High-risk genes involved in common septal defects of congenital heart disease.

The septation defect is one of the main categories of congenital heart disease (CHD). They can affect the septation of the atria leading to atrial septal defect (ASD), septation of ventricles leading to ventricular septal defect (VSD), and formation of the central part of the heart leading to atrioventricular septal defect (AVSD). Disruption of critical genetic factors involved in the proper development of the heart structure leads to CHD manifestation. Because of this, to identify the high-risk genes involved in common septal defects, a comprehensive search of the literature with the help of databases and the WebGestalt analysis tool was performed. The high-risk genes identified in the analysis were checked in 16 Indian whole-exome sequenced samples, including 13 VSD and three Tetralogy of Fallot for in silico validation. This data revealed three variations in GATA4, i.e., c.C1223A at exon 6: c.C602A and c.C1220A at exon 7; and one variation in MYH6, i.e., c.G3883C at exon 28 in two VSD cases. This study supports previously published studies that suggested GATA4 and MYH6 as the high-risk genes responsible for septal defects. Thus, this study contributes to a better understanding of the genes involved in heart development by identifying the high-risk genes and interacting proteins in the pathway.

Effect of Structural Changes in Proteins Derived from GATA4 Nonsynonymous Single Nucleotide Polymorphisms in Congenital Heart Disease.

Congenital heart disease is the most common type of birth defect. The single nucleotide polymorphism in GATA4 is associated with various congenital heart disease phenotypes. In the present study, we analysed the nonsynonymous single nucleotide polymorphism of GATA4, which are involved in congenital heart disease by predicting the changes in protein structures. Total of 49 nonsynonymous single nucleotide polymorphisms of GATA4 was screened from congenital heart disease patients of Mysore and also globally reported nonsynonymous single nucleotide polymorphisms. To understand the role of nonsynonymous single nucleotide polymorphisms, we mutated the sequence and translated into amino acids. Further the mutated protein secondary structure is predicted and tertiary structure is predicted using homology modeling. The quantitative evaluation of protein structure quality was verified with Volume Area Dihedral Angle Reporter server. Results revealed the secondary, tertiary structural changes along with changes in free energy of folding, volume and accessible surface area. Thus, the structural changes in the mutated proteins impaired the normal function of GATA4.

Divergence of the gene aly in experimentally evolved cytoraces, the members of the nasuta-albomicans complex of Drosophila.

We generated cytoraces by crossing the chromosomal races (Drosophila nasuta nasuta and Drosophila nasuta albomicans) of the nasuta subgroup of Drosophila and maintained the offspring over many generations through sibling mating. These cytoraces, along with their parents, are members of the nasuta-albomicans complex of Drosophila. The gene always early (aly) is one of the rapidly evolving genes in the genus Drosophila and plays a central role in regulating meiosis. Here we examined the rate of molecular evolution of aly in cytoraces of Drosophila and demonstrated that the rate of substitutions amongst cytoraces is around eight times greater than their parents and even amongst species of subgenera. Thus, the presence of positive selection in the laboratory-derived cytoraces based on the analysis of the synonymous and nonsynonymous substitution rates of aly suggests the rapid evolution in cytoraces.

INN-toxin, a highly lethal peptide from the venom of Indian cobra (Naja naja) venom-Isolation, characterization and pharmacological actions.

A novel toxic polypeptide, INN-toxin, is purified from the venom of Naja naja using combination of gel-permeation and ion-exchange chromatography. It has a molecular mass of 6951.6Da as determined by MALDI-TOF/MS and the N-terminal sequence of LKXNKLVPLF. It showed both neurotoxic as well as cytotoxic activities. INN-toxin is lethal to mice with a LD(50) of 1.2mg/kg body weight. IgY raised in chicks against basic peptide pool neutralized the toxicity of INN-toxin. INN-toxin did not inhibit cholinesterase activity. It is toxic to Ehrlich ascites tumor (EAT) cells, but it is not toxic to leukocyte culture. The toxin appears to be specific in its mode of action. Interaction of N-bromosuccinamide (NBS) with the peptide resulted in the modification of tryptophan residues and loss of lethal toxicity of INN-toxin.

Hybridization and introgression of the genomes of Drosophila nasuta and Drosophila albomicans: evolution of new karyotypes.

Drosophila nasuta (2n = 8) and Drosophila albomicans (2n = 6) are cross-fertile allopatric sibling chromosomal races of the nasuta subgroup of Drosophila. Hybrids of these races can be maintained for any number of generations. Some of the introgressed hybrid lineages of D. nasuta and D. albomicans, after passing through a transient phase of karyotypic polymorphism, ended up with a stable karyotype whose composition is different from those of the parental races. Such hybrid populations were called cytoraces, in which the chromosomes of D. nasuta and D. albomicans are represented in different combinations. The karyotypic composition of 16 such cytoraces have been presented and discussed with reference to evolutionary strategies such as balancing selection, directional selection, and sex-specific effect on different components of the evolving karyotypes.

Incipient sexual isolation in the nasuta-albomicans complex of Drosophila: mating preference in male-, female- and multiple-choice mating experiments.

Interracial divergence is an important facet of speciation. The nasuta-albomicans complex of Drosophila with sixteen morphologically identical, karyotypically different but cross-fertile races is an excellent system to study a few dimensions of raciation. Drosophila nasuta nasuta, Drosophila nasuta albomicans, Cytorace 1, Cytorace 2, Cytorace 3 and Cytorace 4 of this subgroup have been subjected to male-, female- and multiple-choice mating experiments. Out of 8456 crosses conducted, 7185 had successful matings. The overall impression is that mating is far from random amongst these six closely related races of the nasuta-albomicans complex. The males of D. n. albomicans, Cytorace 1 and Cytorace 4 in male-choice, the females of Cytorace 1 and Cytorace 2 in female-choice, and the males and females of D. n. nasuta, D. n. albomicans, Cytorace 1 and Cytorace 4 against the males and females of Cytorace 2 in multiple-choice experiments, had significantly more homogamic matings than expected. Thus in this study of evolutionary experimentation on raciation under laboratory conditions, we have documented the initiation of preference for conspecific matings among closely related and independently evolving members of the nasuta-albomicans complex of Drosophila.

Autoradiographic study of transcription and dosage compensation in the sex and neo-sex chromosome of Drosophila nasuta nasuta and Drosophila nasuta albomicans.

Cellular autoradiography is used to study the transcription patterns of the polytene X chromosomes in Drosophila nasuta nasuta and D. n. albomicans. D. n. nasuta, with 2n = 8, includes a pair of complete heteromorphic sex chromosomes, whereas D. n. albomicans, with 2n = 6, has a pair of metacentric neo-sex chromosomes representing incomplete heteromorphic sex chromosomes. The neo-X chromosome has two euchromatic arms, one representing the ancestral X while the other represents the ancestral autosome 3 chromosomes. The metacentric neo-Y chromosome has one arm with a complete heterochromatic ancestral Y and the other arm with a euchromatic ancestral autosome 3. The transcription study has revealed that the X chromosome in D. n. nasuta is hyperactive, suggesting complete dosage compensation, while in the neo-X chromosome of D. n. albomicans the ancestral X chromosome is hyperactive and the ancestral autosome 3, which is part of the neo-sex chromosome, is similar to any other autosomes. This finding shows dosage compensation on one arm (XLx/-) of the neo-X chromosome, while the other arm (XR3/YR3) is not dosage compensated and has yet to acquire the dosage compensatory mechanism.

Incipient sexual isolation in the nasuta-albomicans complex of Drosophila: no-choice experiments.

Drosophila nasuta nasuta and Drosophila nasuta albomicans are cross-fertile races of Drosophila. Hybridization between these races in the laboratory has given rise to new races (Cytoraces), among which karyotypic composition differs from one another and also from those of the parental races. In this study, we search for the evidence of incipient reproductive isolation among the parental races and four Cytoraces by assessing the fraction of no-matings, mating latency and copulation duration in all possible types of homo- and heterogamic crosses (N = 4184). In no-choice conditions, the latency time (time to initiation of copulation) is lower in homogamic crosses than in heterogamic crosses for both parental races and Cytoraces. Latency time and copulation duration are negatively correlated, whereas fraction of no matings is positively correlated with latency time. Thus these six closely related races of the nasuta-albomicans complex show the initiation of the earliest stages of pre-zygotic isolation, manifested as a tendency for matings to be initiated earlier and more often, and for a longer duration, among homogamic rather than heterogamic individuals

Patterns of replication in the neo-sex chromosomes of Drosophila nasuta albomicans.

Drosophila nasuta albomicans (with 2n = 6), contains a pair of metacentric neo-sex chromosomes. Phylogenetically these are products of centric fusion between ancestral sex (X, Y) chromosomes and an autosome (chromosome 3). The polytene chromosome complement of males with a neo-X- and neo-Y-chromosomes has revealed asynchrony in replication between the two arms of the neo-sex chromosomes. The arm which represents the ancestral X-chromosome is faster replicating than the arm which represents ancestral autosome. The latter arm of the neo-sex chromosome is synchronous with other autosomes of the complement. We conclude that one arm of the neo-X/Y is still mimicking the features of an autosome while the other arm has the features of a classical X/Y-chromosome. This X-autosome translocation differs from the other evolutionary X-autosome translocations known in certain species of Drosophila.

Racial divergence in abdominal bristles among parental races and newly evolved cytoraces of nasuta-albomicans complex of Drosophila.

Cytoraces are the products of interracial hybridization between Drosophila nasuta nasuta and D. nasuta albomicans. These races differ from their parents in the chromosome composition, mating preference, certain fitness phenotypes and also a few morphophenotypic traits. Now, these cytoraces are passing through 330 generations. Racial divergence in the 4th and 5th abdominal bristles among the parental races and the newly evolved cytorace 1 and 2 is reported. The results revealed that the parental races have more number of bristles than newly evolved cytoraces. Thus, these cytoraces are evolved/evolving with reduced abdominal bristle number and better fitness.

Does evolution reduce the body size? A study of the four members of newly evolved nasuta-albomicans complex of Drosophila.

Our long range interracial hybridization experiments between a pair of cross fertile races, Drosophila nasuta (2n = 8) and D. albomicans (2n = 6) have resulted in the evolution of two new karyotypic strains under laboratory conditions, which are named as Cytorace 1 and Cytorace 2. These Cytoraces harbor chromosomes from both parents. Here, we compare the body size of the parental races and newly evolved Cytoraces and the relationship between the body size and fitness. Analysis reveals that the parental races have reduced fertility and are larger in body size than newly evolved Cytoraces. Thus, the newly evolved Cytoraces show reduced body size and better fitness in the course of their evolution.

A direct screen identifies new flight muscle mutants on the Drosophila second chromosome.

An ethyl methanesulfonate mutagenesis of Drosophila melanogaster was undertaken, and >3000 mutagenized second chromosomes were generated. More than 800 homozygous viable lines were established, and adults were screened directly under polarized light for muscle defects. A total of 16 mutant strains in which the indirect flight muscles were reduced in volume or disorganized or were otherwise abnormal were identified. These fell into seven recessive and one semidominant complementation groups. Five of these eight complementation groups, including the semidominant mutation, have been mapped using chromosomal deficiencies and meiotic recombination. Two complementation groups mapped close to the Myosin heavy chain gene, but they are shown to be in different loci. Developmental analysis of three mutations showed that two of these are involved in the early stages of adult myogenesis while the other showed late defects. This is the first report of results from a systematic and direct screen for recessive flight muscle defects. This mutant screen identifies genes affecting the flight muscles, which are distinct from those identified when screening for flightlessness.

Pattern of sexual isolation between parental races (Drosophila nasuta nasuta and D.n. albomicans) and the newly evolved races (Cytorace I and II).

D.n. nasuta, D.n. albomicans, Cytorace I and Cytorace II [Chromosoma, 93 (1986) 243] are closely related strains of the nasuta subgroup. Through male choice and female choice experiments, the pattern of sexual isolation if any, among them was analysed. Differential mating preference of males and females of these races suggests that they are passing through the process of anagenesis. Therefore, this, as an evolutionary experiment under laboratory conditions, offers a rare opportunity to witness different pattern and levels of divergence among four cytogentically parsimonious races of Drosophila.

Supernumerary chromosomes in Drosophila nasuta albomicana.

Supernumerary chromosomes have been detected in the karyotype of D.n.albomicana. Their number varies from one to three. They are the smallest elements in the karyotype. Karyotypes of D.n.albomicana with and without supernumerary chromosomes have been presented.