Association Study of rs1333040 and rs1004638 Polymorphisms in the 9p21 Locus with Coronary Artery Disease in Southwest of Iran

Background:

Coronary artery disease (CAD) is a multifactorial and heterogenic disease. Recently, genome-wide association studies have reported that rs1333040 (C/T) and rs1004638 (A/T) single nucleotide polymorphisms (SNPs) in the 9p21 locus have very strong association with CAD. This study aimed to examine these associations in Southwest of Iran.

Methods:

Blood samples were collected from 200 CAD patients and 110 healthy individuals with no CAD. The association of two SNPs with CAD was evaluated by PCR and restriction fragment length polymorphism.

Results:

Chi-square test showed no association between rs1333040 SNP and CAD (X²: 4.66, df: 2, P=0.09). Also, there was no association between rs1004638 SNP and CAD (X²: 0.27, df: 2, P=0.88).

Conclusion:

No association was observed between rs1333040 and rs1004638 SNPs in the 9P21 region and CAD in Southwest of Iran. Key Words: Coronary artery disease, Single nucleotide polymorphisms, Genetic association study, Iran     

The role of Ile3434Thr XRCC7 gene polymorphism in Differentiated Thyroid Cancer risk in an Iranian population

Background

The aim of this study was to understand any association between differentiated thyroid carcinoma (DTC) and Ile3434Thr XRCC7 gene polymorphism (GenBank accession number: rs7830743). DTC is the most prevalent thyroid neoplasm, which includes papillary and follicular cell carcinoma. XRCC7 gene encodes a protein that functions in non-homologous end joining DNA repair pathway. Non-synonymous polymorphisms in this gene may alter DNA repair capacity of the cell and change the risk of developing cancers.

Methods

DTC patients (n = 173) and cancer free individuals (n = 204) were enrolled in a case-control study. The Ile3434Thr polymorphic alleles were discriminated by using amplification refractory mutation system-PCR method. The frequencies of this single nucleotide polymorphism in case and control groups were compared. Also, risk ratio for developing DTC in dichotomized genotypes was estimated by multivariate logistic regression analysis.

Results

Dichotomized genotypes into those with and without the 3434Thr allele showed that this allele was associated with DTC (OR [odd ratio]: 1.89, 95% confidence interval (CI) = 1.29-2.79, P<0.001). Also, TC genotype was significantly associated with increased risk of DTC (OR: 2.42, 95% CI = 1.55-3.81, P = 0.0001) in individuals carrying this genotype.

Conclusion

Allele 3434Thr in XRCC7 gene might be associated with differentiated thyroid cancer risk. Further studies with larger samples are needed to verify these initial findings. Key Words: DNA repair enzymes, Thyroid neoplasms, Genetic polymorphism     

OsSUV3 transgenic rice maintains higher endogenous levels of plant hormones that mitigates adverse effects of salinity and sustains crop productivity

Background

The SUV3 (suppressor of Var 3) gene encodes a DNA and RNA helicase, which is localized in the mitochondria. Plant SUV3 has not yet been characterized in detail. However, the Arabidopsis ortholog of SUV3 (AT4G14790) has been shown to be involved in embryo sac development. Previously, we have reported that rice SUV3 functions as DNA and RNA helicase and provides salinity stress tolerance by maintaining photosynthesis and antioxidant machinery. Here, we report further analysis of the transgenic OsSUV3 rice plants under salt stress.

Findings

The transgenic OsSUV3 overexpressing rice T₁ lines showed significantly higher endogenous content of plant hormones viz., gibberellic acid (GA₃), zeatin (Z) and indole-3-acetic acid (IAA) in leaf, stem and root as compared to wild-type (WT), vector control (VC) and antisense (AS) plants under salt (200 mM NaCl) stress condition. A similar trend of endogenous plant hormones profile was also reflected in the T₂ generation of OsSUV3 transgenic rice under defined parameters and stress condition.

Conclusions

In response to stress, OsSUV3 rice plants maintained plant hormone levels that regulate the expression of several stress-induced genes and reduce adverse effects of salt on plant growth and development and therefore sustains crop productivity.     

Metabolomics-Based Study of Logarithmic and Stationary Phases of Promastigotes in Leishmania major by 1H NMR Spectroscopy

Background:

Cutaneous leishmaniasis is one of the most important parasitic diseases in humans. In this disease, one of the responsible organisms is Leishmania major, which is transmitted by sandfly vector. There are specific differences in biochemical profiles and metabolite pathways in logarithmic and stationary phases of Leishmania parasites. In the present study, ¹H NMR spectroscopy was used to examine the metabolites outliers in the logarithmic and stationary phases of promastigotes in L. major to enlighten more about the transmission mechanism in metacyclogenesis of L. major.

Methods:

Promastigote was cultured, logarithmic and stationary phases were separated by the peanut agglutinin, and cell metabolites were extracted. ¹H NMR spectroscopy was applied, and outliers were analyzed using principal component analysis.

Results:

The most altered metabolites in stationary and logarithmic phases were limited to citraconic acid, isopropylmalic acid, L-leucine, ornithine, caprylic acid, capric acid, and acetic acid.

Conclusion:

¹H NMR spectroscopy could play an important role in the characterization of metabolites in biochemical pathways during a metacyclogenesis process. These metabolites and their pathways can help in exploiting a transmission mechanism in metacyclogenesis, and outcoming data might be used in the metabolic network reconstruction of L. major modeling. Key Words: Leishmania major, Metabolomics, Principal component analysis     

Nomenclature report on rice WRKY's - Conflict regarding gene names and its solution

Background

Since whole genome sequences of rice were made publically accessible, the number of articles on new rice genes has increased remarkably. The Committee on Gene Symbolization, Nomenclature and Linkage (CGSNL) of the Rice Genetics Cooperative published the gene nomenclature system for rice and encouraged researchers to follow the rules before publishing their results. The CGSNL provides an on-line registration system for newly identified rice genes to prevent conflicts and/or duplication of gene name in journal articles.

Findings

Recently, the CGSNL surveyed genes in the rice WRKY family in published journal articles and found several duplicated gene names.

Conclusions

To discuss and resolve inconsistencies in WRKY gene nomenclature, the rice WRKY working group was established and redefined the nomenclature. This report announces the conclusion.     

Different Morphologies of Leishmania major Amastigotes with No Molecular Diversity in a Neglected Endemic Area of Zoonotic Cutaneous Leishmaniasis in Iran

Background:

Molecular diversity of Leishmania major and its morphological changes have become a controversial issue among researchers. Some aspects of polymorphic shapes of amastigotes in clinical manifestations along with molecular variation were evaluated among suspected patients of some exceptional zoonotic cutaneous leishmaniasis locations in Northern Khuzestan, Southwestern Iran.

Methods:

Suspected patients (n = 165) were sampled in zoonotic cutaneous leishmaniasis foci over two consecutive years during 2012-2014. Prepared smears were stained, scaled and measured by ocular micrometer. DNA was extracted from smears; ITS-rDNA and Cytochrome b (Cyt b) markers were amplified, and PCR products were digested by BsuR1 restriction enzyme. Then the RFLP and sequencing were employed.

Results:

Only L. major was identified in patients containing regular amastigotes'' shapes (oval or round) with a size of 2-4 µm in each of classical wet, dry, mixed lesions. Meanwhile, irregular shapes (spindle, pear, or cigarette) were observed separately in non-classical wet lesions with more than 4 µm. Interestingly, a few amastigotes with an external flagellum were observed in some lesions. All sequenced ITS-rDNA and Cyt b genes of L. major did not show any molecular variation (χ ² P > 0.05), including only one common haplotype (GenBank access no. {"type":"entrez-nucleotide","attrs":{"text":"EF413075","term_id":"151936076","term_text":"EF413075"}}EF413075).

Conclusion:

Findings proved that unlike other endemic foci, there is not a meaningful correlation between phenotypic and genotypic features of L. major isolates. This study is considered as the first comprehensive report to incriminate morphometric shapes of L. major amastigotes, which enhances our knowledge concerning their relevance with various clinical appearances and genotypic traits. Key Words: Leishmania major, Nuclear gene, Mitochondrial gene, Amastigote shapes, Iran     

Isolation of a novel mutant gene for soil-surface rooting in rice (Oryza sativa L.)

Background

Root system architecture is an important trait affecting the uptake of nutrients and water by crops. Shallower root systems preferentially take up nutrients from the topsoil and help avoid unfavorable environments in deeper soil layers. We have found a soil-surface rooting mutant from an M₂ population that was regenerated from seed calli of a japonica rice cultivar, Nipponbare. In this study, we examined the genetic and physiological characteristics of this mutant.

Results

The primary roots of the mutant showed no gravitropic response from the seedling stage on, whereas the gravitropic response of the shoots was normal. Segregation analyses by using an F₂ population derived from a cross between the soil-surface rooting mutant and wild-type Nipponbare indicated that the trait was controlled by a single recessive gene, designated as sor1. Fine mapping by using an F₂ population derived from a cross between the mutant and an indica rice cultivar, Kasalath, revealed that sor1 was located within a 136-kb region between the simple sequence repeat markers RM16254 and 2935-6 on the terminal region of the short arm of chromosome 4, where 13 putative open reading frames (ORFs) were found. We sequenced these ORFs and detected a 33-bp deletion in one of them, Os04g0101800. Transgenic plants of the mutant transformed with the genomic fragment carrying the Os04g0101800 sequence from Nipponbare showed normal gravitropic responses and no soil-surface rooting.

Conclusion

These results suggest that sor1, a rice mutant causing soil-surface rooting and altered root gravitropic response, is allelic to Os04g0101800, and that a 33-bp deletion in the coding region of this gene causes the mutant phenotypes.     

Genetic interaction between rice PLASTOCHRON genes and the gibberellin pathway in leaf development

Background

The rice PLASTOCHRON (PLA) genes PLA1 and PLA2 regulate leaf maturation and the temporal pattern of leaf initiation. Although the function of PLA genes in the leaf initiation process has been analyzed, little is known about how they affect leaf growth. Previously, we suggested that PLA1 and PLA2 function downstream of the gibberellin (GA) signal transduction pathway. In the present study, we examined the phenotype of a double mutant of pla and slender rice 1 (slr1), which is a constitutive GA response mutant. By analyzing these double mutants, we discuss the relationship between PLA-related and GA-dependent pathways and the possible function of PLA genes in leaf growth.

Findings

Single slr1 and pla mutants exhibited elongated and dwarf phenotypes in the vegetative stage, respectively. The stature and leaf size of the pla1/slr1 and pla2/slr1 double mutants were intermediate between those of the pla and slr1 single mutants. However, the effects of slr1 on leaf elongation were markedly suppressed in the pla1 and pla2 mutant backgrounds. On the other hand, the change in cell length in the double mutants was almost the same as that in the single mutants. An expression analysis of genes involved in GA biosynthesis and catabolism indicated that feedback regulation functioned normally in the pla/slr1 double mutants.

Conclusions

Our genetic results confirm that PLA genes regulate leaf growth downstream of the GA pathway. Our findings also suggest that PLA1 and PLA2 are partly required for GA-dependent leaf elongation, mainly by affecting cellular proliferation.     

A Novel Mutation in the Aprataxin (APTX) Gene in an Iranian Individual Suffering Early-Onset Ataxia with Oculomotor Apraxia Type 1(AOA1) Disease

Background

Ataxia with oculomotor apraxia type 1 (AOA1) shows early onset with autosomal recessive inheritance and is caused by a mutation in the aprataxin (APTX) gene encoding for the APTX protein.

Methods

In this study, a 7-year-old girl born of a first-cousin consanguineous marriage was described with early-onset progressive ataxia and AOA, with increased cholesterol concentration and decreased albumin concentration in serum. PCR and direct DNA sequencing was performed after DNA extraction.

Results

Sequencing analysis revealed a novel homozygous deletion in c.643 and A>T single nucleotide polymorphism in c.641 in exon 6 of the APTX gene [ENST00000379825].

Conclusion

It seems that this region of exon 6 is probably a hot spot; however, no deletions have been reported in exon 6 yet. Key Words: Ataxia oculomotor apraxia 1 (AOA1), aprataxin (APTX), Iranian     

Quality Control of Widely Used Therapeutic Recombinant Proteins by a Novel Real-Time PCR Approach

Background:

Existence of bacterial host-cell DNA contamination in biopharmaceutical products is a potential risk factor for patients receiving these drugs. Hence, the quantity of contamination must be controlled under the regulatory standards. Although different methods such as hybridization assays have been employed to determine DNA impurities, these methods are labor intensive and rather expensive. In this study, a rapid real-time PCR test was served as a method of choice to quantify the E. coli host- cell DNA contamination in widely used recombinant streptokinase (rSK), and alpha interferon (IFN-α) preparations.

Methods:

A specific primer pair was designed to amplify a sequence inside the E. coli 16S rRNA gene. Serial dilutions of DNA extracted from E. coli host cells, along with DNA extracted from Active Pharmaceutical Ingredients of rSK, and IFN-α samples were subjected to an optimized real-time PCR assay based on SYBR Green chemistry.

Results:

The test enabled us to detect a small quantity of genomic DNA contamination as low as 0.0002 pg in recombinant protein-based drugs. For the first time, this study showed that DNA contamination in rSK and IFN-α preparation manufactured in Pasteur Institute of Iran is much lower than the safety limit suggested by the US FDA.

Conclusion:

Real-time PCR is a reliable test for rapid detection of host-cell DNA contamination, which is a major impurity of therapeutic recombinant proteins to keep manufacturers’ minds on refining drugs, and provides consumers with safer biopharmaceuticals. Key Words: DNA contamination, Real-time PCR, Streptokinase, Interferon-alpha (IFN-α)     

Molecular Genetic Analysis of the Variable Number of Tandem-Repeat Alleles at the Phenylalanine Hydroxylase Gene in Iranian Azeri Turkish Population

Background:

The variable numbers of tandem-repeat (VNTR) alleles at the phenylalanine hydroxylase (PAH) gene have been used in carrier detection and prenatal diagnosis in phenylketonuria families. This study was carried out to analyze VNTR alleles at the PAH gene in Iranian Azeri Turkish population.

Methods:

In this study, 200 alleles from general population were studied by PCR.

Results:

The frequencies of VNTR alleles were 45%, 46%, 2%, 3%, 1%, and 3% in studied group regarding 3, 8, 9, 11, 12, and 13 repeat copies, respectively. Statistically significant differences were not found between expected and observed frequencies of VNTR genotypes (P > 0.05).

Conclusions:

VNTR alleles with three and eight repeats were frequent, and the VNTR alleles with 13 repeats showed 3% frequency in the tested group. This study is the first report on tested population genetic structure using VNTR alleles at the PAH gene. Key Words: Phenylalanine hydroxylase, Population genetics, Variable numbers of tandem-repeat     

Impact of Methylenetetrahydrofolate Reductase C677T Polymorphism on the Risk of Gastric Cancer and Its Interaction with Helicobacter pylori Infection

Background

Attempts for early detection of gastric cancer have recently focused on host''s genetic susceptibility factors and gene-environment interactions. We have, herein, studied the association of MTHFR C677T single nucleotide polymorphism (SNP) and its interaction with Helicobacter pylori infection, smoking, age and gender on the risk of gastric cancer among an Iranian population.

Methods

Gastric cancer patients (n = 450) and cancer-free controls (n = 780) were studied for serum H. pylori-specific IgG antibodies by ELISA and MTHFR C677T polymorphism (SNP) by PCR-RFLP. Demographic and life style data were collected through patient interviews. Unconditional logistic regression model estimated odds ratio (OR) and the corresponding 95% confidence intervals (CI).

Results

The interactions of MTHFR genotype with H. pylori infection (P = 0.03), age (P = 0.049) and gender (P = 0.007) were statistically significant. Accordingly, MTHFR C677T carriers who were also positive for H. pylori infection exhibited 80% (OR = 1.8, 95% CI = 1.0-2.9) significant excess risk of non-cardia gastric cancer. Furthermore, subjects over the age of 50 or female subjects carrying MTHFR C677T SNP showed 40 (OR = 1.4, 95% CI = 1.0-2.0) and 100 (OR = 2.0, 95% CI = 1.2-3.2) percent increased risk of gastric cancer, respectively.

Conclusion

MTHFR C677T SNP seems to increase the risk of gastric cancer and the effect is significantly inflated by interactions with H. pylori infection, age and gender. Key Words: Helicobacter pylori, Smoking, Gender identity, Age group, Methylenetetrahydrofolate reductase     

Protective Effect of Vitamin E against Diabetes-Induced Oxidized LDL and Aorta Cell Wall Proliferation in Rat

Background:

Hyperlipidemia and oxidized-low-density lipoproteins (Ox-LDL) are important independent cardiovascular risk factors that have been shown to stimulate vascular smooth muscle cell (VSMC) proliferation. The purpose of the present study was to investigate the effect of vitamin E on Ox-LDL, lipid profile, C-reactive protein (CRP), and VSMC proliferation of rat aorta.

Methods:

Male Wistar rats (n = 32) were divided into four groups namely: sham (SH), control (C), non-treated diabetic, and vitamin E-treated diabetic (VETD) groups. Ox-LDL, lipid profile, CRP and VSMC proliferation of aorta were measured after 42 days.

Results:

The results revealed that along with a significant increase in VSMC proliferation, the amount of CRP, Ox-LDL, and lipid profiles in diabetic rats. VSMC proliferation was significantly ameliorated, and elevated CRP, Ox-LDL, and lipid profiles were also restored to those of shams in VETD.

Conclusions:

These findings strongly support the idea that diabetes induces Ox-LDL-mediated oxidative stress and VSMC proliferation in aorta of rat and imply that vitamin E has a strong protective effect as an antioxidant. Key Words: Ox-LDL, Vitamin E, Diabetes, VSMC proliferation     

Genome duplication improves rice root resistance to salt stress

Background

Salinity is a stressful environmental factor that limits the productivity of crop plants, and roots form the major interface between plants and various abiotic stresses. Rice is a salt-sensitive crop and its polyploid shows advantages in terms of stress resistance. The objective of this study was to investigate the effects of genome duplication on rice root resistance to salt stress.

Results

Both diploid rice (HN2026-2x and Nipponbare-2x) and their corresponding tetraploid rice (HN2026-4x and Nipponbare-4x) were cultured in half-strength Murashige and Skoog medium with 150 mM NaCl for 3 and 5 days. Accumulations of proline, soluble sugar, malondialdehyde (MDA), Na⁺ content, H⁺ (proton) flux at root tips, and the microstructure and ultrastructure in rice roots were examined. We found that tetraploid rice showed less root growth inhibition, accumulated higher proline content and lower MDA content, and exhibited a higher frequency of normal epidermal cells than diploid rice. In addition, a protective gap appeared between the cortex and pericycle cells in tetraploid rice. Next, ultrastructural analysis showed that genome duplication improved membrane, organelle, and nuclei stability. Furthermore, Na⁺ in tetraploid rice roots significantly decreased while root tip H⁺ efflux in tetraploid rice significantly increased.

Conclusions

Our results suggest that genome duplication improves root resistance to salt stress, and that enhanced proton transport to the root surface may play a role in reducing Na⁺ entrance into the roots.     

Association of Lecithin Cholesterol Acyltransferase rs5923 Polymorphism in Iranian Individuals with Extremely Low High-Density Lipoprotein Cholesterol: Tehran Lipid and Glucose Study

Background:

The serum concentration of high-density lipoprotein cholesterol (HDL-C) is one of the important heritable risk factors for cardiovascular disease and is a target for therapeutic intervention. In this study, we aimed to evaluate the effects of lecithin cholesterol acyltransferase (LCAT) gene polymorphism rs5923 on LCAT enzyme activity and serum HDL-C concentration.

Methods:

The study population was selected from consecutive individuals with HDL-C ≤ 5^th percentile (n = 73) and extremely high HDL-C ≥ 95^th percentile (n = 57) who had participated in the Tehran Lipid and Glucose Study. The rs5923 polymorphism was genotyped using direct sequencing. LCAT activity was measured by fluorometric assay kit, and lipid concentrations were measured using the enzymatic colorimetric method.

Results:

The genotype frequencies were significantly different between the high HDL-C group (CC 94.7%, CT 5.3%) and the low HDL-C group (CC 83.6%, CT 16.4%) (P = 0.048). The T-allele frequencies in subjects with low and high HDL-C were 0.082 and 0.026, respectively (P = 0.16). The association of the single-nucleotide polymorphism rs5923 with low HDL-C was not statistically significant after adjustment for age, sex, and BMI (odd ratio = 2.65, 95% confidence interval = 0.32-21.5, P = 0.36, regression logistic analysis). Also, the effects of LCAT enzyme activity did not depend on the HDL-C level (P = 0.24).

Conclusion:

rs5923 polymorphism is not associated with low HDL-C levels in Iranian population. Key Words: Polymorphism, Single nucleotide, Lipoproteins     

Nuclear and chloroplast diversity and phenotypic distribution of rice (Oryza sativa L.) germplasm from the democratic people’s republic of Korea (DPRK; North Korea)

Background

Rice accounts for 43% of staple food production in the Democratic People’s Republic of Korea (DPRK). The most widely planted rice varieties were developed from a limited number of ancestral lines that were repeatedly used as parents in breeding programs. However, detailed pedigrees are not publicly available and little is known about the genetic, phenotypic, and geographical variation of DPRK varieties.

Results

We evaluated 80 O. sativa accessions from the DPRK, consisting of 67 improved varieties and 13 landraces. Based on nuclear SSR analysis, we divide the varieties into two genetic groups: Group 1 corresponds to the temperate japonica subpopulation and represents 78.75% of the accessions, while Group 2 shares recent ancestry with indica varieties. Interestingly, members of Group 1 are less diverse than Group 2 at the nuclear level, but are more diverse at the chloroplast level. All Group 2 varieties share a single Japonica maternal-haplotype, while Group 1 varieties trace maternal ancestry to both Japonica and Indica. Phenotypically, members of Group 1 have shorter grains than Group 2, and varieties from breeding programs have thicker and wider grains than landraces. Improved varieties in Group 1 also show similar and/or better levels of cold tolerance for most traits, except for spikelet number per panicle. Finally, geographic analysis demonstrates that the majority of genetic variation is located within regions that have the most intensive rice cultivation, including the Western territories near the capital city Pyungyang. This is consistent with the conscious and highly centralized role of human selection in determining local dispersion patterns of rice in the DPRK.

Conclusions

Diversity studies of DPRK rice germplasm revealed two genetic groups. The most widely planted group has a narrow genetic base and would benefit from the introduction of new genetic variation from cold tolerant landraces, wild accessions, and/or cultivated gene pools to enhance yield potential and performance.     

Effects of Peptone Supplementation in Different Culture Media on Growth,Metabolic Pathway and Productivity of CHO DG44 Cells; a New Insight into Amino Acid Profiles

Background:

The optimization of bioprocess conditions towards improved growth profile and productivity yield is considered of great importance in biopharmaceutical manufacturing. Peptones as efficient sources of nutrients have been studied for their effect on media development; however, their role on metabolic pathway is not well understood.

Methods:

In the present study, the effect of different concentration of peptones on a recombinant Chinese hamster ovary (CHO) cell line grown in three serum-free suspension cultures was determined. Six peptones of different origins and available amino acid profiles were investigated regarding their impact on cell growth, productivity, and metabolic pathways changes.

Results:

In optimized feeding strategies, increases of 136% and 159% in volumetric productivity (for a low-nutrient culture media) and 55% (for a high-nutrient culture media) were achieved. Furthermore, particular sources of peptones with specific amino acid profile developed preferential results for each different culture medium. Two peptones, SoyA2SC and SoyE-110, were the only hydrolysates that showed production improvement in all three media. Casein Peptone plus Tryptone N1 and SoyA3SC showed different improved results based on their implemented concentration for each individual basal medium.

Conclusion:

The amino acid profile of peptones may provide clues to identify the most effective feeding strategies for recombinant CHO cells.Key Words: CHO cells, Culture media, Peptones, Recombinant proteins