Differential fates of introns in gene expression due to global alternative splicing.

The discovery of introns over four decades ago revealed a new vision of genes and their interrupted arrangement. Throughout the years, it has appeared that introns play essential roles in the regulation of gene expression. Unique processing of excised introns through the formation of lariats suggests a widespread role for these molecules in the structure and function of cells. In addition to rapid destruction, these lariats may linger on in the nucleus or may even be exported to the cytoplasm, where they remain stable circular RNAs (circRNAs). Alternative splicing (AS) is a source of diversity in mature transcripts harboring retained introns (RI-mRNAs). Such RNAs may contain one or more entire retained intron(s) (RIs), but they may also have intron fragments resulting from sequential excision of smaller subfragments via recursive splicing (RS), which is characteristic of long introns. There are many potential fates of RI-mRNAs, including their downregulation via nuclear and cytoplasmic surveillance systems and the generation of new protein isoforms with potentially different functions. Various reports have linked the presence of such unprocessed transcripts in mammals to important roles in normal development and in disease-related conditions. In certain human neurological-neuromuscular disorders, including myotonic dystrophy type 2 (DM2), frontotemporal dementia/amyotrophic lateral sclerosis (FTD/ALS) and Duchenne muscular dystrophy (DMD), peculiar processing of long introns has been identified and is associated with their pathogenic effects. In this review, we discuss different mechanisms involved in the processing of introns during AS and the functions of these large sections of the genome in our biology.

Disrupting the Molecular Pathway in Myotonic Dystrophy.

Myotonic dystrophy is the most common muscular dystrophy in adults. It consists of two forms: type 1 (DM1) and type 2 (DM2). DM1 is associated with a trinucleotide repeat expansion mutation, which is transcribed but not translated into protein. The mutant RNA remains in the nucleus, which leads to a series of downstream abnormalities. DM1 is widely considered to be an RNA-based disorder. Thus, we consider three areas of the RNA pathway that may offer targeting opportunities to disrupt the production, stability, and degradation of the mutant RNA.

Induction of Senescence in Cancer Cells by a Novel Combination of Cucurbitacin B and Withanone: Molecular Mechanism and Therapeutic Potential.

Cancer, an uncontrolled proliferation syndrome, is treated with synthetic chemotherapeutic drugs that are associated with severe adverse effects. Development and application of new natural compounds is warranted to deal with the exponentially increasing incidence of cancer worldwide. Keeping selective toxicity to cancer cells as a priority criterion, we developed a combination of Cucurbitacin B and Withanone, and analyzed its anticancer potential using non-small cell lung cancer cells. We demonstrate that the selective cytotoxicity of the combination, called CucWi-N, to cancer cells is mediated by induction of cellular senescence that was characterized by decrease in Lamin A/C, CDK2, CDK4, Cyclin D, Cyclin E, phosphorylated RB, mortalin and increase in p53 and CARF proteins. It compromised cancer cell migration that was mediated by decrease in mortalin, hnRNP-K, vascular endothelial growth factor, matrix metalloproteinase 2, and fibronectin. We provide in silico, molecular dynamics and experimental data to support that CucWi-N (i) possesses high capability to target mortalin-p53 interaction and hnRNP-K proteins, (ii) triggers replicative senescence and inhibits metastatic potential of the cancer cells, and (iii) inhibits tumor progression and metastasis in vivo. We propose that CucWi-N is a potential natural anticancer drug that warrants further mechanistic and clinical studies.

Caffeic acid phenethyl ester (CAPE) possesses pro-hypoxia and anti-stress activities: bioinformatics and experimental evidences.

Honeybee propolis and its bioactive component, caffeic acid phenethyl ester (CAPE), are known for a variety of therapeutic potentials. By recruiting a cell-based reporter assay for screening of hypoxia-modulating natural drugs, we identified CAPE as a pro-hypoxia factor. In silico studies were used to probe the capacity of CAPE to interact with potential hypoxia-responsive proteins. CAPE could not dock into hypoxia inducing factor (HIF-1), the master regulator of hypoxia response pathway. On the other hand, it was predicted to bind to factor inhibiting HIF (FIH-1). The active site residue (Asp201) of FIH-1α was involved in hydrogen bond formation with CAPE and its analogue, caffeic acid methyl ester (CAME), especially in the presence of Fe and 2-oxoglutaric acid (OGA). We provide experimental evidence that the low doses of CAPE, that did not cause cytotoxicity or anti-migratory effect, activated HIF-1α and inhibited stress-induced protein aggregation, a common cause of age-related pathologies. Furthermore, by structural homology search, we explored and found candidate compounds that possess stronger FIH-1 binding capacity. These compounds could be promising candidates for modulating therapeutic potential of CAPE, and its recruitment in treatment of protein aggregation-based disorders.

Fragment-Based Ligand Designing.

Fragment-based drug design strategies have been used in drug discovery since it was first demonstrated using experimental structural biology techniques such as nuclear magnetic resonance (NMR) and X-ray crystallography. The underlying idea is that existing or new chemical entities with known desirable properties may serve both as tool compounds and as starting points for hit-to-lead expansion. Despite the recent advancements, there remain challenges to overcome, such as assembly of the synthetically feasible structures, development of scoring functions to correlate structure and their activities, and fine tuning of the promising molecules. This chapter first covers the theoretical background needed to understand the concepts and the challenges related to the field of study, followed by the description of important protocols and related software. Case studies are presented to demonstrate practical applications.

In silico identification and construction of microbial gene clusters associated with biodegradation of xenobiotic compounds.

Chemical substances not showing any importance in existence of biological systems and causing serious health hazards may be designated as Xenobiotic compound. Elimination or degradation of these unwanted substances is a major issue of concern for current time research. Process of biodegradation is a very important aspect of current research as discussed in current manuscript. Current study focuses on the detailed mining of data for the construction of microbial consortia for wide range of xenobiotics compounds. Intensive literature search was done for the construction of this library. Desired data was retrieved from NCBI in fasta format. Data was analysed through homology approaches by using BLAST. This homology based searched enriched with a great vision that not only bacterial population but many other cheap and potential sources are available for different xenobiotic degradation. Though it was focused that bacterial population covers a major part of biodegradation which is near about 90.6% but algae and fungi are also showing promising future in degradation of some important xenobiotic compounds. Analysis of data reveals that Pseudomonas putida has potential for degrading maximum compounds. Establishment of correlation through cluster analysis signifies that Pseudomonas putida, Aspergillus niger and Skeletonema costatum can have combined traits that can be used in finding out actual evolutionary relationship between these species. These findings may also givea new outcome in terms of much cheaper and eco-friendly source in the area of biodegradation of specified xenobiotic compounds.

Function and structure-based screening of compounds, peptides and proteins to identify drug candidates.

Drug discovery in simple words is all about finding small molecular compounds that possess the potential to interact with specific bio-macromolecules, mainly proteins, thereby bringing a desired effect in the functioning of the target molecules. Virtual screening of large compound libraries using computational approaches has come up as a great alternative to cost and labor-intensive high-throughput screening carried out in laboratories. Virtual high-throughput screening enormously reduces the number of compounds for systematic analysis using biochemical assays before entering the clinical trials. Here, we first give a brief overview of the rationale behind virtual screening, types of virtual screening - structure-based, ligand-based and inverse virtual screening, and challenges that need to be addressed to improve the existing strategies. Subsequently, we describe the methodology adopted for virtual screening of small molecules, peptides and proteins. Finally, we use few case studies to provide a better insight to the application of computer-aided high-throughput screening.