Volatility of internal eliminated segments in germ line genes of hypotrichous ciliates.

Germ line micronuclear genes in ciliated protozoa contain two types of interrupting sequences. Some genes contain introns, but internal eliminated segments (IESs) are much more prevalent. IESs are AT-rich DNA segments that separate macronucleus-destined segments (MDSs) in micronuclear genes. All IESs are excised and destroyed when a micronucleus develops into a macronucleus after each cell mating. IESs have no discernible function. Therefore, an investigation of the behavior of IESs in evolution has been undertaken to assess their possible significance. The IESs in the micronuclear gene encoding the beta-subunit of the telomere-binding protein (beta-TP) are not conserved in number, position, sequence, or length during the evolution of four oxytrichid ciliates. In contrast, the scrambled pattern of MDSs and IESs of the micronuclear actin I gene has been conserved during evolution; however, the precise positions, sequences, and lengths of the IESs differ among species, and in some organisms the actin I gene contains an additional IES and MDS. Corresponding IESs in the actin I genes among the different organisms have shifted positions by 1 to 14 bp, presumably by a mutation-shifting mechanism, creating differences in the repeat sequences flanking IESs. Thus, conservation of a particular repeat sequence among species is not required for IES excision. The changes in IES number and position in the beta-TP genes among ciliates are in sharp contrast to the stability of the intron position. Therefore, IESs are volatile, hypermutable elements that are inserted, removed, shifted, and modified continuously in the germ line through evolutionary time.     

Global gene expression analysis to identify molecular markers of uterine receptivity and embryo implantation

Infertility and spontaneous pregnancy losses are an enduring problem to women's health. The establishment of pregnancy depends on successful implantation, where a complex series of interactions occurs between the heterogeneous cell types of the uterus and blastocyst. Although a number of genes are implicated in embryo-uterine interactions during implantation, genetic evidence suggests that only a small number of them are critical to this process. To obtain a global view and identify novel pathways of implantation, we used a dual screening strategy to analyze the expression of nearly 10,000 mouse genes by microarray analysis. Comparison of implantation and interimplantation sites by a conservative statistical approach revealed 36 up-regulated genes and 27 down-regulated genes at the implantation site. We also compared the uterine gene expression profile of progesterone-treated, delayed implanting mice to that of mice in which delayed implantation was terminated by estrogen. The results show up-regulation of 128 genes and down-regulation of 101 genes after termination of the delayed implantation. A combined analysis of these experiments showed specific up-regulation of 27 genes both at the implantation site and during uterine activation, representing a broad diversity of molecular functions. In contrast, the majority of genes that were decreased in the combined analysis were related to host immunity or the immune response, suggesting the importance of these genes in regulating the uterine environment for the implanting blastocyst. Collectively, we identified genes with recognized roles in implantation, genes with potential roles in this process, and genes whose functions have yet to be defined in this event. The identification of unique genetic markers for the onset of implantation signifies that genome-wide analysis coupled with functional assays is a promising approach to resolve the molecular pathways required for successful implantation.     

The unique Phe–His dyad of 2-ketopropyl coenzyme M oxidoreductase/carboxylase selectively promotes carboxylation and S–C bond cleavage

The 2-ketopropyl-coenzyme M oxidoreductase/carboxylase (2-KPCC) enzyme is the only member of the disulfide oxidoreductase (DSOR) family of enzymes, which are important for reductively cleaving S–S bonds, to have carboxylation activity. 2-KPCC catalyzes the conversion of 2-ketopropyl-coenzyme M to acetoacetate, which is used as a carbon source, in a controlled reaction to exclude protons. A conserved His–Glu motif present in DSORs is key in the protonation step; however, in 2-KPCC, the dyad is substituted by Phe–His. Here, we propose that this difference is important for coupling carboxylation with C–S bond cleavage. We substituted the Phe–His dyad in 2-KPCC to be more DSOR like, replacing the phenylalanine with histidine (F501H) and the histidine with glutamate (H506E), and solved crystal structures of F501H and the double variant F501H_H506E. We found that F501 protects the enolacetone intermediate from protons and that the F501H variant strongly promotes protonation. We also provided evidence for the involvement of the H506 residue in stabilizing the developing charge during the formation of acetoacetate, which acts as a product inhibitor in the WT but not the H506E variant enzymes. Finally, we determined that the F501H substitution promotes a DSOR-like charge transfer interaction with flavin adenine dinucleotide, eliminating the need for cysteine as an internal base. Taken together, these results indicate that the 2-KPCC dyad is responsible for selectively promoting carboxylation and inhibiting protonation in the formation of acetoacetate.     

Régénération des îlots de langerhans,au cours de la correction du diabète expérimental du rat par bréphoplastie pancréatique

Résumé La greffe de pancréas foetal chez le rat alloxanisé (une injection d'alloxane de 100–250 mg/kg) corrige immédiatement et définitivement le diabète. La sécrétion d'insuline est, au début, assurée par le greffon puisque dans les îlots du pancréas de l'hôte l'alloxane a provoqué la destruction totale des cellules B. Durant les deux premières semaines qui suivent la bréphoblastie, les îlots sont le lieu d'une prolifération massive de cellules A; les premières cellules B néoformées apparaissent vers le 15^e jour mais la proportion normale des cellules AB n'est rétablie que 1 1/2 à 2 mois après l'implantation de la greffe. Au fur et à mesure de la régénération des cellules B, la fonction insulinique des îlots du pancréas de l'hôte se substitue à celle du greffon qui dégénère progressivement.Chez les rats pancréatectomisés et greffés, la sécrétion d'insuline est également assurée par le greffon pendant le 1^er mois environ. La régénération du pancréas à partir de reliquats pancréatiques laissés dans la région de la confluence des canaux de Wirsung et biliaire, aboutit, à 3 1/2 mois, à une polynésie d'îlots, souvent volumineux et irréguliers, formés presqu'exclusivement de cellules B. La glycémie restant constamment normale, la sécrétion d'insuline est ici encore dans une première phase, assumée par le greffon qui dégénère, par la suite, au fur et à mesure que les îlots du régénérat sont capables de secréter de l'insuline en quantité suffisante pour assurer l'équilibre glycémique.La prolifération et la néogenèse des cellules A, comme celles des cellules B, se font essentiellement aux dépens des cellules des acini exocrines qui perdent leurs caractères de cellules exocrines (disparition des granulations de zymogène et de la réserve de RNA) et prolifèrent en gros bourgeons plasmodiaux A ou B. La différenciation de cellules endocrines se fait également, mais plus rarement, à partir de l'épithélium des petits canalicules secrétoires sous acineux.Bréphoplastie = greffe d'organe foetal; terme créé par R. M. May.     

STUDIES ON THE MECHANISM OF HYDROGEN TRANSPORT IN ANIMAL TISSUES : VI. INHIBITOR STUDIES WITH SUCCINIC DEHYDROGENASE

1. The mechanism of succinic dehydrogenase action was studied by means of inhibitors. 2. The enzyme is inhibited by a large number of diverse compounds whose only common denominator appears to be their ability to react with SH groups. These compounds include quinonoid structures, sulfhydryl reagents, sulfhydryl compounds, copper, zinc, selenite, and arsenite. 3. In contrast to the above inhibitors, the action of malonate does not appear to involve sulfhydryl groups and is explained on the basis of its affinity for the enzyme groups which react with the carboxyl groups of succinate. 4. The action of malonate and the sulfhydryl reactants is mutually exclusive, and this fact suggests the conclusion that the sulfhydryl group of the enzyme is located between the carboxyl affinity points. 5. On the basis of the deduced structure of the succinate-activating center of the enzyme, it is suggested that the enzyme may function by oscillating between the EnSH and EnS· forms, rather than by a thiol-disulfide equilibrium.     

Coral bleaching response index: a new tool to standardize and compare susceptibility to thermal bleaching

As coral bleaching events become more frequent and intense, our ability to predict and mitigate future events depends upon our capacity to interpret patterns within previous episodes. Responses to thermal stress vary among coral species; however the diversity of coral assemblages, environmental conditions, assessment protocols, and severity criteria applied in the global effort to document bleaching patterns creates challenges for the development of a systemic metric of taxon‐specific response. Here, we describe and validate a novel framework to standardize bleaching response records and estimate their measurement uncertainties. Taxon‐specific bleaching and mortality records (2036) of 374 coral taxa (during 1982–2006) at 316 sites were standardized to average percent tissue area affected and a taxon‐specific bleaching response index (taxon‐BRI) was calculated by averaging taxon‐specific response over all sites where a taxon was present. Differential bleaching among corals was widely variable (mean taxon‐BRI = 25.06 ± 18.44%, ±SE). Coral response may differ because holobionts are biologically different (intrinsic factors), they were exposed to different environmental conditions (extrinsic factors), or inconsistencies in reporting (measurement uncertainty). We found that both extrinsic and intrinsic factors have comparable influence within a given site and event (60% and 40% of bleaching response variance of all records explained, respectively). However, when responses of individual taxa are averaged across sites to obtain taxon‐BRI, differential response was primarily driven by intrinsic differences among taxa (65% of taxon‐BRI variance explained), not conditions across sites (6% explained), nor measurement uncertainty (29% explained). Thus, taxon‐BRI is a robust metric of intrinsic susceptibility of coral taxa. Taxon‐BRI provides a broadly applicable framework for standardization and error estimation for disparate historical records and collection of novel data, allowing for unprecedented accuracy in parameterization of mechanistic and predictive models and conservation plans.     

Nitric oxide production and absorption in trachea, bronchi, bronchioles, and respiratory bronchioles of humans

Different volumesof dead-space gas were collected and analyzed for nitric oxide (NO)content, either immediately after inspiration or after a period ofbreath holding on clean air or NO mixtures. This allowed calculation ofNO equilibrium, NO production, and NO absorption. In seven young,healthy, adult nonsmokers, the mean NO equilibrium values in parts perbillion (ppb) were 56 ± 11 (SE) in the trachea, 37 ± 6 in thebronchi, 21 ± 3 in the bronchioles, and 16 ± 2 in therespiratory bronchioles. At any given NO concentration, the NOabsorption rate (in nl/min) equaled the NO concentration (in ppb) timesA (the absorption coefficient inl/min). A values (in l/min) were 0.11 ± 0.01 in the trachea, 0.17 ± 0.04 in the bronchi, 0.66 ± 0.09 in the bronchioles, and 1.35 ± 0.32 in the respiratorybronchioles. NO equilibrium concentrations and production rates in one74-yr-old subject were three to five times as high as those found inthe young subjects. Mouth equilibrium NO concentrations were 3 and 6 parts per million in two subjects who had oral production rates of 6 and 23 nl/min, respectively. In conclusion, production and absorptionof NO occur throughout the first 450 ml of the airways.