Identification of Two Novel Mutations in PKHD1 Gene from Two Families with Polycystic Kidney Disease by Whole Exome Sequencing

Background Polycystic kidney disease (PKD) is an autosomal recessive disorder resulting from mutations in the PKHD1 gene on chromosome 6 (6p12), a large gene spanning 470 kb of genomic DNA.ObjectiveThe aim of the present study was to report newly identified mutations in the PKHD1 gene in two Iranian families with PKD.Materials and Methods Genetic alterations of a 3-month-old boy and a 27-year-old girl with PKD were evaluated using whole-exome sequencing. The PCR direct sequencing was performed to analyse the co-segregation of the variants with the disease in the family. Finally, the molecular function of the identified novel mutations was evaluated by in silico study. ResultsIn the 3 month-old boy, a novel homozygous frameshift mutation was detected in the PKHD1 gene, which can cause PKD. Moreover, we identified three novel heterozygous missense mutations in ATIC, VPS13B, and TP53RK genes. In the 27-year-old woman, with two recurrent abortions history and two infant mortalities at early weeks due to metabolic and/or renal disease, we detected a novel missense mutation on PKHD1 gene and a novel mutation in ETFDH gene.Conclusion In general, we have identified two novel mutations in the PKHD1 gene. These molecular findings can help accurately correlate genotype and phenotype in families with such disease in order to reduce patient births through preoperative genetic diagnosis or better management of disorders.     

Proteomics reveals new salt responsive proteins associated with rice plasma membrane

The signaling processes in plants that initiate cellular responses to biotic and abiotic factors are believed to be located in the plasma membrane (PM). A better understanding of the PM proteome response to environmental stresses might lead to new strategies for improving stress-tolerant crops. A sub-cellular proteomics approach was applied to monitor changes in abundance of PM-associated protein in response to salinity, a key abiotic stress affecting rice productivity worldwide. Proteome was extracted from a root plasma-membrane-rich fraction of a rice salt tolerant variety, IR651, grown under saline and normal conditions. Comparative two-dimensional electrophoresis revealed that 24 proteins were differentially expressed in response to salt stress. From these, eight proteins were identified by mass spectrometry analysis. Most of the proteins identified are likely to be PM-associated and are known to be involved in several important mechanisms of plant adaptation to salt stress. These include regulation of PM pumps and channels, membrane structure, oxidative stress defense, signal transduction, protein folding, and the methyl cycle. To investigate the correlation between mRNA and protein level in response to salinity, we performed quantitative Real-Time PCR analysis of three genes that were salt responsive at the protein level, including 1,4-Benzoquinone reductase, a putative remorin and a hypersensitive induced response protein. No concordance was detected between the changes in levels of gene and protein expression. Our results indicate that the proteomics approach is suitable for expression analysis of membrane associated proteins under salt stress.     

Synthesis of New Benzimidazole‐1,2,3‐triazole Hybrids as Tyrosinase Inhibitors

A novel series of benzimidazole‐1,2,3‐triazole hybrids containing substituted benzyl moieties were designed, synthesized and evaluated for their inhibitory activity against mushroom tyrosinase. The results indicated that 2‐(4‐{[1‐(3,4‐dichlorobenzyl)‐1H‐1,2,3‐triazol‐4‐yl]methoxy}phenyl)‐1H‐benzimidazole ( 6g ) and 2‐(4‐{[1‐(4‐bromobenzyl)‐1H‐1,2,3‐triazol‐4‐yl]methoxy}phenyl)‐1H‐benzimidazole ( 6h ) exhibited effective inhibitory activity with IC₅₀ values of 9.42 and 10.34 μm , respectively, comparable to that of kojic acid as the reference drug (IC₅₀ = 9.28 μm ). Kinetic study of compound 6g confirmed mixed‐type inhibitory activity towards tyrosinase indicating that it can bind to free enzyme as well as enzyme‐substrate complex. Also, molecular docking analysis was performed to determine the binding mode of the most potent compounds ( 6g and 6h ) in the active site of tyrosinase. Consequently, 6g and 6h derivatives might serve as promising candidates in cosmetics, medicine or food industry, and development of such compounds may be of an interest.     

Gene polymorphisms of macrophage migration inhibitory factor affect susceptibility to chronic hepatitis B virus infection in an Iranian cohort

Macrophage migration inhibitory factor (MIF), a key proinflammatory mediator, plays important roles in chronic diseases. In this study, an attempt was made to clarify the associations between some functional polymorphisms such as MIF‐173 G/C, MIF 95 bp and 189 bp insertion/deletion (I/D) polymorphisms and chronic hepatitis B virus (HBV) infection. Polymorphisms were assessed in 221 HBV patients and 200 normal subjects. MIF‐173 G/C and MIF 95 bp and 189 bp I/D polymorphisms were genotyped using PCR–RFLP and PCR, respectively. When allele and genotype frequencies of the variants were compared between patients and controls by the χ² test, it was found that the frequency of MIF‐173 G/C genotypes differed significantly between patients with chronic HBV and healthy controls (P < 0.05). Carriers of the MIF ‐173‐C allele were at significantly higher risk of HBV infection than carriers of the MIF ‐173‐G allele (P = 0.009, OR = 1.549, 95% CI = 1.114 ? 2.155). Moreover, 95 bp I/D polymorphism was not associated with CP and the 185 bp I/D variant was not polymorphic in our group of subjects. The frequency of haplotypes did not differ significantly between groups (χ² = 11.391, P = 0.181). Our results suggest that MIF ‐173 G/C variant increases the risk of HBV in Iranian subjects. Further studies with larger sample sizes and different ethnicities are required to validate our findings.

     

Persistence and tissue distribution of DNA in normal and immunodeficient mice inoculated with polyomavirus VP1 pseudocapsid complexes or polyomavirus

Introduction of DNA into normal and immunodeficient mice, alone or in complex with VP1 pseudocapsids, has been compared to DNA transfer by viral infection. Similar to natural infection and in contrast to plasmid alone, VP1 pseudocapsids efficiently introduced DNA, which remained for months in normal mice and possibly longer in B- and T-cell-deficient mice.     

Hydrolysis of pyrethroids by carboxylesterases from Lucilia cuprina and Drosophila melanogaster with active sites modified by in vitro mutagenesis

The cloned genes encoding carboxylesterase E3 in the blowfly Lucilia cuprina and its orthologue in Drosophila melanogaster were expressed in Sf9 cells transfected with recombinant baculovirus. Resistance of L. cuprina to organophosphorus insecticides is due to mutations in the E3 gene that enhance the enzyme's ability to hydrolyse insecticides. Previous in vitro mutagenesis and expression of these modifications (G137D, in the oxyanion hole and W251L, in the acyl pocket) have confirmed their functional significance. We have systematically substituted these and nearby amino acids by others expected to affect the hydrolysis of pyrethroid insecticides. Most mutations of G137 markedly decreased pyrethroid hydrolysis. W251L was the most effective of five substitutions at this position. It increased activity with trans permethrin 10-fold, and the more insecticidal cis permethrin >130-fold, thereby decreasing the trans:cis hydrolysis ratio to only 2, compared with >25 in the wild-type enzyme. Other mutations near the bottom of the catalytic cleft generally enhanced pyrethroid hydrolysis, the most effective being F309L, also in the presumptive acyl binding pocket, which enhanced trans permethrin hydrolysis even more than W251L. In these assays with racemic 1RS cis and 1RS trans permethrin, two phases were apparent, one being much faster suggesting preferential hydrolysis of one enantiomer in each pair as found previously with other esterases. Complementary assays with individual enantiomers of deltamethrin and the dibromo analogue of cis permethrin showed that the wild type and most mutants showed a marked preference for the least insecticidal 1S configuration, but this was reversed by the F309L substitution. The W251L/F309L double mutant was best overall in hydrolysing the most insecticidal 1R cis isomers. The results are discussed in relation to likely steric effects on enzyme-substrate interactions, cross-resistance between pyrethroids and malathion, and the potential for bioremediation of pyrethroid residues.     

Comparison of pathogenicity of Clavibacter michiganensis subsp. insidiosus and Pseudomonas viridiflava strains on alfalfa

Hamedan province of Iran is a suitable niche for alfalfa growth but many diseases including alfalfa bacterial wilt, bacterial crown and root rot diseases cause economic crop losses. Bacterial wilt is caused by Clavibacter michiganensis subsp. insidiosus, and bacterial crown and root rot diseases are caused by Pseudomonas viridiflava. In this study, we investigated the pathogenicity of C. michiganensis subsp. insidiosus and P. viridiflava strains collected from the main alfalfa growing areas of Hamedan province. Pathogenicity of the virulent strains was tested on alfalfa and the bacterial strains caused symptoms, and data were collected about stem length, root length, wet weight and dry weight of the infected plantlets. The data about the pathogenicity of C. michiganensis subsp. insidiosus and P. viridiflava on alfalfa were compared with each other and were analysed by SAS software and Dunkan's test. Resulted data showed more pathogenicity of C. michiganensis subsp. insidiosus than P. viridiflava on alfalfa. These data also showed that both of these bacteria produced the most losses on wet weight and dry weight of alfalfa plantlets.     

A Comparison between the Nephrotoxic Profile of Gentamicin and Gentamicin Nanoparticles in Mice

Aminoglycoside antibiotics are widely used against Gram‐negative infections. On the other hand, nephrotoxicity is a deleterious side effect associated with aminoglycoside therapy. Gentamicin is the most nephrotoxic aminoglycoside. Because of serious health complications ensue the nephrotoxicity induced by aminoglycosides, finding new therapeutic strategies against this problem has a great clinical value. This study has attempted to compare the nephrotoxic properties of gentamicin and a new nanosized formulation of this drug in a mice model. Animals were treated with gentamicin (100 mg/kg, i.p. for eight consecutive days) and nanogentamicin (100 mg/kg, i.p. for eight consecutive days). Blood urea nitrogen (BUN), plasma creatinine levels, and histopathological changes of kidney proximal tubule were monitored. It was found that gentamicin caused severe degeneration of kidney proximal tubule cells and an increase in serum creatinine and BUN. No severe injury was observed after nanogentamicin administration. This study proved that nanosized gentamicin is less nephrotoxic.     

The effect of different iron concentrations on lead accumulation in hydroponically grown <Emphasis Type="Italic">Matthiola flavida</Emphasis> Boiss

The general relationship between heavy metals and mineral nutrition of plants grown in polluted environments is one of the most important factors for modifying the toxic properties of these metals. To study the effect of iron and lead pollution on the growth of Matthiola flavida a factorial research was undertaken in the form of a completely randomized design with four replications in hydroponic culture. After germination, seedlings were transferred to a hydroponic culture. During the pre-treatment step, a series of plants contained enough iron (+Fe), whereas the second series was without iron (?Fe). After the pre-treatment step, both series of plants were treated with three levels of iron as FeEDDHA and two levels of lead as Pb(NO₃)₂. The results showed that in both series of plants, lead reduced the root growth, shoot height, shoot and root dry weight. For 5 µM lead concentration, with increasing concentration of iron in the nutrient solution, concentration of lead in the roots and shoots decreased. For 1 µM concentration of lead, increasing the iron concentration in the nutrient solution reduced the concentration of lead in the roots, but had no significant effect on the amount of lead in shoots. Lead accumulation in shoots and roots of plants with Fe was more than plants without Fe. Also, in low concentrations of iron, the amount of iron in shoot increased with the increase of lead concentration in the nutrient solution. The results showed that in lead pollution, iron has a positive effect on investigated traits.     

How is the relationship between TWIST-1 and BCR-ABL1 gene expressions in chronic myeloid leukaemia patients?

Background: The activation and increased expression of BCR-ABL1 lead to malignant chronic myelogenous leukaemia (CML) cells, as well as the resistance to antitumour agents and apoptosis inducers. Moreover, TWIST-1 protein is a prognostic factor of leukemogenesis, and its level is raised in CML patients with cytogenetic resistance to imatinib. So, there is a likely relationship between BCR-ABL1 and TWIST-1 genes.

Objective: The aim of the study was to assess the relationship between TWIST-1 and BCR-ABL1 expressions.

Methods: Peripheral blood samples were obtained from 44 CML patients under treatment and also from ten healthy subjects as normal controls. The expression of TWIST-1 and BCR-ABL1 genes was measured using real-time PCR, and ABL1 was used as the reference gene. The gene expression was evaluated by REST software.

Results: The expression levels of TWIST-1 and BCR-ABL1 genes in CML patients was changed 40.23?±?177.75-fold and 6?±?18-fold, respectively.

Discussion: No significant relationship was observed between the expressions of TWIST-1 and BCR-ABL1 genes. All patients with TWIST-1 expression levels?≥100-fold had failure of response to treatment.

Conclusion: The probability of the relationship between BCR-ABL1 and TWIST-1 is still debatable, and the average of TWIST-1 expression has been higher in patients without response to treatment. Definitive conclusion needs further investigations.