Selective ion tracing and MSn analysis of peptide digests from FSBA-treated kinases for the analysis of protein ATP-binding sites

Kinases play a key role in many cellular processes by catalyzing the transfer of phosphoryl groups from ATP to a broad number of substrates, including amino acids on target proteins. The reagent 5'-fluorosulfonylbenzoyl-5'-adenosine (FSBA) has been widely used to identify ATP-binding sites in kinases since it reacts with nucleophilic amino acids occurring within these motifs, determining a mass increase of 433 Da. In this study, we present a versatile MS approach that has been developed to recognize labeled peptides generated after enzymatic digestion of FSBA-treated kinases. Using selective ion tracing and MS(2)/MS(3) experiments, we were able to easily identify peptides occurring at protein ATP-binding sites, also affording a complete characterization of the modified amino acids. This methodology may be used in the development of future parent ion scanning-based applications directed to large scale analysis of kinases within complex protein mixtures.     

Immunohistochemical observations on tubulin and alpha actinin during mitosis of cultured fibroblasts

In this work we studied alpha-actinin and tubulin sites in rabbit fibroblasts in culture. Antibodies anti-alpha-actinin were used for indirect PAP-reaction while antibodies anti-tubulin were used for indirect immunofluorescence method. The observations were carried out by light microscopy, phase-contrast and interference-contrast microscopy, with regard to actinin, and by fluorescence microscopy, with regard to tubulin. During the early mitotic phase, alpha-actinin is localized all over the cell membrane of the fibroblasts, forming a sort strong protective cap, while during diacinesis it forms only rings, localizing below the cell membrane and the philopodia. Thus tubulin forms the bundle fibres during mitotic phases.     

Renpenning syndrome maps to Xp11.

Mutations in genes on the X chromosome are believed to be responsible for the excess of males among individuals with mental retardation. Such genes are numerous, certainly >100, and cause both syndromal and nonsyndromal types of mental retardation. Clinical and molecular studies have been conducted on the Mennonite family with X-linked mental retardation (XLMR) reported, in 1962, by Renpenning et al. The clinical phenotype includes severe mental retardation, microcephaly, up-slanting palpebral fissures, small testes, and stature shorter than that of nonaffected males. Major malformations, neuromuscular abnormalities, and behavioral disturbances were not seen. Longevity is not impaired. Carrier females do not show heterozygote manifestations. The syndrome maps to Xp11.2-p11.4, with a maximum LOD score of 3.21 (recombination fraction 0) for markers between DXS1039 and DXS1068. Renpenning syndrome (also known as "MRXS8"; gene RENS1, MIM 309500) shares phenotypic manifestations with several other XLMR syndromes, notably the Sutherland-Haan syndrome. In none of these entities has the responsible gene been isolated; hence, the possibility that two or more of them may be allelic cannot be excluded at present.     

Different Relationship Between Electron Transport and CO2 Assimilation in two Zea mays Cultivars as Influenced by Increasing Irradiance

Gas exchange and fluorescence parameters were measured simultaneously in two Zea mays L. cultivars (Liri and 121C D8) to assess the relationship between the quantum yield of electron transport (_PS2) and the quantum yield of CO₂ assimilation (_CO2) in response to photosynthetic photon flux density (PPFD). The cv. Liri was grown under controlled environmental conditions in a climate chamber (CC) while cv. 121C D8 was grown in CC as well as outdoors (OT). By exposing the two maize cultivars grown in CC to an increasing PPFD, higher photosynthetic and photochemical rates were evidenced in cv. Liri than in cv. 121C D8. In Liri plants the _PS2/_CO2 ratio increased progressively up to 27 with increasing PPFD. This suggests that the reductive power was more utilised in non-assimilatory processes than in CO₂ assimilation at high PPFD. On the contrary, by exposing 121C D8 plants to increasing PPFD, _PS2/_CO2 was fairly constant (around 11–13), indicating that the electron transport rate was tightly down regulated by CO₂ assimilation. Although no significant differences were found between _PS2/_CO2 of the 121C D8 maize grown under CC and OT by exposing them to high PPFD, the photosynthetic rate and photochemical rates were higher in OT maize plants.     

Full light and soil drought constrain plant growth in Mediterranean cliffs: the case of Primula palinuri Petagna

In any biogeographical region, cliffs shelter a larger number of relict species than do the surrounding areas. To develop effective conservation strategies, experimental studies are needed. We focused on Mediterranean coastal cliffs and on Primula palinuri Petagna, an endangered narrow-endemic plant, included in the IUCN Red List and chosen as a key species. The hypothesis was that light intensity and soil water availability could affect survival and growth of juvenile plants. To avoid the bias of other cliff environmental factors and to overcome problems of cliff site inaccessibility, we performed an ex-situ experiment. The effect of two light regimes and two soil water availability conditions on survival and morpho-functional traits were analysed. Plants grown under reduced light intensity and wet soil showed no reduction in survival and performed very well in terms of growth and storage accumulation. By contrast, full light and dry soil proved to be detrimental conditions. Overall results suggested that cliff environmental conditions play an important role in conservation of narrow-endemic species. The decreasing trend of the already severely fragmented populations of P. palinuri and the ongoing increasing aridity is likely to contribute to a further reduction of the already narrow distribution this cliff relict species.     

A Comparison of Tension Headache Sufferers and Nonpain Controls on the State-Trait Anger Expression Inventory: An Exploratory Study with Implications for Applied Psychophysiologists

Studies that have examined the relationship between personality characteristics and tension headache have arrived at conflicting and, for the most part, negative results. In recent years, a number of investigators have begun examining the relationship between anger and psychophysiological disorders, focusing mostly on anger which is suppressed or held in rather than expressed behaviorally. The present study explored the relationship between anger in 59 tension headache subjects and compared their results to 33 nonpain controls. Materials consisted of the revised research edition of the Spielberger State-Trait Anger Expression Inventory. As predicted, tension headache sufferers were found to have significantly more anger held inward than nonpain controls. Implications for applied psychophysiology treatment and future research directions are discussed.     

Alternative splicing of the estrogen receptor primary transcript normally occurs in estrogen receptor positive tissues and cell lines

Several laboratories have described estrogen receptor mRNA variants created by skipping internal exons. Some of the putative proteins encoded for by these variants have been functionally characterized by transfection analyses. The variant lacking exon 5 would lead, if translated, to a truncated receptor which shows dominant positive transactivation activity in the absence of hormone. It has been postulated that the variant could account for anti-estrogen resistant tumor growth and for expression of the progesterone receptor in estrogen receptor negative tumors. In order to understand the possible role this and other variants may have in the tumorigenesis of mammary tissue we have carried out a thorough analysis of variants expressed in a tumor cell line (MCF-7), in a tumor sample and in a sample of normal breast tissue derived from mammary reduction surgery. We performed rt-PCR analyses followed by hybridization with exon specific oligonucleotide probes. By these means we have detected nine different variants co-expressed in MCF-7 cells and at least the major variants were equally expressed in normal and neoplastic breast tissue. The same is true for the variant lacking exon 5 which, however, resulted to be a variant of low expression in the three samples analyzed. Variant formation appeared to be restricted to the estrogen receptor messenger since several other members of the superfamily of nuclear receptors did not show variant formation. We also have analyzed the effect of the most abundantly expressed variant, the exon 4 lacking variant, on normal estrogen receptor function, on the growth and on the response to estradiol and to tamoxifen of MCF-7 cells. Although over-expressed at high levels this variant has, if any, only marginal effects on the expression of endogenous estrogen regulated genes and on growth and response to the hormone and its antagonist. Although the lack of function of this variant cannot be extrapolated to other variants, their involvement in tumor formation appears rather unlikely since they are also expressed in normal tissue and the single variant is expressed in addition to many others, some of which might have opposing effects. Variant formation is, however, specific for the estrogen receptor and apparently regulated with tissue specificity as our expression analysis in normal mouse tissues shows. Therefore the variants probably have a physiological significance yet to be discovered.     

Immunobiosis and probiosis: antimicrobial activity of lactic acid bacteria with a focus on their antiviral and antifungal properties

Lactic acid bacteria (LAB), a heterogeneous group of bacteria that produce lactic acid as the main product of carbohydrate degradation, play an important role in the production and protection of fermented foods. Moreover, beside the technological use of these microorganisms added to control and steer food fermentations, their beneficial healthy properties are largely overt. Thus, numerous LAB strains have obtained the probiotic status, which entails the ability to maintain and promote a good health of consumers. In particular, increasing consideration is being focused on probiotic microorganisms that can improve the human immune response against dangerous viral and fungal enemies. For such beneficial microbes, the term “immunobiotics” has been coined. Together with an indirect host-mediated adverse effect against undesirable microorganisms, also a direct antagonistic activity of several LAB strains has been largely demonstrated. The purpose of this review is to provide a fullest possible overview of the antiviral and antifungal activities ascribed to probiotic LAB. The interest in this research field is substantiated by a large number of studies exploring the potential application of these beneficial microorganisms both as biopreservatives and immune-enhancers, aiming to reduce and/or eliminate the use of chemical agents to prevent the development of pathogenic, infectious, and/or degrading causes.