Signaling via the G protein alpha subunit FGA2 is necessary for pathogenesis in Fusarium oxysporum

Cloning and disruption of fga1, the gene encoding the G protein alpha subunit FGA1 in phytopathogenic fungus Fusarium oxysporum, has been reported previously, and the fga1 disruptants showed altered colony morphology, increased heat resistance, reduced conidiation and pathogenicity. To further evaluate the role of G protein signaling in this fungus, cloning of fga2, which encodes the second Galpha protein FGA2, was performed by PCR methods. The deduced primary structure of FGA2 (355 amino acid residues) showed high identity with other Galpha proteins, which belong to class III of fungal Galpha proteins. Disruption of fga2 led to higher heat resistance, similar to the fga1 disruptants, but pathogenicity was completely lost, unlike the fga1 disruptants. Alteration of colony morphology and conidiation, which was observed in the fga1 disruptants, was not observed in the fga2 disruptants. The fga1/fga2 double disruptants showed phenotypic alterations similar to the fga1 or fga2 single disruptants, but increase of heat resistance was much more pronounced than in each single disruptant.     

'Dipeptidyl peptidase-IV activity and/or structure homologs' (DASH) in growth-modulated glioma cell lines

Dipeptidyl peptidase-IV has been demonstrated to play a role in cancer biology by many authors. Since then, additional proteins possessing similar enzymatic activity have been described and their role in cancerogenesis has been hypothesized. To assess the complexity of these 'Dipeptidyl peptidase-IV activity and/or structure homologs' (DASH) in glioma cells, we have studied their presence in cell lines of different degree of transformation. Our results provide evidence of cell line-specific expression and distribution of dipeptidyl peptidase-IV enzyme activity-bearing molecules and their dynamics associated with cell growth conditions. The biologic outcome of DASH pattern of composition probably depends on the regulatory peptides/DASH substrates in the cellular environment.     

Aging and β3-adrenergic stimulation alter mitochondrial lipidome of adipose tissue

Mitochondrial abundance and thermogenic capacity are two imperative components that distinguish brown, beige and white adipose tissues. Most importantly, the lipid composition is vital for maintaining the quantity, quality and function of mitochondria. Therefore, we employed quantitative lipidomics to probe the mitochondrial lipidome of adipose tissues. The mitochondrial lipidome reveals β3-adrenergic stimulation and aging drastically altered the levels of phosphatidylcholine (PC)/phosphatidylethanolamine (PE) ratio and acyl chain desaturation. Precisely, PC_36:2 and PE_38:4 levels correlate with the increased brown and beige fat activity in young mice. While aging increased lysoPC species in white adipose tissue (WAT) mitochondria, CL-316,243 administration reduced lysoPC species and increased lyso-PE_{18:1 and 18:2} content during WAT browning. Also, non-thermogenic mitochondria accumulate sphingomyelin (SM), phosphatidylserine (PS), phosphatidic acid (PA) and ether-linked PC (ePC). Similarly, enrichment of phosphatidylglycerol (PG) and cardiolipin (CL) levels are associated with thermogenic mitochondria. Also, our in vitro experiment supports that blocking the de novo sphingolipid synthesis pathway by myriocin, SPT1 inhibitor increased the thermogenic capacity and oxygen consumption rate in mature adipocytes. Overall, our study suggests mitochondria of brown, beige and white adipose tissues own a unique pattern of lipid molecular species and their levels are altered by aging and CL-316,243 administration.     

A Prospective Study of the Causes of Febrile Illness Requiring Hospitalization in Children in Cambodia

BackgroundFebrile illnesses are pre-eminent contributors to morbidity and mortality among children in South-East Asia but the causes are poorly understood. We determined the causes of fever in children hospitalised in Siem Reap province, Cambodia.ConclusionWe identified a microbiological cause of fever in almost 50% of episodes in this large study of community-acquired febrile illness in hospitalized children in Cambodia. The range of pathogens, antimicrobial susceptibility, and co-morbidities associated with mortality described will be of use in the development of rational guidelines for infectious disease treatment and control in Cambodia and South-East Asia.     

Arabidopsis Heterotrimeric G-Proteins Play a Critical Role in Host and Nonhost Resistance against Pseudomonas syringae Pathogens

Heterotrimeric G-proteins have been proposed to be involved in many aspects of plant disease resistance but their precise role in mediating nonhost disease resistance is not well understood. We evaluated the roles of specific subunits of heterotrimeric G-proteins using knock-out mutants of Arabidopsis Gα, Gβ and Gγ subunits in response to host and nonhost Pseudomonas pathogens. Plants lacking functional Gα, Gβ and Gγ1Gγ2 proteins displayed enhanced bacterial growth and disease susceptibility in response to host and nonhost pathogens. Mutations of single Gγ subunits Gγ1, Gγ2 and Gγ3 did not alter bacterial disease resistance. Some specificity of subunit usage was observed when comparing host pathogen versus nonhost pathogen. Overexpression of both Gα and Gβ led to reduced bacterial multiplication of nonhost pathogen P. syringae pv. tabaci whereas overexpression of Gβ, but not of Gα, resulted in reduced bacterial growth of host pathogen P. syringae pv. maculicola, compared to wild-type Col-0. Moreover, the regulation of stomatal aperture by bacterial pathogens was altered in Gα and Gβ mutants but not in any of the single or double Gγ mutants. Taken together, these data substantiate the critical role of heterotrimeric G-proteins in plant innate immunity and stomatal modulation in response to P. syringae.     

Flexible functional interactions between G‐protein subunits contribute to the specificity of plant responses

Plants being sessile integrate information from a variety of endogenous and external cues simultaneously to optimize growth and development. This necessitates the signaling networks in plants to be highly dynamic and flexible. One such network involves heterotrimeric G‐proteins comprised of Gα, Gβ, and Gγ subunits, which influence many aspects of growth, development, and stress response pathways. In plants such as Arabidopsis, a relatively simple repertoire of G‐proteins comprised of one canonical and three extra‐large Gα, one Gβ and three Gγ subunits exists. Because the Gβ and Gγ proteins form obligate dimers, the phenotypes of plants lacking the sole Gβ or all Gγ genes are similar, as expected. However, Gα proteins can exist either as monomers or in a complex with Gβγ, and the details of combinatorial genetic and physiological interactions of different Gα proteins with the sole Gβ remain unexplored. To evaluate such flexible, signal‐dependent interactions and their contribution toward eliciting a specific response, we have generated Arabidopsis mutants lacking specific combinations of Gα and Gβ genes, performed extensive phenotypic analysis, and evaluated the results in the context of subunit usage and interaction specificity. Our data show that multiple mechanistic modes, and in some cases complex epistatic relationships, exist depending on the signal‐dependent interactions between the Gα and Gβ proteins. This suggests that, despite their limited numbers, the inherent flexibility of plant G‐protein networks provides for the adaptability needed to survive under continuously changing environments.     

The aqueous garlic, onion and leek extracts release nitric oxide from S-nitrosoglutathione and prolong relaxation of aortic rings

Garlic, onion and leek have beneficial effects in treatment of numerous health disorders. The aim of the present study was to investigate underlying molecular mechanisms. To test the potency of the aqueous garlic, onion and leek extracts to release NO from GSNO we have measured NO oxidation product, NO(2)-, by the Griess reagent method. Further, we studied the ability of garlic extract to relax noradrenaline-precontracted rat aortic rings in the presence of GSNO and effects of garlic extract on electrical properties of rat heart intracellular chloride channels. We have observed that: i) garlic, onion and leek extracts released NO from GSNO in the order: garlic > onion > leek; ii) the ability of garlic extract to release NO was pH-dependent (8.0 > 7.4 > 6.0) and potentiated by thiols (Cys > GSH = N-acetyl-cysteine > oxidized glutathione) at concentration 100 μmol/l; iii) the garlic extract (0.045 mg/ml) prolonged relaxation time of aortic rings induced by GSNO (50 nmol/l) and inhibited intracellular chloride channels. We suggest that NO-releasing properties of the garlic, onion and leek extracts and their interaction with Cys and GSH are involved in NO-signalling pathway which contributes to some of its numerous beneficial biological effects.     

Molecular Pathology of Rare Bleeding Disorders (RBDs) in India: A Systematic Review

Background

Though rare in occurrence, patients with rare bleeding disorders (RBDs) are highly heterogeneous and may manifest with severe bleeding diathesis. Due to the high rate of consanguinity in many caste groups, these autosomal recessive bleeding disorders which are of rare occurrence in populations across the world, may not be as rare in India.

Objectives

To comprehensively analyze the frequency and nature of mutations in Indian patients with RBDs.

Methods

Pubmed search was used (www.pubmed.com) to explore the published literature from India on RBDs using the key words “rare bleeding disorders”, “mutations”, “India”, “fibrinogen”, “afibrinogenemia”, “factor II deficiency”, “prothrombin” “factor VII deficiency”, “factor V deficiency”, “factor X deficiency”, “factor XI deficiency”, “combined factor V and VIII deficiency”, “factor XIII deficiency”, “Bernard Soulier syndrome” and “Glanzmanns thrombasthenia” in different combinations. A total of 60 relevant articles could be retrieved. The distribution of mutations from India was compared with that of the world literature by referring to the Human Gene Mutation Database (HGMD) (www.hgmd.org).

Results

Taken together, 181 mutations in 270 patients with different RBDs have been reported from India. Though the types of mutations reported from India and their percentage distribution with respect to the world data are largely similar, yet much higher percentage of small deletions, duplication mutations, insertions, indels were observed in this analysis. Besides the identification of novel mutations and polymorphisms, several common mutations have also been reported, which will allow to develop a strategy for mutation screening in Indian patients with RBDs.

Conclusion

There is a need for a consortium of Institutions working on the molecular pathology of RBDs in India. This will facilitate a quicker and cheaper diagnosis of RBDs besides its utility in first trimester prenatal diagnosis of the affected families.     

How to narrow down chromosomal breakpoints in small and large derivative chromosomes – a new probe set

Here a new fluorescence in situ hybridization (FISH-) based probe set is presented and its possible applications are highlighted in 34 exemplary clinical cases. The so-called pericentric-ladder-FISH (PCL-FISH) probe set enables a characterization of chromosomal breakpoints especially in small supernumerary marker chromosomes (sSMC), but can also be applied successfully in large inborn or acquired derivative chromosomes. PCL-FISH was established as 24 different chromosome-specific probe sets and can be used in two- up multicolor-FISH approaches. PCL-FISH enables the determination of a chromosomal breakpoint with a resolution between 1 and ～10 megabasepairs and is based on locus-specific bacterial artificial chromosome (BAC) probes. Results obtained on 29 sSMC cases and five larger derivative chromosomes are presented and discussed. To confirm the reliability of PCL-FISH, eight of the 29 sSMC cases were studied by array-comparative genomic hybridization (aCGH); the used sSMC-specific DNA was obtained by glass-needle based microdissection and DOP-PCR-amplification. Overall, PCL-FISH leads to a better resolution than most FISH-banding approaches and is a good tool to narrow down chromosomal breakpoints.