Epidemiology of pelvic floor disorders between urban and rural female inhabitants

The aim of this study was to investigate the prevalence of pelvic organ prolapse in urban and rural women and to identify possible related factors. They were 1749 participants; one thousand four hundred seventeen (81%) urban women and 332 rural residents (19%). The urban and rural women were congruently regarding to age, parity, using oral contraceptives and postmenopausal status. The urban women were more often obese (p < 0.01), estrogen replacement users (p < 0.001), smokers (p < 0.001), with mild (p < 0.001) and high (p < 0.001) education, and they were often divorced (p < 0.05) than rural women. Rural women were more often alcohol consumers (p < 0.001), with low level of education (p < 0.001) and more often married (p < 0.05) than rural examinees. There were no association between the presence of prolapse and: weight, menopausal status, oral contraceptives and estrogen replacement using, smoking, alcohol consuming and marital status. There were not observed differences in prevalence of prior hysterectomy, urinary incontinence, uroinfectio, sexual and bowel dysfunction between both groups. The prevalence of cystocele, rectocele and uterine prolapse were similar among urban and rural participants. In conclusion, a more complete picture of factors associated with genital prolapse would include in investigation, such as molecular and genetic ones.     

Genome-wide RNAi screening identifies human proteins with a regulatory function in the early secretory pathway

The secretory pathway in mammalian cells has evolved to facilitate the transfer of cargo molecules to internal and cell surface membranes. Use of automated microscopy-based genome-wide RNA interference screens in cultured human cells allowed us to identify 554 proteins influencing secretion. Cloning, fluorescent-tagging and subcellular localization analysis of 179 of these proteins revealed that more than two-thirds localize to either the cytoplasm or membranes of the secretory and endocytic pathways. The depletion of 143 of them resulted in perturbations in the organization of the COPII and/or COPI vesicular coat complexes of the early secretory pathway, or the morphology of the Golgi complex. Network analyses revealed a so far unappreciated link between early secretory pathway function, small GTP-binding protein regulation, actin cytoskeleton organization and EGF-receptor-mediated signalling. This work provides an important resource for an integrative understanding of global cellular organization and regulation of the secretory pathway in mammalian cells.     

S-methylmethionine plays a major role in phloem sulfur transport and is synthesized by a novel type of methyltransferase.

All flowering plants produce S-methylmethionine (SMM) from Met and have a separate mechanism to convert SMM back to Met. The functions of SMM and the reasons for its interconversion with Met are not known. In this study, by using the aphid stylet collection method together with mass spectral and radiolabeling analyses, we established that l-SMM is a major constituent of the phloem sap moving to wheat ears. The SMM level in the phloem ( approximately 2% of free amino acids) was 1.5-fold that of glutathione, indicating that SMM could contribute approximately half the sulfur needed for grain protein synthesis. Similarly, l-SMM was a prominently labeled product in phloem exudates obtained by EDTA treatment of detached leaves from plants of the Poaceae, Fabaceae, Asteraceae, Brassicaceae, and Cucurbitaceae that were given l-(35)S-Met. cDNA clones for the enzyme that catalyzes SMM synthesis (S-adenosylMet:Met S-methyltransferase; EC 2.1.1.12) were isolated from Wollastonia biflora, maize, and Arabidopsis. The deduced amino acid sequences revealed the expected methyltransferase domain ( approximately 300 residues at the N terminus), plus an 800-residue C-terminal region sharing significant similarity with aminotransferases and other pyridoxal 5'-phosphate-dependent enzymes. These results indicate that SMM has a previously unrecognized but often major role in sulfur transport in flowering plants and that evolution of SMM synthesis in this group involved a gene fusion event. The resulting bipartite enzyme is unlike any other known methyltransferase.     

Identification and characterization of the missing phosphatase on the riboflavin biosynthesis pathway in Arabidopsis thaliana

Despite the importance of riboflavin as the direct precursor of the cofactors flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN), the physiologically relevant catalyst dephosphorylating the riboflavin biosynthesis pathway intermediate 5‐amino‐6‐ribitylamino‐2,4(1H,3H) pyrimidinedione 5′‐phosphate (ARPP) has not been characterized from any organism. By using as the query sequence a previously identified plastidial FMN hydrolase AtcpFHy1 (At1g79790), belonging to the haloacid dehalogenase (HAD) superfamily, seven candidates for the missing ARPP phosphatase were found, cloned, recombinantly expressed, and purified. Activity screening showed that the enzymes encoded by AtcpFHy1, At4g11570, and At4g25840 catalyze dephosphorylation of ARPP. AtcpFHy1 was renamed AtcpFHy/PyrP1, At4g11570 and At4g25840 were named AtPyrP2 and AtGpp1/PyrP3, respectively. Subcellular localization in planta indicated that AtPyrP2 was localized in plastids and AtGpp1/PyrP3 in mitochondria. Biochemical characterization of AtcpFHy/PyrP1 and AtPyrP2 showed that they have similar K_m values for the substrate ARPP, with AtcpFHy/PyrP1 having higher catalytic efficiency. Screening of 21 phosphorylated substrates showed that AtPyrP2 is specific for ARPP. Molecular weights of AtcpFHy/PyrP1 and AtPyrP2 were estimated at 46 and 72 kDa, suggesting dimers. pH and temperature optima for AtcpFHy/PyrP1 and AtPyrP2 were ~7.0–8.5 and 40–50°C. T‐DNA knockout of AtcpFHy/PyrP1 did not affect the flavin profile of the transgenic plants, whereas silencing of AtPyrP2 decreased accumulation of riboflavin, FMN, and FAD. Our results strongly support AtPyrP2 as the missing phosphatase on the riboflavin biosynthesis pathway in Arabidopsis thaliana. The identification of this enzyme closes a long‐standing gap in understanding of the riboflavin biosynthesis in plants.     

A dual regulatory role of Arabidopsis calreticulin-2 in plant innate immunity

Calreticulin (CRT) is an endoplasmic reticulum-resident calcium-binding molecular chaperone that is highly conserved in multi-cellular eukaryotes. Higher plants contain two distinct groups of CRTs: CRT1/CRT2 and CRT3 isoforms. Previous studies have shown that bacterial elongation factor Tu receptor (EFR), a pattern-recognition receptor that is responsible for pathogen-associated molecular pattern-triggered immunity, is a substrate for Arabidopsis CRT3, suggesting a role for CRT3 in regulating plant defense against pathogens. Here we report that Arabidopsis CRT2 is another regulator of plant innate immunity. Despite significantly increased salicylic acid levels and constitutive expression of the systemic acquired resistance-associated marker genes PR1, PR2 and PR5, transgenic plants over-expressing CRT2 displayed reduced resistance to virulent Pseudomonas syringae pv. tomato DC3000 (PstDC3000). A (45)Ca(2+) overlay assay and a domain-swapping experiment further demonstrated that the negatively charged C-terminal tail of CRT2 is responsible for its high calcium-binding capacity and function in regulating the endogenous salicylic acid level. In addition, over-expression of the His173 mutant of CRT2 greatly enhanced plant defense against PstDC3000, supporting the existence of a self-inhibition mechanism that can counteract the effects of salicylic acid-dependent immune responses. These results suggest that CRT2 functions through its N-terminal domain(s) as a self-modulator that can possibly prevent the salicylic acid-mediated runaway defense responses triggered by its C-terminal calcium-buffering activity in response to pathogen invasion.     

CD‐sensitive Zn‐porphyrin tweezer host‐guest complexes,part 2: Cis‐ and trans‐3‐hydroxy‐4‐aryl/alkyl‐β‐lactams. A case study

This article describes an application of the host‐guest chiral recognition approach called tweezer methodology for the determination of the absolute configuration of 3‐hydroxy‐β‐lactams. These substrates represent challenging cases due to their chemical reactivity, the presence of multiple stereogenic centers and several functional groups which offer various possibilities of binding to the Zn‐porphyrin host. OPLS‐2005, the force field used in this work to predict the interporphyrin twist, modeled correctly the host‐guest complexation mechanism and revealed conformational details of the bound substrates. The computational study also suggested that in cases where an increase in the magnitude of the stereodifferentiation and an intense experimental CD are observed, the bound conformation of the conjugates are hydrogen bonded. The present investigation provides evidence that when the tweezer method is assisted by the OPLS‐2005 based computational approach, it can be successfully applied to the configurational and conformational elucidation of multi‐functional compounds with multiple stereogenic centers. Chirality, 2010. © 2009 Wiley‐Liss, Inc.     

The folate precursor p-aminobenzoate is reversibly converted to its glucose ester in the plant cytosol

Plants synthesize p-aminobenzoate (pABA) in chloroplasts and use it for folate synthesis in mitochondria. It has generally been supposed that pABA exists as the free acid in plant cells and that it moves between organelles in this form. Here we show that fruits and leaves of tomato and leaves of a diverse range of other plants have a high capacity to convert exogenously supplied pABA to its beta-D-glucopyranosyl ester (pABA-Glc), whereas yeast and Escherichia coli do not. High performance liquid chromatography analysis indicated that much of the endogenous pABA in fruit and leaf tissues is esterified and that the total pool of pABA (free plus esterified) varies greatly between tissues (from 0.2 to 11 nmol g-1 of fresh weight). UDP-glucose:pABA glucosyltransferase activity was readily detected in fruit and leaf extracts, and the reaction was found to be freely reversible. p-Aminobenzoic acid beta-D-glucopyranosyl ester esterase activity was also detected in extracts. Subcellular fractionation indicated that the glucosyltransferase and esterase activities are predominantly if not solely cytosolic. Taken together, these results show that reversible formation of pABA-Glc in the cytosol is interposed between pABA production in chloroplasts and pABA consumption in mitochondria. As pABA is a hydrophobic weak acid, its uncharged form is membrane-permeant, and its anion is consequently prone to distribute itself spontaneously among subcellular compartments according to their pH. Esterification of pABA may eliminate such errant behavior and provide a readily reclaimable storage form of pABA as well as a substrate for membrane transporters.