Does the channel for nascent peptide exist inside the ribosome? Immune electron microscopy study

MS2 phage RNA-directed synthesis of an N-terminal polypeptide of the phage coat protein on Escherichia coli 70 S ribosomes was initiated in a cell-free system with the N-dinitrophenyl derivative of methionyl-tRNAFMet) and performed in the absence of tyrosine, lysine, cysteine and methionine. As a result, the translating ribosomes carried peptides up to 42 amino acid residues in length with the dinitrophenyl hapten at the N-ends. Using the immune electron microscopy technique the positions of the nascent peptide N-ends on the 70 S ribosomes have been visualized. It has been found that (i) the N-ends of nascent peptides of these lengths are accessible to antibodies, (ii) the exit site of a nascent peptide is the pocket between the base of the central protuberance and the L1 ridge on the 50 S subunit, i.e. presumably its peptidyl transferase center, and (iii) the further pathway of a nascent peptide seems to proceed along the groove on the external surface of the 50 S subunit.     

The natriuretic peptide system in eels: a key endocrine system for euryhalinity?

The natriuretic peptide system of a euryhaline teleost, the Japanese eel (Anguilla japonica), consists of three types of hormones [atrial natriuretic peptide (ANP), ventricular natriuretic peptide (VNP), and C-type natriuretic peptide (CNP)] and four types of receptors [natriuretic peptide receptors (NPR)-A, -B, -C, and -D]. Although ANP is recognized as a volume-regulating hormone that extrudes both Na(+) and water in mammals, ANP more specifically extrudes Na(+) in eels. Accumulating evidence shows that ANP is secreted in response to hypernatremia and acts to inhibit the uptake and to stimulate the excretion of Na(+) but not water, thereby promoting seawater (SW) adaptation. In fact, ANP is secreted immediately after transfer of eels to SW and ameliorates sudden increases in plasma Na(+) concentration through inhibition of drinking and intestinal absorption of NaCl. ANP also stimulates the secretion of cortisol, a long-acting hormone for SW adaptation, whereas ANP itself disappears quickly from the circulation. Thus ANP is a primary hormone responsible for the initial phase of SW adaptation. By contrast, CNP appears to be a hormone involved in freshwater (FW) adaptation. Recent data show that the gene expression of CNP and its specific receptor, NPR-B, is much enhanced in FW eels. In fact, CNP infusion increases (22)Na uptake from the environment in FW eels. These results show that ANP and CNP, despite high sequence identity, have opposite effects on salinity adaptation in eels. This difference apparently originates from the difference in their specific receptors, ANP for NPR-A and CNP for NPR-B. VNP may compensate the effects of ANP and CNP for adaptation to respective media, because it has high affinity to both receptors. On the basis of these data, the authors suggest that the natriuretic peptide system is a key endocrine system that allows this euryhaline fish to adapt to diverse osmotic environments, particularly in the initial phase of adaptation.     

Does the hematopoietic stem cell exist?

The hypothesis concerning the existence of a self-maintaining, i. e. "immortal" hemopoietic stem cell has no conclusive evidence. The data demonstrating the existence of hemopoietic cells with very high proliferative potentials, which are formed only during embryogenesis are discussed. Clonal expansion of these cells can support hemopoiesis during the entire life of the adult organism.     

Convergent occurrence of the developmental hourglass in plant and animal embryogenesis?

Background The remarkable similarity of animal embryos at particular stages of development led to the proposal of a developmental hourglass. In this model, early events in development are less conserved across species but lead to a highly conserved ‘phylotypic period’. Beyond this stage, the model suggests that development once again becomes less conserved, leading to the diversity of forms. Recent comparative studies of gene expression in animal groups have provided strong support for the hourglass model. How and why might such an hourglass pattern be generated? More importantly, how might early acting events in development evolve while still maintaining a later conserved stage?Scope The discovery that an hourglass pattern may also exist in the embryogenesis of plants provides comparative data that may help us explain this phenomenon. Whether the developmental hourglass occurs in plants, and what this means for our understanding of embryogenesis in plants and animals is discussed. Models by which conserved early-acting genes might change their functional role in the evolution of gene networks, how networks buffer these changes, and how that might constrain, or confer diversity, of the body plan are also discused.Conclusions Evidence of a morphological and molecular hourglass in plant and animal embryogenesis suggests convergent evolution. This convergence is likely due to developmental constraints imposed upon embryogenesis by the need to produce a viable embryo with an established body plan, controlled by the architecture of the underlying gene regulatory networks. As the body plan is largely laid down during the middle phases of embryo development in plants and animals, then it is perhaps not surprising this stage represents the narrow waist of the hourglass where the gene regulatory networks are the oldest and most robust and integrated, limiting species diversity and constraining morphological space.     

Does an animal peptide:N-glycanase have the dual role as an enzyme and a carbohydrate-binding protein?

Recently, we have reported purification and characterization of a de-N-glycosylating enzyme, peptide:N-glycanase (PNGase) found in C3H mouse fibroblast L-929 cells, and designated L-929 PNGase [Suzuki T, Seko A, Kitajima K, Inoue Y, Inoue S (1994)J Biol Chem 269, 17611–18]. The unique properties of L-929 PNGase are that the enzyme had a high affinity to the substrate glycopeptide (e.g.K _m=114 µm for fetuin derived glycopentapeptide) and that the PNGase-catalysed reaction is strongly inhibited by the released free oligosaccharides but not by the free peptides formed, suggesting that L-929 PNGase is able to bind to a certain type of carbohydrate chain. In this study, we report the new findings of the mannan-binding property of L-929 PNGase; the de-N-glycosylating enzyme activity of L-929 PNGase was inhibited by yeast mannan and triomannose, Man1 3(Man1 6)Man, but not by mannose and -methyl-d-mannoside. Furthermore, L-929 PNGase was revealed to bind to the glycan moiety of yeast mannan by using mannan-conjugated Sepharose 4B gel as a ligand, suggesting that L-929 PNGase could serve not only as an enzyme but also as a carbohydrate recognition proteinin vivo. Such dual properties found for animal-derived L-929 PNGase are unique and are not shared with other previously characterized plant- and bacterial-origin PNGases — PNGase A and PNGase F, respectively.Abbreviations GLC gas liquid chromatography - GlcNAc-Asn 2-acetamido-1--(l-aspartamido)-1,2-dideoxy-d-glucose - BSA bovine serum albumin - DTT dithiothreitol - EDTA ethylenediaminetetraacetic acid - Gal d-galactose - GlcNAc N-acetyl-d-glucosamine - Man d-mannose; triomannose, Man1 3(Man1 6)Man; - MES 2-(N-morphorino)ethanesulfonic acid - NeuAc N-acetyl-neuraminic acid - PNGase peptide:N ⁴-(N-accetyl-glucosaminyl)asparagine amidase (peptide:N-glycanase,EC 3.5.1.52) - PNP p-nitrophenyl     

Interactions between atrial natriuretic peptide and the renin–angiotensin system during salt-sensitivity exhibited by the proANP gene-disrupted mouse

To understand the involvement of the systemic and cardiac components of the renin–angiotensin system (RAS) in the development of cardiac hypertrophy induced by salt intake, the present study analyzed the effect of high dietary salt (8.0% NaCl) in mice possessing a full complement (+/+) or ablation (−/−) of atrial natriuretic peptide (ANP). A 3 week treatment of 8.0% NaCl was able to induce cardiac hypertrophy in both genotypes, though exaggerated hypertrophy was noted in the ANP −/− mouse. Although a marked decrease in angiotensin II (Ang II) plasma levels in both genotypes fed a high salt diet was observed, systemic RAS mRNA components were altered only in the ANP −/− animals and remained unchanged in ANP +/+ mice. Decreased Ang II plasma levels were better correlated with decreases in angiotensinogen protein expression observed in both genotypes. High salt had no effect on cardiac RAS mRNA components in the ANP −/− animals, but did cause a significant decrease in some cardiac RAS mRNA components in ANP +/+ mice. As expected, high salt was able to increase plasma ANP levels and ventricular mRNA expression of ANP (ANP +/+ mice only) and B-type NP in both genotypes. The latter peptides are key cardiac markers of hypertrophy whose increased expression correlate well with the physical salt-induced cardiac alterations observed in this study. These findings suggest that although the RAS does not play a key role in salt-induced cardiac hypertrophy, ANP is an important determinant of the degree of salt-sensitivity observed in the proANP gene-disrupted animal. (Mol Cell Biochem 276: 121–131, 2005)     

Does a superficial musculoaponeurotic system exist in the face and neck? An anatomical study by the tissue plastination technique

An exact knowledge of the subcutaneous layers in the different regions of the face and neck is important in several surgical disciplines. In the parotid region, a superficial musculoaponeurotic system (SMAS) has been described. The existence of a SMAS as a guiding structure for the surgeon in the other regions of the face and neck has been discussed but is controversial. Therefore, the authors investigated the development of the subcutaneous connective-tissue layers in the different facial regions and in the neck. They studied these regions in 22 human fetuses using the technique of plastination histology and in three newborn and three adult specimens using sheet plastination. In addition, they dissected the neck and face in 10 fresh adult cadavers to identify the SMAS as in the surgical situation. The results show that no SMAS could be detected in any facial regions other than the parotid region. In the parotid region, it is thick and attached to the parotid sheath. However, it becomes very thin, discontinuous, and undissectable in the cheek area. No SMAS can be found in the neck, in which the authors are the first to describe a fascia covering both sides of the platysma. This fascia has close topographical connections to the subcutaneous layers of the adjoining regions. On the basis of these findings, the surgical pathways have to be defined regionally in the face. A "platysma fascia" can be considered as a surgical landmark in the neck. Therefore, the authors conclude that it is not justified to generalize a SMAS as a surgical guiding structure.     

Does functional redundancy exist?

Functional redundancy has often been assumed as an intuitive null hypothesis in biodiversity experiments, but theory based on the classical Lotka-Volterra competition model shows that functional redundancy sensu stricto is incompatible with stable coexistence. Stable coexistence requires differences between species which lead to functional complementarity and differences between the yields of mixtures and monocultures. Only a weaker version of functional redundancy, i.e. that mixture yields lie within the range of variation of monoculture yields, is compatible with stable coexistence in Lotka-Volterra systems. Spatial and temporal environmental variability may provide room for functional redundancy at small spatial and temporal scales, but is not expected to do so at the larger scales at which environmental variations help maintain coexistence. Neutral coexistence of equivalent competitors, non-linear per capita growth rates, and lack of correlation between functional impact and biomass may provide the basis for the existence of functional redundancy in natural ecosystems. Overall, there is a striking parallel between the conditions that allow stable coexistence and those that allow overyielding.     

Does active transport exist?

Conclusion Since its publication, the two-membrane theory has been generally accepted as a prototype of a transporting epithelium. It should be remembered, however, that whereas it gives a quantitative measure of the active sodium transport, the coupling ratio between Na transport and K recycling has to be estimated by other methods (see Skou, 1957). We are reminded of our initial conclusion: that active transport can only be observed with certainty in systems where net transport of the species in question is going on.Also, the procedures described in the foregoing are powerful only for tight epithelia where paracellular shunt paths can be neglected.The finding (Ussing, 1978; Sten-Knudsen & Ussing, 1981) that the flux ratio is time independent, i.e., constant from the first appearance of the tracers on the receiving side, makes it possible, at least theoretically, to analyze multipathway systems like leaky epithelia (see, for example, Ussing & Eskesen, 1989 and Ussing & Nedergaard, 1993).     

Does plant richness influence animal richness?: the mammals of Catalonia (NE Spain)

Although it has long been held that plant diversity must influence animal diversity, the nature of this relationship remains poorly understood at large spatial scales. We compare the species richness patterns of vascular plants and mammals in north‐eastern Spain using a 100‐km² grain size to examine patterns of covariation. We found that the total mammal richness pattern, as well as those of herbivores and carnivores considered separately, only weakly corresponded to the pattern of plants. Rather, mammal richness was best described by climatic variables incorporating water inputs, and after adding these variables to multiple regression models, plant and mammal richness were virtually independent. We conclude that the observed association, although weak, is explained by shared responses of both groups to climate, and thus, plant richness has no influence on the richness pattern of Catalan mammals.     

Does a proton-pumping ATPase exist in the tonoplast?

In order to account for the accumulation of metabolites in plant vacuoles, the existence of a proton-pumping ATPase has been widely suggested in the literature. The demonstration of such a tonoplast-bound ATPase was merely based on the characterization of a nitrate-sensitive microsomal fraction. In some examples, this ATPase activity has been evidenced on vacuole preparations obtained under conditions which were criticized by Boller. The application of the reverse phase high-performance liquid chromatography method (RP-HPLC) to the simultaneous separation of adenine nucleotides, in the presence of tonoplast vesicles isolated from Catharanthus roseus, showed results not necessarily correlated with the ATPase hypothesis. Moreover, in light of the H+-quenching of quinacrine fluorescence observed during ATP hydrolysis by vacuoles or tonoplast vesicles, the existence of a proton-pumping ATPase may be questioned.     

Is there a link between salt-intake and atrial natriuretic peptide system during hypertension induced by nitric oxide blockade?

Long-term nitric oxide (NO) blockade is known to induce a severe and progressive hypertension. The influence of the salt-intake on atrial natriuretic peptide (ANP) system in this hypertension model is unknown. The aim of this study was to evaluate ANP plasma levels, content and mRNA in atria of male Wistar rats chronically treated with oral Nomega-nitro-L-arginine methyl ester (L-NAME) after 4 weeks of high-salt diet. The high-salt diet induced an increase (P < 0.05) in ANP plasma levels in normotensive rats and no significant changes in hypertensive animals. We observed a significant increase in the ANP content in the left and right atria of hypertensive rats (P < 0.001) when compared to normotensive ones. However, no significant changes were observed during high-salt diet in normotensive and hypertensive animals. Northern blot analysis revealed that ANP gene expression is higher in the right and left atria of hypertensive rats when compared to normotensive rats. However, we found no significant changes in ANP mRNA of rats treated with high-salt diet in normotensive and hypertensive rats when compared to low-salt diet. The present observations indicate no interaction between salt-intake and activation of the ANP system during chronic nitric oxide synthase (NOS) inhibition.     

Does the circadian system regulate lactation?

Environmental variables such as photoperiod, heat, stress, nutrition and other external factors have profound effects on quality and quantity of a dairy cow's milk. The way in which the environment interacts with genotype to impact milk production is unknown; however, evidence from our laboratory suggests that circadian clocks play a role. Daily and seasonal endocrine rhythms are coordinated in mammals by the master circadian clock in the hypothalamus. Peripheral clocks are distributed in every organ and coordinated by signals from the master clock. We and others have shown that there is a circadian clock in the mammary gland. Approximately 7% of the genes expressed during lactation had circadian patterns including core clock and metabolic genes. Amplitude changes occurred in the core mammary clock genes during the transition from pregnancy to lactation and were coordinated with changes in molecular clocks among multiple tissues. In vitro studies using a bovine mammary cell line showed that external stimulation synchronized mammary clocks, and expression of the core clock gene, BMAL1, was induced by lactogens. Female clock/clock mutant mice, which have disrupted circadian rhythms, have impaired mammary development and their offspring failed to thrive suggesting that the dam's milk production was not adequate enough to nourish their young. We envision that, in mammals, during the transition from pregnancy to lactation the master clock is modified by environmental and physiological cues that it receives, including photoperiod length. In turn, the master clock coordinates changes in endocrine milieu that signals peripheral tissues. In dairy cows, it is clear that changes in photoperiod during the dry period and/or during lactation influences milk production. We believe that the photoperiod effect on milk production is mediated, in part by the 'setting' of the master clock with light, which modifies peripheral circadian clocks including the mammary core clock and subsequently impacts milk yield and may impact milk composition.