Effects of free proline accumulation in petunias under drought stress

Petunias (Petunia hybrida cv. 'Mitchell') accumulate free proline (Pro) under drought-stress conditions. It is therefore believed that Pro acts as an osmoprotectant in plants subjected to drought conditions. Petunia plants were transformed by Delta(1)-pyrroline-5-carboxylate synthetase genes (AtP5CS from Arabidopsis thaliana L. or OsP5CS from Oryza sativa L.). The transgenic plants accumulated Pro and their drought tolerance was tested. The Pro content amounted to 0.57-1.01% of the total amino acids in the transgenic plants, or 1.5-2.6 times that in wild-type plants grown under normal conditions. The transgenic plant lines tolerated 14 d of drought stress, which confirms that both P5CS transgenes had full functionality. Exogenous L-Pro treatment caused the plants to accumulate Pro; plants treated with 5 mM L-Pro accumulated up to 18 times more free Pro than untreated plants. Exogenous L-Pro restricted the growth of wild-type petunias more than that of Arabidopsis plants. The capacity for free Pro accumulation might depend on the plant species. The growth of petunia plants was influenced not only by the Pro concentration in the plants, but by the ratio of the Pro content to the total amino acids, because the growth of the transgenic petunia plants appeared normal.     

Air-liquid interface promotes invasive growth of laryngeal squamous cell carcinoma with or without hypoxia

Air-liquid interface (ALI) is a microenvironment of aerodigestive tract. In our previous study, ALI promoted invasive growth of laryngeal squamous cell carcinoma (SCC); but its mechanism was unclear. Hypoxia is also related to cancer spread. Here we show that ALI with or without hypoxia accelerated invasive growth of laryngeal SCC cells, using collagen gel invasion assay. Submerged condition (SMC) without ALI did not induce the invasion with or without hypoxia. ALI enhanced expression of the following growth-, invasion-, and motility-related molecules in the cells with or without hypoxia more greatly than SMC: c-Met, Ras, mitogen-activated protein kinase cascade proteins (Raf-1, MEK-1, and ERK-1/2), matrix metalloproteinase-1, and filamin A. The data indicate that ALI promotes invasive growth of SCC cells by enhancement of the invasive growth-related molecules above, through mechanisms that differ from hypoxia, suggesting that ALI microenvironment should be taken into account for the study of cancer biology.     

Structure and thermal history dependent phase behavior of hydrated synthetic sphingomyelin analogue: 1,2-dimyristamido-1,2-deoxyphosphatidylcholine

The physical properties of hydrated multilamellar sample of 1,2-dimyristamido-1,2-deoxyphosphatidylcholine (DDPC) were investigated by means of differential scanning calorimetry (DSC), static X-ray diffraction, and simultaneous DSC and X-ray diffraction. The DDPC is a synthetic sphingomyelin analogue and has two amide bonds in its hydrophobic parts. This paper reports on metastable phase behavior of the hydrated DDPC sample. By cooling from a chain-melted state at the rates of greater than 4 degrees C min(-1), hydrated DDPC bilayers form a metastable gel phase. In the gel phase, the hydrophobic chains are tilted with respect to the bilayer normal, as like the gel phase of glycero-phosphatidylcholines. By heating, the metastable gel phase is transformed in to a stable phase associated with an exothermic heat event at 18.3 degrees C (DeltaH=14.6 kJ mol(-1)) and then the stable phase is transformed into a liquid-crystalline phase at 25.6 degrees C (DeltaH=42 kJ mol(-1)). The incubation at 17 degrees C for more than 1 h also induces the formation of the stable phase. In the stable phase, the hydrophobic chains are packed into highly ordered crystal-like structure. However, the X-ray diffraction pattern of the stable phase suggested that the entire DDPC molecules do not form a two-dimensional molecular ordered lattice, differing from normal subgel phase of glycero-phosphatidylcholines. The structure and phase behavior of DDPC revealed by the present study are discussed from the viewpoint of hydrogen bonds.     

Design and expression of cysteine-bearing hydrophobic polypeptides and their self-assembling properties with bacteriochlorophyll a derivatives as a mimic of bacterial photosynthetic antenna complexes. Effect of steric confinement and orientation of the polypeptides on the pigment/polypeptide assembly process

A series of cysteine-bearing hydrophobic polypeptides analogous to a light-harvesting one betapolypeptide (LH1beta) from the LH1 complex from the purple photosynthetic bacterium, Rhodobacter sphaeroides, was synthesized using an Escherichia coli expression system. The cysteine was placed in the C- or N-terminal regions of the polypeptide to investigate the influence of steric confinement and orientation of the polypeptides via disulfide linkages as they were self-assembled with zinc-substituted bacteriochlorophyll a ([Zn]-BChl a). The polypeptides were expressed as water-soluble fusion proteins with maltose-binding protein (MBP). The fusion proteins formed a subunit-type complex with the [Zn]-BChl a in an n-octyl-beta-d-glucopyranoside (OG) micellar solution regardless of the cross-links or the cleavage of the cysteines, judging from absorption, CD, and fluorescence spectra. Following treatment with trypsin, the polypeptides were detached from the MBP portion. Such trypsin-digested polypeptides formed a subunit-type LH complex at 25 degrees C, which also showed that the disulfide linkage was not crucial for the subunit formation. When a polypeptide having cysteine on the C-terminus was assembled at 4 degrees C, the Qy absorption band was remarkably red-shifted to approximately 836 nm, suggesting that the cleavage of the large MBP portion liberates the polypeptides to form the progressive type of complex similar to LH1-type complex. The trypsin-treated polypeptides bearing cysteines in both terminal regions, which are randomly cross-linked, did not form the LH1-type complex under oxidative conditions but did form the complex under reductive conditions. This observation suggests that the polypeptide orientation strongly influences the LH1-type complex formation. The progressive assembly from the subunit to the holo-LH1-type complex following cleavage of MBP portion in a lipid bilayer is also briefly discussed.     

Anti-citrullinated collagen type I antibody is a target of autoimmunity in rheumatoid arthritis

Rheumatoid arthritis (RA) is one of the most common autoimmune diseases, but its autoimmune mechanisms are not clearly understood. Recently, anti-citrullinated peptide antibodies have been specifically observed in sera of RA patients. Furthermore, we identified RA-susceptible variant in a gene encoding citrullinating enzyme, peptidylarginine deiminase type 4 (PADI4). Therefore, we hypothesized that proteins which are modified in RA synovium by PADI4 act as autoantigens. Subsequently, we obtained human collagen type I (huCI) as one of the autoantigens using a RA synoviocyte cDNA library by immunoscreening. We also investigated that the levels of anti-citrullinated huCI were significantly higher in RA patient sera than in normal control sera with high specificity (99%) and positively correlated with the levels of anti-cyclic citrullinated peptide (anti-CCP) antibodies. We concluded that huCI is a novel substrate protein of PADIs and that citrullinated huCI is a candidate autoantigen of RA.     

Pnlip encoding pancreatic lipase is possible candidate for obesity QTL in the OLETF rat

The Otsuka Long-Evans Tokushima Fatty (OLETF) rat exhibits polygenic obesity, and one of quantitative trait loci (QTLs) responsible for a susceptibility to obesity in the OLETF, Nidd6/of, has been mapped to the approximately 10-cM genomic region between D1Rat166 and D1Rat90 on chromosome 1 in (OLETF x normal) F2 intercross. In this study, we have attempted to identify the causal gene for the Nidd6/of QTL. A Nidd6/of congenic strain, constructed by introgressing the OLETF allele on the mapped Nidd6/of region in the normal F344 rat strain, confirmed the existence of the Nidd6/of as obesity QTL. The Nidd6/of region was refined to a approximately 2.3-cM genomic region between D1Rat225 and D1Rat90, using informative recombinants selected from (Nidd6/of congenic x F344) F1 x Nidd6/of congenic backcross progenies. Among 46 genes located within the approximately 2.3-cM region, pancreatic lipase gene, Pnlip, was regarded as the most prominent and physiologically relevant positional candidate for the Nidd6/of QTL. We found that Pnlip possesses an OLETF allele-specific increase of mRNA levels in the pancreas, and that the OLETF allele is longer in variable number of tandem repeat (VNTR) within the 5'-flanking region than normal alleles. We further showed that the Nidd6/of QTL completely cosegregates with Pnlip VNTR in the informative recombinants from (Nidd6/of congenic x F344) F1 x Nidd6/of congenic backcross progenies. These results suggest that Pnlip is possible candidate for the Nidd6/of QTL.     

Gastric inhibitory polypeptide modulates adiposity and fat oxidation under diminished insulin action

Gut hormone gastric inhibitory polypeptide (GIP) stimulates insulin secretion from pancreatic β-cells upon ingestion of nutrients. Inhibition of GIP signaling prevents the onset of obesity and consequent insulin resistance induced by high-fat diet. In this study, we investigated the role of GIP in accumulation of triglycerides into adipocytes and in fat oxidation peripherally using insulin receptor substrate (IRS)-1-deficient mice and revealed that IRS-1^−/−GIPR^−/− mice exhibited both reduced adiposity and ameliorated insulin resistance. Furthermore, increased gene expression of CD36 and UCP2 in liver, and increased expression and enzyme activity of 3-hydroxyacyl-CoA dehydrogenase in skeletal muscle of IRS-1^−/−GIPR^−/− mice might contribute to the lower respiratory quotient and the higher fat oxidation in light phase. These results suggest that GIP plays a crucial role in switching from fat oxidation to fat accumulation under the diminished insulin action as a potential target for secondary prevention of insulin resistance.     

Differences in growth trajectory and strategy of two sympatric congeneric species in an Indonesian floodplain forest

Whole-plant development trajectories and sapling leaf displays were compared for two sympatric congeneric species, Pterospermum diversifolium and P. javanicum, in a tropical floodplain forest in East Kalimantan, Indonesia. We assessed their growth strategies and developed hypotheses for their coexistence within the community. Pterospermum diversifolium retains a monoaxial growth habit that promotes quick stem elongation; thus, it is taller when branches are initiated than is P. javanicum. The species differed significantly in height growth and total crown expansion per unit increment of biomass: monoaxial P. diversifolium saplings devote more effort to stem elongation, whereas branched P. javanicum saplings devote more effort to branch expansion. Monoaxial P. diversifolium sustained more severe self-shading than P. javanicum. The sapling growth strategy of P. diversifolium appears to be dynamic, emphasizing the opportunistic use of light following a disturbance, whereas that of P. javanicum appears to be static, optimizing leaf display for current light conditions. The advantages of these strategies depend on context, and the two species may coexist within a community by adopting different regeneration niches based on differing understory light conditions: P. diversifolium is favored over P. javanicum at high light levels, but the opposite is true at low light levels.