RETRACTED ARTICLE: Antituberculars which target decaprenylphosphoryl-β-D-ribofuranose 2′-oxidase DprE1: state of art

Multidrug resistance is a major barrier in the battle against tuberculosis and still a leading cause of death worldwide. In order to fight this pathogen, two routes are practicable: vaccination or drug treatment. Vaccination against Mycobacterium tuberculosis with the current vaccine Mycobacterium bovis Bacillus Calmette–Guerin is partially successful, being its efficacy variable. A few new tuberculosis vaccines are now in various phases of clinical trials. The emergence of multidrug-resistant strains of M. tuberculosis gave the impulse to discover new effective antitubercular drugs, a few of which are in clinical development. Here we focus on three different classes of very promising antitubercular drugs recently discovered (benzothiazinones, dinitrobenzamides, and benzoquinoxalines) that share the same cellular target: a subunit of the heteromeric decaprenylphosphoryl-β-d-ribose 2′-epimerase, encoded by the dprE1 (or Rv3790) gene. This enzyme is involved in the biosynthesis of d-arabinose which is crucial for the synthesis of the mycobacterial cell wall and essential for the pathogen’s survival.     

PI3K and ERK 1-2 regulate early stages during head regeneration in hydra

Different signaling systems coordinate and regulate the development of a multicellular organism. In hydra, the canonical Wnt pathway and the signal transduction pathways mediated by PKC and Src regulate early stages of head formation. In this paper, we present evidence for the participation of a third pathway, the PI3K-PKB pathway, involved in this process. The data presented here are consistent with the participation of ERK 1-2 as a point of convergence for the transduction pathways mediated by PKC, Src and PI3K for the regulation of the regeneration of the head in hydra. The specific developmental point regulated by them appears to be the commitment of tissue at the apical end of the regenerate to form the head organizer.     

p85 Associates with Unphosphorylated PTEN and the PTEN-Associated Complex

The lipid phosphatase PTEN functions as a tumor suppressor by dephosphorylating the D3 position of phosphoinositide-3,4,5-trisphosphate, thereby negatively regulating the phosphoinositide 3-kinase (PI3K)/AKT signaling pathway. In mammalian cells, PTEN exists either as a monomer or as a part of a >600-kDa complex (the PTEN-associated complex [PAC]). Previous studies suggest that the antagonism of PI3K/AKT signaling by PTEN may be mediated by a nonphosphorylated form of the protein resident within the multiprotein complex. Here we show that PTEN associates with p85, the regulatory subunit of PI3K. Using newly generated antibodies, we demonstrate that this PTEN-p85 association involves the unphosphorylated form of PTEN engaged within the PAC and also includes the p110β isoform of PI3K. The PTEN-p85 association is enhanced by trastuzumab treatment and linked to a decline in AKT phosphorylation in some ERBB2-amplified breast cancer cell lines. Together, these results suggest that integration of p85 into the PAC may provide a novel means of downregulating the PI3K/AKT pathway.The phosphoinositide 3-kinase (PI3K)/AKT signaling pathway regulates glucose/nutrient homeostasis and cell survival and plays a central role in both normal metabolism and cancer. The PTEN tumor suppressor gene (29, 30, 54) negatively regulates the PI3K/AKT pathway by dephosphorylating the D3 hydroxyl subunit of phosphoinositide-3,4,5-trisphosphate, a key membrane phosphatidylinositol generated by PI3K (34). PTEN undergoes genetic or epigenetic inactivation in many malignancies, including glioblastoma, melanoma, and endometrial, prostate, and breast cancers, among others (6, 13, 22, 23, 47, 49-51, 55, 68). Similarly, germ line mutations of PTEN are associated with the development of hamartomatous neoplasias such as Cowden disease and Bannayan-Zonana syndrome (17, 21, 41).The tumor suppressor function of PTEN undergoes dynamic regulation involving both C-terminal phosphorylation and protein-protein interactions. Phosphorylation of serine and threonine residues at the PTEN C-terminal tail, mediated by kinases such as CK2 and glycogen synthase kinase 3β, alters its conformational structure and association with PDZ domain-containing proteins and attenuates PTEN enzymatic activity (1, 11, 20, 32, 45, 61-63, 66, 67, 71). Conversely, PTEN function is promoted in large part through its stabilization in unphosphorylated form by incorporation into a high-molecular-weight protein complex (the PTEN-associated complex [PAC]) (66). We first demonstrated the existence of the PAC through gel filtration studies of rat liver extracts, which identified PTEN within a high-molecular-mass peak (>600 kDa), as well as a low-molecular-mass peak (40 to 100 kDa) in which PTEN is monomeric and phosphorylated (66). Subsequently, several PDZ domain-containing proteins were shown to interact with PTEN, including MAGI-1b, MAGI-2, MAGI-3, ghDLG, hMAST205, MSP58/MCRS1, NHERF1, and NHERF2, which mediate indirect binding with platelet-derived growth factor (PDGF) receptor β (25, 36, 42, 57, 66). More recently, LKB1, a serine/threonine kinase tumor suppressor (7), was also found to interact with and phosphorylate PTEN in vitro (36). In aggregate, these data suggest that PTEN functional output is controlled by a complex interplay of protein interactions and regulation of C-terminal phosphorylation.Beyond these interactions, there is also evidence to support additional regulatory mechanisms by which the tumor suppressor function of PTEN is mediated. The herpesvirus-associated ubiquitin-specific protease was shown to interact directly with PTEN and promote its nuclear entry (53). Both ubiquitination and relocalization into the nucleus constitute important PTEN regulatory mechanisms (53, 64). In many tumors, PTEN nuclear exclusion has been associated with poor cancer prognosis and more aggressive cancer development (15, 44, 56). Moreover, successful treatment of acute promyelocytic leukemia was shown to be associated with an increase in monoubiquitinylation and relocation of PTEN into the nucleus (53).Like PTEN, the p85 regulatory subunit of PI3K serves as a prominent modulator of PI3K/AKT signaling. p85, which exists in three isoforms (α, β, and γ), targets the catalytic (110-kDa) PI3K subunit to the membrane, which brings it into proximity with membrane-associated phosphatidylinositol lipids. In the steady state, p85 forms a tight association with the catalytic PI3K subunit, usually p110α or p110β in nonhematopoietic cells, with p110δ predominating in leukocytes (19). Consistent with this notion, p85 and p110 exist in equimolar ratios in a wide variety of mammalian cell lines and tissues (19), although some studies have suggested a role for free p85 in cell signaling (33, 65).Several recent lines of evidence have begun to support a possible regulatory relationship between PTEN and p85 (reviewed in references 3 and 53). For example, liver-specific deletion of PIK3R1, which encodes the p85α regulatory subunit, reduces both the activation of PI3K and PTEN enzymatic activity in this context. As a result, p85α-deficient hepatic cells express elevated levels of phosphoinositide trisphosphate and exhibit prolonged AKT activation (60). In addition, both PTEN and p85 are regulated by small GTPase proteins such as RhoA, but PTEN coimmunoprecipitates with the RhoA effector Rock only in the presence of PI3K (18, 31, 37). Although only correlative in nature, these findings may suggest a possible role for PTEN in p85 regulation or vice versa, in addition to its known function as a direct antagonist of the PI3K/AKT pathway (3, 9, 52, 57, 60).In the present study, we demonstrate an endogenous association between p85 and PTEN. Using newly generated antibodies that selectively recognize the PTEN C-terminal tail in its unphosphorylated form, we demonstrate that this PTEN-p85 association preferentially involves the unphosphorylated form of PTEN. The specificity of this interaction was confirmed using multiple antibodies and through studies of both human cancer cells and murine embryonic fibroblasts (MEFs) deficient for specific p85 subunits. This association, which also engages p110β, is enhanced by trastuzumab treatment and correlates with diminished AKT phosphorylation. These results support a functional role for the PTEN-p85 association that may have important biological and therapeutic implications for PI3K/AKT pathway regulation.     

Dream work with children: Perceptions and practices of school mental health professionals.

In this study, 49 public school mental health practitioners (school counselors, school psychologists, and school social workers) completed a survey about working with students’ dreams. The majority of these practitioners reported having at least one student bring up dreams during counseling, more frequently with troubling dreams and nightmares or when coping with grief. Results showed that practitioners were less likely to talk about dreams with students who had been identified with an adjustment disorder, psychosis, or eating disorder; those who were oppositional or ill; and those who struggled with substance abuse problems. Although most practitioners did not feel competent working with children’s dreams and reported minimal training in dream work, they were interested in learning more about children’s dreams and potential uses of dream work in supportive counseling. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Recent Emergence of Clonal Group O25b:K1:H4-B2-ST131 ibeA Strains among Escherichia coli Poultry Isolates,Including CTX-M-9-Producing Strains,and Comparison with Clinical Human Isolates

To discern the possible spread of the Escherichia coli O25b:H4-ST131 clonal group in poultry and the zoonotic potential of avian strains, we made a retrospective search of our strain collection and compared the findings for those strains with the findings for current strains. Thus, we have characterized a collection of 19 avian O25b:H4-ST131 E. coli strains isolated from 1995 to 2010 which, interestingly, harbored the ibeA gene. Using this virulence gene as a criterion for selection, we compared those 19 avian strains with 33 human O25b:H4-ST131 ibeA-positive E. coli strains obtained from patients with extraintestinal infections (1993 to 2009). All 52 O25b:H4-ST131 ibeA-positive E. coli strains shared the fimH, kpsMII, malX, and usp genes but showed statistically significant differences in nine virulence factors, namely, papGIII, cdtB, sat, and kpsMII K5, which were associated with human strains, and iroN, kpsMII K1, cvaC, iss, and tsh, which were associated with strains of avian origin. The XbaI macrorestriction profiles of the 52 E. coli O25b:H4-ST131 ibeA-positive strains revealed 11 clusters (clusters I to XI) of >85% similarity, with four clusters including strains of human and avian origin. Cluster VII (90.9% similarity) grouped 10 strains (7 avian and 3 human strains) that mostly produced CTX-M-9 and that also shared the same virulence profile. Finally, we compared the macrorestriction profiles of the 12 CTX-M-9-producing O25b:H4-ST131 ibeA strains (7 avian and 5 human strains) identified among the 52 strains with those of 15 human O25b:H4-ST131 CTX-M-14-, CTX-M-15-, and CTX-M-32-producing strains that proved to be negative for ibeA and showed that they clearly differed in the level of similarity from the CTX-M-9-producing strains. In conclusion, E. coli clonal group O25b:H4-ST131 ibeA has recently emerged among avian isolates with the new acquisition of the K1 capsule antigen and includes CTX-M-9-producing strains. This clonal group represents a real zoonotic risk that has crossed the barrier between human and avian hosts.Strains of the extensively antimicrobial-resistant Escherichia coli clonal group of sequence type (ST) 131 (ST131) belonging to serotype O25b:H4 have recently been recognized to be important human pathogens worldwide (9, 33). Although it is commonly associated with the dissemination of CTX-M-15 extended-spectrum cephalosporin resistance, E. coli O25b:H4-ST131 also occurs as a fluoroquinolone (FQ)-resistant but cephalosporin-susceptible pathogen (5, 22, 26, 27). Currently, it is assumed that O25b:H4-ST131 strains circulate not only among humans but also among animal hosts (13, 21, 37), which would contribute to the ongoing global emergence of O25b:H4-ST131, in the case of regular transmission between animals and humans. Even though CTX-M-15 is the most widely distributed extended-spectrum beta-lactamase (ESBL) linked to this clonal group, other, different variants of CTX-M have recently been reported, such as CTX-M-9, CTX-M-14, and CTX-M-32 (4, 34, 36, 39). Noteworthy was the detection, for the first time on poultry farms, of this clonal group producing CTX-M-9 that had macrorestriction profiles and virulence genes very similar to those observed in clinical human isolates (10).Extraintestinal pathogenic E. coli (ExPEC) strains, which include avian pathogenic E. coli (APEC) and human uropathogenic E. coli (UPEC), septicemic E. coli, and newborn meningitis-causing E. coli (NMEC) strains, exhibit considerable genome diversity and have a wide range of virulence-associated factors (12, 18). While infections caused by APEC strains initially start as a respiratory tract disease which evolves to a systemic infection of the internal organs and, finally, to sepsis, the most frequent origin of human sepsis is urinary tract infection (UTI), especially pyelonephritis (2, 3, 11). However, APEC strains have been recognized to share common traits with human isolates (29, 30, 31), including the K1 capsule antigen (23, 24, 29) and the ibeA gene (14). In addition, retail chicken products have been found to carry nalidixic-resistant ExPEC strains (17, 19), and although it is drug susceptible, an E. coli strain belonging to the O25b:H4-ST131 clonal group has even recently been detected in retail chicken (41), supporting the urgent necessity for the implementation of food control measures.The aim of the present study was to discern the possible spread of the O25b:H4-ST131 clonal group, especially CTX-M-9-producing strains, in poultry and the zoonotic potential of avian isolates. For this purpose, we made a retrospective search of our human and avian strain collections and compared the findings for those strains with the findings for current strains. Identification of this emerging clone among avian sources and comparison of the clone with clinical human isolates will shed new light on the epidemiology of the O25b:H4-ST131 clonal group.     

Biological and structural characterization of the Mycobacterium smegmatis nitroreductase NfnB,and its role in benzothiazinone resistance

Tuberculosis is still a leading cause of death in developing countries, for which there is an urgent need for new pharmacological agents. The synthesis of the novel antimycobacterial drug class of benzothiazinones (BTZs) and the identification of their cellular target as DprE1 (Rv3790), a component of the decaprenylphosphoryl‐β‐d ‐ribose 2′‐epimerase complex, have been reported recently. Here, we describe the identification and characterization of a novel resistance mechanism to BTZ in Mycobacterium smegmatis. The overexpression of the nitroreductase NfnB leads to the inactivation of the drug by reduction of a critical nitro‐group to an amino‐group. The direct involvement of NfnB in the inactivation of the lead compound BTZ043 was demonstrated by enzymology, microbiological assays and gene knockout experiments. We also report the crystal structure of NfnB in complex with the essential cofactor flavin mononucleotide, and show that a common amino acid stretch between NfnB and DprE1 is likely to be essential for the interaction with BTZ. We performed docking analysis of NfnB‐BTZ in order to understand their interaction and the mechanism of nitroreduction. Although Mycobacterium tuberculosis seems to lack nitroreductases able to inactivate these drugs, our findings are valuable for the design of new BTZ molecules, which may be more effective in vivo.     

An integrated view of gene expression and solute profiles of Arabidopsis tumors: a genome-wide approach

Transformation of plant cells with T-DNA of virulent agrobacteria is one of the most extreme triggers of developmental changes in higher plants. For rapid growth and development of resulting tumors, specific changes in the gene expression profile and metabolic adaptations are required. Increased transport and metabolic fluxes are critical preconditions for growth and tumor development. A functional genomics approach, using the Affymetrix whole genome microarray (approximately 22,800 genes), was applied to measure changes in gene expression. The solute pattern of Arabidopsis thaliana tumors and uninfected plant tissues was compared with the respective gene expression profile. Increased levels of anions, sugars, and amino acids were correlated with changes in the gene expression of specific enzymes and solute transporters. The expression profile of genes pivotal for energy metabolism, such as those involved in photosynthesis, mitochondrial electron transport, and fermentation, suggested that tumors produce C and N compounds heterotrophically and gain energy mainly anaerobically. Thus, understanding of gene-to-metabolite networks in plant tumors promotes the identification of mechanisms that control tumor development.     

Oxidative catalysts for the transformation of phenolic pollutants: a brief review

Phenolics are often produced as wastes by several industrial and agricultural activities. Many of these compounds and their derivatives are extremely dangerous to living organisms, because they are highly toxic and thus represent a serious environmental concern.

Conventional remediation methods of phenol-polluted systems have some disadvantages due to high cost, time-consuming procedures and formation of toxic residues. Conversely, the use of oxidative catalysts, both enzymatic or inorganic, is a promising alternative technology to address the clean up of such wastes. Oxidative enzymes and inorganic compounds, both naturally occurring in soil, behave as biotic and abiotic catalysts and support the transformation of phenolic compounds. The complete mineralization of phenolic pollutants as well as the formation of polymeric products, often less toxic than their precursors, may occur.

The present paper gives a brief review of many aspects concerning the properties of biotic and abiotic catalytic agents effective in the transformation of phenolic compounds. The main mechanisms of the processes as well as their feasibility for catalytic practical applications will be addressed. Examples of their potentiality in the detoxification of phenol-polluted systems will be provided, as well.