1,3,3-trimethyl-3H-indolium iodide (9) and 2-[2-[4-(diethylamino)phenyl]ethenyl]-1-butyl-3,3-dimethyl-3H-indolium iodide (11) display excellent affinities to Tau-aggregates (IC₅₀ values of 5.1 and 1.4 nM, respectively) in the displacement of Thiazin Red R. These probes have good solubility in distilled water and low or no cytotoxicity in zebrafish embryo and liver hepatocellular carcinoma cell assays.     

Transgenic rice grains expressing a heterologous ρ-hydroxyphenylpyruvate dioxygenase shift tocopherol synthesis from the γ to the α isoform without increasing absolute tocopherol levels

We generated transgenic rice plants overexpressing Arabidopsis thaliana ρ-hydroxyphenylpyruvate dioxygenase (HPPD), which catalyzes the first committed step in vitamin E biosynthesis. Transgenic grains accumulated marginally higher levels of total tocochromanols than controls, reflecting a small increase in absolute tocotrienol synthesis (but no change in the relative abundance of the α and γ isoforms). In contrast, there was no change in the absolute tocopherol level, but a significant shift from the γ to the α isoform. These data confirm HPPD is not rate limiting, and that increasing flux through the early pathway reveals downstream bottlenecks that act as metabolic tipping points.     

DLK1(PREF1) is a negative regulator of adipogenesis in CD105⁺/CD90⁺/CD34⁺/CD31⁻/FABP4⁻ adipose-derived stromal cells from subcutaneous abdominal fat pats of adult women

The main physiological function of adipose-derived stromal/progenitor cells (ASC) is to differentiate into adipocytes. ASC are most likely localized at perivascular sites in adipose tissues and retain the capacity to differentiate into multiple cell types. Although cell surface markers for ASC have been described, there is no complete consensus on the antigen expression pattern that will precisely define these cells. DLK1(PREF1) is an established marker for mouse adipocyte progenitors which inhibits adipogenesis. This suggests that DLK1(PREF1) could be a useful marker to characterize human ASC. The DLK1(PREF1) status of human ASC is however unknown. In the present study we isolated ASC from the heterogeneous stromal vascular fraction of subcutaneous abdominal fat pats of adult women. These cells were selected by their plastic adherence and expanded to passage 5. The ASC were characterized as relatively homogenous cell population with the capacity to differentiate in vitro into adipocytes, chondrocytes, and osteoblasts and the immunophenotype CD105?/CD90?/CD34?/CD31?/FABP4?. The ASC were positive for DLK1(PREF1) which was well expressed in proliferating and density arrested cells but downregulated in the course of adipogenic differentiation. To investigate whether DLK1(PREF1) plays a role in the regulation of adipogenesis in these cells RNAi-mediated knockdown experiments were conducted. Knockdown of DLK1(PREF1) in differentiating ASC resulted in a significant increase of the expression of the adipogenic key regulator PPARγ2 and of the terminal adipogenic differentiation marker FABP4. We conclude that DLK1(PREF1) is well expressed in human ASC and acts as a negative regulator of adipogenesis. Moreover, DLK1(PREF1) could be a functional marker contributing to the characterization of human ASC.     

Association of genetic variation in the NR1H4 gene, encoding the nuclear bile acid receptor FXR, with inflammatory bowel disease

ABSTRACT: BACKGROUND: Pathogenesis of inflammatory bowel diseases (IBD), ulcerative colitis (UC) and Crohn's disease (CD), involves interaction between environmental factors and inappropriate immune responses in the intestine of genetically predisposed individuals. Bile acids and their nuclear receptor, FXR, regulate inflammatory responses and barrier function in the intestinal tract. METHODS: We studied the association of five variants (rs3863377, rs7138843, rs56163822, rs35724, rs10860603) of the NR1H4 gene encoding FXR with IBD. 1138 individuals (591 non-IBD, 203 UC, 344 CD) were genotyped for five NR1H4 genetic variants with TaqMan SNP Genotyping Assays. RESULTS: We observed that the NR1H4 SNP rs3863377 is significantly less frequent in IBD cases than in non-IBD controls (allele frequencies: P = 0.004; wild-type vs. SNP carrier genotype frequencies: P = 0.008), whereas the variant rs56163822 is less prevalent in non-IBD controls (allele frequencies: P = 0.027; wild-type vs. SNP carrier genotype frequencies: P = 0.035). The global haplotype distribution between IBD and control patients was significantly different (P = 0.003). This also held true for the comparison between non-IBD and UC groups (P = 0.004), but not for the comparison between non-IBD and CD groups (P = 0.079). CONCLUSIONS: We show that genetic variation in FXR is associated with IBD, further emphasizing the link between bile acid signaling and intestinal inflammation.     

Comprehensive proteome analysis in Cenococcum geophilum Fr. as a tool to discover drought-related proteins

Cenococcum geophilum is a widely distributed ectomycorrhizal fungus potentially playing a significant role in resistance and resilience mechanisms of its tree hosts exposed to drought stress. In this study, we performed a large scale protein analysis in pure cultures of C. geophilum in order to gain first global insights into the proteome assembly of this fungus. Using 1-D gel electrophoresis coupled with ESI-MS/MS, we indentified 638 unique proteins. Most of these proteins were related to the metabolic and cellular processes, and the transport machinery of cells. In a second step, we examined the influence of water deprivation on the proteome of C. geophilum pure cultures at three time points of gradually imposed drought. The results indicated that 12 proteins were differentially abundant in mycelia subjected to drought compared to controls. The induced responses in C. geophilum point towards regulation of osmotic stress, maintainance of cell integrity, and counteracting increased levels of reactive oxygen species formed during water deprivation.