Intervarietal differences in some metabolic functions associated with protein accumulation in rice grains

Summary Total nitrogen, amino nitrogen, glutamic acid dehydrogenase (GDH) activity and incorporation of ³H-uridine and ¹⁴C-amino acids into RNA and proteins, respectively, were compared in the developing grains of three high-protein stocks (IR-480-5-9, GMPR-51 and Erythroceros) and a high-yielding, medium-protein cultivar IR-8. The above parameters were also independently studied in the developing grains of IR-8 grown at 0, 60 and 120 kgN/ha. In addition, mobilization of nitrogen from flag leaf during kernel development was compared in a separate experiment. Higher protein concentration, both in high-protein stocks and in IR-8 grown at 120 kgN/ha, was associated with increased levels of: soluble amino nitrogen, GDH activity, ³H-uridine and ¹⁴C-amino acid incorporation. Significant variation was found among the high protein stocks in mobilization of nitrogen from flag leaf.Research partly supported by International Atomic Energy Agency Research Contract 1035     

Genetic engineering approaches to improve the bioavailability and the level of iron in rice grains

Iron deficiency is the most widespread micronutrient deficiency world-wide. A major cause is the poor absorption of iron from cereal and legume-based diets high in phytic acid. We have explored three approaches for increasing the amount of iron absorbed from rice-based meals. We first introduced a ferritin gene from Phaseolus vulgaris into rice grains, increasing their iron content up to two-fold. To increase iron bioavailability, we introduced a thermotolerant phytase from Aspergillus fumigatus into the rice endosperm. In addition, as cysteine peptides are considered a major enhancer of iron absorption, we overexpressed the endogenous cysteine-rich metallothionein-like protein. The content of cysteine residues increased about seven-fold and the phytase level in the grains about 130-fold, giving a phytase activity sufficient to completely degrade phytic acid in a simulated digestion experiment. High phytase rice, with an increased iron content and rich in cysteine-peptide, has the potential to greatly improve iron nutrition in rice-eating populations. Received: 15 April 2000 / Accepted: 12 May 2000     

Iodine and IFN-gamma synergistically enhance intercellular adhesion molecule 1 expression on NOD.H2h4 mouse thyrocytes

NOD.H2(h4) mice spontaneously develop autoimmune lymphocytic thyroiditis that mimics human Hashimoto's thyroiditis, a disease where iodine, IFN-gamma, and adhesion molecules have all been implicated in the pathogenesis. To study how iodine and IFN-gamma modulate the expression of ICAM-1, we analyzed NOD.H2(h4) thyrocytes in baseline conditions (day 0) and at several time points following supplementation of iodine in the drinking water. On day 0, a small percentage ( approximately 10%) of thyrocytes constitutively expressed ICAM-1. The expression gradually increased to 13, 25, and 41% on days 7, 14 and 28, respectively, returning to baseline (9%) on day 35. The initial ICAM-1 kinetics was paralleled by thyroidal infiltration of CD45(+) hemopoietic cells, which increased from an average of 4% on day 0 to an average of 13, 21, and 24% on days 14, 28, and 35, respectively. To distinguish whether the observed ICAM-1 increase was a direct effect of iodine or a consequence of the immune infiltrate, we treated mouse primary thyrocyte cultures with 0.01 mM sodium iodine and showed a 3-fold increased ICAM-1 expression. To assess interaction between IFN-gamma and iodine, we analyzed CD45 and ICAM-1expression on thyrocytes from NOD.H2(h4) wild-type and NOD.H2(h4) thyr-IFN-gamma transgenic littermates. Strikingly, IFN-gamma interacted synergistically with iodine to enhance ICAM-1 expression on thyrocytes. These findings suggest that iodine and IFN-gamma cooperate to promote thyroidal expression of ICAM-1 in this mouse model of thyroiditis, highlighting the complex interplay present in the pathogenesis of Hashimoto's thyroiditis.     

Sugar starvation induces the central vacuolation with coordinated increase in expression of tonoplast intrinsic protein genes in suspension-cultured rice cells

An efficient monitoring of cytoplasmic vacuolation by fluorescein diacetate (FDA) staining of protoplasts revealed that the major populations of suspension-cultured rice cells undergo a rapid vacuolation upon glucosedepletion. As aquaporin-family proteins, tonoplast intrinsic proteins (TIPs) are known to play in regulating the water balance across the vacuolar membrane. Glucose starvation increased expression of every member of OsTIP family, leading to an enhancement of the total expression by up to 110-fold, which is well matched with an expansion of the vacuolar structure induced by starvation. OsTIPs 1;1, 2;2 and 3;1 are the three most prominently expressed OsTIPs in starved conditions due to their highest responsiveness to sugar deprivation. Feeding experiments with various sugars and glucose analogs indicated that sugar regulated expression of the three major OsTIPs is likely mediated by a hexokinasedependent pathway. Alleviation of sugar-induced suppression of OsTIP expression by co-treatment with the uncoupler of ATP synthesis suggests that sugar signaling for OsTIP regulation is also cross-talked by the energy-deficit conditions. Intriguingly, starvation-induced central vacuolation was effectively prevented by mannose and 2-deoxyglucose, but not by 3-O-methylglucose. These results imply that the hexokinase is able to trigger the signaling to suppress the central vacuolation, independently of fueling the energy metabolism.     

TGF-beta and IL-13 synergistically increase eotaxin-1 production in human airway fibroblasts

Chronic diseases may involve an "innate" response followed by an adaptive immune response, of a Th1 or Th2 variety. Little is known regarding the interactions of these responses. We hypothesized that TGF-beta1 (innate response factor associated with wound repair) in combination with IL-13 (Th2 factor) might augment inflammatory processes associated with asthma. Airway fibroblasts were cultured from asthmatic subjects and normal controls. These fibroblasts were exposed to TGF-beta1 and IL-13 alone or in combination, and eotaxin-1 expression and production were evaluated. At 48 h, eotaxin-1 production was markedly increased with the combination of TGF-beta1 and IL-13 (p < 0.0001) compared with either stimulus alone. mRNA increased slightly at 1 h with IL-13 or TGF-beta1 plus IL13, peaked, and became significantly increased over IL-13 alone at 24 h. Protein was measurable from 6 h with IL-13 and TGF-beta1 plus IL-13, but greater levels were measured over time with the combination. Actinomycin ablated the increase in mRNA and protein seen with IL-13 alone and with TGF-beta1 plus IL-13. Cycloheximide blocked the increase in mRNA at 6 h in both conditions, but also blocked the increase at 24 h with TGF-beta1 plus IL-13. STAT-6 was rapidly activated with both IL-13 and the combination, without difference. Finally, eotaxin-1-positive fibroblasts were identified in severe asthma biopsies in greater numbers than in normals. These results support the concept that interactions of innate and adaptive immune systems may be important in promoting the tissue eosinophilia of asthma, particularly in those with more severe disease.     

Chemically synthesized pathogen-associated molecular patterns increase the expression of peptidoglycan recognition proteins via toll-like receptors, NOD1 and NOD2 in human oral epithelial cells

Peptidoglycan recognition proteins (PGRPs), a novel family of pattern recognition molecules (PRMs) in innate immunity conserved from insects to mammals, recognize bacterial cell wall peptidoglycan (PGN) and are suggested to act as anti-bacterial factors. In humans, four kinds of PGRPs (PGRP-L, -Ialpha, -Ibeta and -S) have been cloned and all four human PGRPs bind PGN. In this study, we examined the possible regulation of the expression of PGRPs in oral epithelial cells upon stimulation with chemically synthesized pathogen-associated molecular patterns (PAMPs) in bacterial cell surface components: Escherichia coli-type tryacyl lipopeptide (Pam3CSSNA), E. coli-type lipid A (LA-15-PP), diaminopimelic acid containing desmuramyl peptide (gamma-D-glutamyl-meso-DAP; iE-DAP), and muramyldipeptide (MDP). These synthetic PAMPs markedly upregulated the mRNA expression of the four PGRPs and cell surface expression of PGRP-Ialpha and -Ibeta, but did not induce either mRNA expression or secretion of inflammatory cytokines, in oral epithelial cells. Suppression of the expression of Toll-like receptor (TLR)2, TLR4, nucleotide-binding oligomerization domain (NOD)1 and NOD2 by RNA interference specifically inhibited the upregulation of PGRP mRNA expression induced by Pam3CSSNA, LA-15-PP, iE-DAP and MDP respectively. These PAMPs definitely activated nuclear factor (NF)-kappaB in the epithelial cells, and suppression of NF-kappaB activation clearly prevented the induction of PGRP mRNA expression induced by these PAMPs in the cells. These findings suggested that bacterial PAMPs induced the expression of PGRPs, but not proinflammatory cytokines, in oral epithelial cells, and the PGRPs might be involved in host defence against bacterial invasion without accompanying inflammatory responses.     

Expression of TERF1 in rice regulates expression of stress-responsive genes and enhances tolerance to drought and high-salinity

Drought and high-salinity are the important constraints that severely affect plant development and crop yield worldwide. It has been established that ethylene response factor (ERF) proteins play important regulatory roles in plant response to abiotic and biotic stresses. Our previous researches have revealed that transgenic tobacco over-expressing TERF1 (encoding a tomato ERF protein) showed enhanced tolerance to abiotic stress. Here, we further investigate the function of TERF1 in transgenic rice. Compared with the wild-type plants, overexpression of TERF1 resulted in an increased tolerance to drought and high-salt in transgenic rice. And the enhanced tolerance may be associated with the accumulation of proline and the decrease of water loss. Furthermore, TERF1 can effectively regulate the expression of stress-related functional genes Lip5, Wcor413-l, OsPrx and OsABA2, as well as regulatory genes OsCDPK7, OsCDPK13 and OsCDPK19 under normal growth conditions. Our analyses of cis-acting elements show that there exist DRE/CRT and/or GCC-box existing in TERF1 targeted gene promoters. Our results revealed that ectopic expression of TERF1 in rice caused a series of molecular and physiological alterations and resulted in the transgenic rice with enhanced tolerance to abiotic stress, indicating that TERF1 might have similar regulatory roles in response to abiotic stress in tobacco and rice. Shumei Gao, Haiwen Zhang and Yun Tian contributed equally to this work.     

NOD2 and CCDC122-LACC1 genes are associated with leprosy susceptibility in Brazilians

Leprosy is a complex disease with phenotypes strongly influenced by genetic variation. A Chinese genome-wide association study (GWAS) depicted novel genes and pathways associated with leprosy susceptibility, only partially replicated by independent studies in different ethnicities. Here, we describe the results of a validation and replication study of the Chinese GWAS in Brazilians, using a stepwise strategy that involved two family-based and three independent case–control samples, resulting in 3,614 individuals enrolled. First, we genotyped a family-based sample for 36 tag single-nucleotide polymorphisms (SNPs) of five genes located in four different candidate loci: CCDC122-LACC1, NOD2, TNFSF15 and RIPK2. Association between leprosy and tag SNPs at NOD2 (rs8057431) and CCDC122-LACC1 (rs4942254) was then replicated in three additional, independent samples (combined OR_AA = 0.49, P = 1.39e?06; OR_CC = 0.72, P = 0.003, respectively). These results clearly implicate the NOD2 pathway in the regulation of leprosy susceptibility across diverse populations.