Two- and three-locus haplotypes of the paraoxonase (PON1) gene are associated with coronary artery disease in Asian Indians

Introduction

In view of the reported association of SNPs in the paraoxonase (PON1) gene with coronary artery disease (CAD), and the absence of conclusive data from India, we investigated the relationship of three SNPs at different loci (‐108C/T, L55M and Q192R) of the PON1 gene and their haplotypes with CAD among people residing in the northern plains of India.

Materials and methods

One hundred and seventy-eight healthy controls and two hundred and four angiographically-proven CAD patients were genotyped using PCR-RFLP.

Results

Of the three SNPs, only the R allele of Q192R polymorphism was associated with CAD (p < 0.05). Two locus haplotypes QT (OR 0.55, p = 0.0004, 95% CI 0.39–0.77, significant) and LQ (odds ratio 0.73, p = 0.03, 95% CI 0.55–0.97, trend) showed protective effects, while haplotypes MR (OR = 5.36, p = 0.0001, 95% CI 2.045–14.049) and MC (OR = 2.71, p = 0.011, 95% CI 1.221–6.046) were associated with increased risk of CAD. MRT, a minor three-locus haplotype also displayed significant association (OR 4.93, 95% CI 1.7–13.5) with the disease. Significance was assessed after applying Bonferroni's correction.

Conclusions

Our study revealed that only one SNP at a single locus but several haplotype combinations of PON1 coding and promoter-region polymorphisms were associated with the risk of or protection against CAD. Thus, haplotype analysis brought better insights into the association of PON1 gene polymorphisms with CAD in Asian Indians.     

Use of a new retrieving adaptor in the cloning of a synthetic human insulin A-chain gene

A DNA duplex encoding the A-chain of human insulin was constructed from eight chemically synthesized oligomers by enzymatic ligation to form a partial duplex followed by repair synthesis to complete the complementary strands. After sequential addition of translation start and stop signal adaptors the assembly was cloned in pBR322. To regenerate the end of the coding sequence by precise removal of extraneous nucleotides a new method using a synthetic retrieval adaptor was developed. The procedure included filling in the cohesive ends of the EcoRI site by repair synthesis, ligating a symmetrical adaptor having an MboII recognition sequence to the resulting blunt end, cutting with MboII and removing the single protruding 3′-nucleotide using the 3′ exonuclease activity of DNA polymerase I. Synthetic oligomers useful for ligation to a synthetic insulin C-chain gene were added to the retrieved end of the gene. Sequence analysis established that retrieval adaptors of this type may be used for precise excision of up to eight nucleotides from the end of a cloned DNA fragment.     

Karyotype and meiotic mechanism in Muga silkmoths,Antheraea compta Roth. and A. assamensis (Helf.) (Lepidoptera: Saturniidae)

The chromosome number in males of Antheraea compta (Wild) and A. assamensis (semi-domesticated) is 30, while in females it is 30 (XY) in A. compta and 29 (XO) in A. assamensis, the latter concomitantly does not reveal sex chromatin in interphase nuclei of both somatic and germ cells. This confirms that the Y chromosome, which forms the sex chromatin in females of A. compta, has been lost in the other species. Meiosis in males shows discrete chiasmata while in females of both species it is achiasmatic. Meiotic details of the species are given and the evolutionary inter-relationship is discussed.     

Transposition studies of mini-Mu plasmids constructed from the chemically synthesized ends of bacteriophage Mu

We describe below the chemical synthesis of the right and left ends of bacteriophage Mu and characterize the activity of these synthetic ends in mini-Mu transposition. Mini-Mu plasmids were constructed which carry the synthetic Mu ends together with the Mu A and B genes under control of the bacteriophage λ p_L promoter. Derepression of p_L leads to a high frequency of mini-Mu transposition (5.6 × 10⁻²) which is dependent on the presence of the Mu ends and the Mu A and B proteins. Five deletion mutants in the Mu ends were tested in the mini-Mu transposition system and their effects on transposition are described.     

Structural Basis and Targeting of the Interaction between Fibroblast Growth Factor-inducible 14 and Tumor Necrosis Factor-like Weak Inducer of Apoptosis

Deregulation of the TNF-like weak inducer of apoptosis (TWEAK)-fibroblast growth factor-inducible 14 (Fn14) signaling pathway is observed in many diseases, including inflammation, autoimmune diseases, and cancer. Activation of Fn14 signaling by TWEAK binding triggers cell invasion and survival and therefore represents an attractive pathway for therapeutic intervention. Based on structural studies of the TWEAK-binding cysteine-rich domain of Fn14, several homology models of TWEAK were built to investigate plausible modes of TWEAK-Fn14 interaction. Two promising models, centered on different anchoring residues of TWEAK (tyrosine 176 and tryptophan 231), were prioritized using a data-driven strategy. Site-directed mutagenesis of TWEAK at Tyr¹⁷⁶, but not Trp²³¹, resulted in the loss of TWEAK binding to Fn14 substantiating Tyr¹⁷⁶ as the anchoring residue. Importantly, mutation of TWEAK at Tyr¹⁷⁶ did not disrupt TWEAK trimerization but failed to induce Fn14-mediated nuclear factor κ-light chain enhancer of activated B cell (NF-κB) signaling. The validated structural models were utilized in a virtual screen to design a targeted library of small molecules predicted to disrupt the TWEAK-Fn14 interaction. 129 small molecules were screened iteratively, with identification of molecules producing up to 37% inhibition of TWEAK-Fn14 binding. In summary, we present a data-driven in silico study revealing key structural elements of the TWEAK-Fn14 interaction, followed by experimental validation, serving as a guide for the design of small molecule inhibitors of the TWEAK-Fn14 ligand-receptor interaction. Our results validate the TWEAK-Fn14 interaction as a chemically tractable target and provide the foundation for further exploration utilizing chemical biology approaches focusing on validating this system as a therapeutic target in invasive cancers.     

Chromatin-Based Classification of Genetically Heterogeneous AMLs into Two Distinct Subtypes with Diverse Stemness Phenotypes

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Computational design and evaluation of β-sheet breaker peptides for destabilizing Alzheimer's amyloid-β42 protofibrils

The β-sheet breaker (BSB) peptides interfere with amyloid fibril assembly and used as therapeutic agents in the treatment of Alzheimer's disease (AD). In this regard, a simple yet effective in silico screening methodology was applied in the present study to evaluate a potential 867 pentapeptide library based on known BSB peptide, LPFFD, for destabilizing Aβ₄₂ protofibrils. The molecular docking based virtual screening was used to filter out pentapeptides having binding affinities stronger than LPFFD. In the next step, binding free energies of the top 10 pentapeptides were evaluated using the MM-PBSA method. The residue-wise binding free energy analysis reveals that two pentapeptides, PVFFE, and PPFYE, bind to the surface of Aβ₄₂ protofibril and another pentapeptide, PPFFE, bind in the core region of Aβ₄₂ protofibril. By employing molecular dynamics simulation as a post filter for the top-hit peptides from MM-PBSA, the pentapeptides, PPFFE, PVFFE, and PPFYE, have been identified as potential BSB peptides for destabilizing Aβ₄₂ protofibril structure. The conformational microstate analysis, a significant decrease in the β-sheet content of Aβ₄₂ protofibril, a loss in the total number of hydrogen bonds in Aβ₄₂ protofibril, Asp23-Lys28 salt bridge destabilization and analysis of the free energy surfaces highlight Aβ₄₂ protofibril structure destabilization in presence of pentapeptides. Among three top-hit pentapeptides, PPFFE displayed the most potent Aβ₄₂ protofibril destabilization effect that shifted the energy minima toward lowest value of β-sheet content as well as lowest number of hydrogen bonds in Aβ₄₂ protofibril. The in silico screening workflow presented in the study highlight an alternative tool for designing novel peptides with enhanced BSB ability as potential therapeutic agents for AD.     

Recent admixture in an Indian population of African ancestry

Identification and study of genetic variation in recently admixed populations not only provides insight into historical population events but also is a powerful approach for mapping disease loci. We studied a population (OG-W-IP) that is of African-Indian origin and has resided in the western part of India for 500 years; members of this population are believed to be descendants of the Bantu-speaking population of Africa. We have carried out this study by using a set of 18,534 autosomal markers common between Indian, CEPH-HGDP, and HapMap populations. Principal-components analysis clearly revealed that the African-Indian population derives its ancestry from Bantu-speaking west-African as well as Indo-European-speaking north and northwest Indian population(s). STRUCTURE and ADMIXTURE analyses show that, overall, the OG-W-IPs derive 58.7% of their genomic ancestry from their African past and have very little inter-individual ancestry variation (8.4%). The extent of linkage disequilibrium also reveals that the admixture event has been recent. Functional annotation of genes encompassing the ancestry-informative markers that are closer in allele frequency to the Indian ancestral population revealed significant enrichment of biological processes, such as ion-channel activity, and cadherins. We briefly examine the implications of determining the genetic diversity of this population, which could provide opportunities for studies involving admixture mapping.     

Activation of DNA damage response signaling in lung adenocarcinoma A549 cells following oxygen beam irradiation

Oxygen beams are high linear energy transfer (LET) radiation characterized by higher relative biological effectiveness than low LET radiation. The aim of the current study was to determine the signaling differences between γ- and oxygen ion-irradiation. Activation of various signaling molecules was looked in A549 lung adenocarcinoma cells irradiated with 2Gy oxygen, 2Gy or 6Gy γ-radiation. Oxygen beam was found to be three times more cytotoxic than γ-radiation. By 4h there was efficient repair of DNA in A549 cells exposed to 2Gy or 6Gy gamma radiation but not in cells exposed to 2Gy oxygen beam as determined by γ-H2AX counting. Number of ATM foci was found to be significantly higher in cells exposed to 2Gy oxygen beam. Percentage of cells showing ATR foci were more with gamma however number of foci per cell were more in case of oxygen beam. Oxygen beam irradiated cells showed phosphorylation of Chk1, Chk2 and p53. Many apoptotic nuclei were seen by DAPI staining in cells exposed to oxygen beam. The noteworthy finding of this study is the activation of the sensor proteins, ATM and ATR by oxygen irradiation and the significant activation of Chk1, Chk2 and p53 only in the oxygen beam irradiated cells.