Annonaceous acetogenins as promising DNA methylation inhibitors to prevent and treat leukemogenesis - an in silico approach.

Leukemia is a haematological malignancy affecting blood and bone marrow, ranking 10th among the other common cancers. DNA methylation is an epigenetic dysregulation that plays a critical role in leukemogenesis. DNA methyltransferases (DNMTs) such as DNMT1, DNMT3A and DNMT3B are the key enzymes catalysing DNA methylation. Inhibition of DNMT1 with secondary metabolites from medicinal plants helps reverse DNA methylation. The present study focuses on inhibiting DNMT1 protein (PDB ID: 3PTA) with annonaceous acetogenins through in-silico studies. The docking and molecular dynamic (MD) simulation study was carried out using Schrödinger Maestro and Desmond, respectively. These compounds' drug likeliness, ADMET properties and bioactivity scores were analysed. About 76 different acetogenins were chosen for this study, among which 17 showed the highest binding energy in the range of -8.312 to -10.266 kcal/mol. The compounds with the highest negative binding energy were found to be annohexocin (-10.266 kcal/mol), isoannonacinone (-10.209 kcal/mol) and annonacin (-9.839 kcal/mol). MD simulation results reveal that annonacin remains stable throughout the simulation time of 100 ns and also binds to the catalytic domain of DNMT1 protein. From the above results, it can be concluded that annonacin has the potential to inhibit the DNA methylation process and prevent leukemogenesis.Communicated by Ramaswamy H. Sarma.

Nuclear receptor: Structure and function.

Ligand-dependent transcription factors are nuclear receptors (NRs) that regulate various critical cellular processes such as reproduction, metabolism, development, etc. NRs are classified into (subgroup 0 to subgroup 6) seven superfamilies based on ligand-binding characteristics. All NRs share a general domain structure (A/B, C, D, and E) with distinct essential functions. NRs as monomers, homodimers, or heterodimers bind to consensus DNA sequences known as Hormone Response Elements (HREs). Furthermore, nuclear receptor-binding efficiency depends on minor differences in the sequences of HREs, spacing between the two half-sites, and the flanking sequence of the response elements. NRs can trans-activate and repress their target genes. In positively regulated genes, ligand-bound NRs recruit coactivators to activate the target gene expression, and unliganded NRs cause transcriptional repression. On the other hand, NRs repress gene expression by different mechanisms: (i) ligand-dependent transcriptional repression, (ii) ligand-independent transcriptional repression. This chapter will briefly explain NR superfamilies, their structures, molecular mechanism of action and their role in pathophysiological conditions, etc. That could enable the discovery of new receptors and their ligands and may elucidate their roles in various physiological processes. In addition, therapeutic agonists and antagonists would be developed to control the dysregulation of nuclear receptor signaling.

CRISPR/Cas9 in epigenetics studies of health and disease.

Epigenetics is the heritable phenotypic changes without altering the genotype. Epigenetic processes are such as histone methylation, acetylation, ubiquitination, sumoylation, phosphorylation, ADP ribosylation, DNA methylation and non-coding RNAs interactions associated with structural changes in chromatin. The change of structure is either open chromatin for "active" state or closed chromatin for "inactive" state, that regulates important biological phenomenon like chromatin condensation, gene expression, DNA repair, cellular development, differentiation and homeostasis, etc. However, dysregulation of epigenetic patterns causes diseases like cancer, diabetes, neurological disorder, infectious diseases, autoimmunity etc. Besides, the most important clinical uses of Epigenetics studies are i. identification of disease biomarkers and ii. development of their therapeutics. Epigenetic therapies include epi-drugs, combinatorial therapy, nanocarriers, plant-derived products that are being used for changing the epigenetic pattern to reverse gene expression. However, the developed epi- drugs cause off-target gene and transposable elements activation; promote mutagenesis and carcinogenesis in normal cells, are the major hurdles regarding their clinical use. Therefore, advanced epigenetic therapeutics are required to develop target-specific epigenetic modifications to reverse gene expression pattern. CRISPR-Cas9 (Clustered Regularly Interspaced Palindrome Repeats-associated protein 9) system-mediated gene activation mechanism paves new methods of target-specific epigenetic therapeutics to cure diseases. In this chapter, we discuss how CRISPR/Cas9 and dCas9 have recently been engineered for epigenome editing. Different strategies have been discussed used for epigenome editing based on their efficacy and complexity. Last but not least we have discussed the limitations, different uses of CRISPR/Cas9 and dCas9 in the area of genetic engineering.

MHC Class II (DRB) Promoter Polymorphism and Its Role in Parasite Control among Malaria Patients.

MHC class II (MHCII) molecules are cell surface glycoproteins that play an important role to develop adaptive immune responses. MHCII-disease association is not restricted to structural variation alone but also may extend to genetic variations, which may modulate gene expression. The observed variations in class II gene expression make it possible that the association of MHCII polymorphism with diseases may relate to the level of gene expression in addition to the restriction of response to Ag. Understanding the extent of, and the mechanisms underlying, transcription factor DNA binding variation is therefore key to elucidate the molecular determinants of complex phenotypes. In this study, we investigated whether single nucleotide polymorphisms in MHCII-DRB regulatory gene may be associated with clinical outcomes of malaria in Plasmodium-infected individuals. To this end, we conducted a case-control study to compare patients who had mild malaria with those patients who had asymptomatic Plasmodium infection. It demonstrates that GTAT haplotype exerts an increased DRB transcriptional activity, resulting in higher DRB expression and subsequently perturbed Ag presentation and T cell activation, higher TLR-mediated innate immune gene expression, and Ag clearance, so low parasitemia in comparison with haplotypes other than GTAT (GTAC, GGGT). Hence, we hypothesized that DRB gene promoter polymorphism might lead to altered DRB gene expression, which could possibly affect the TLR-triggered innate immune responses in malaria patients. These genetic findings may contribute to the understanding of the pathogenesis of malaria and will facilitate the rational vaccine design for malaria.

Correction: 3, 3'5 Triiodo L Thyronine Induces Apoptosis in Human Breast Cancer MCF-7cells, Repressing SMP30 Expression through Negative Thyroid Response Elements.

[This corrects the article DOI: 10.1371/journal.pone.0020861.].

3, 3'5 Triiodo L thyronine induces apoptosis in human breast cancer MCF-7 cells, repressing SMP30 expression through negative thyroid response elements.

BACKGROUND: Thyroid hormones regulate cell proliferation, differentiation as well as apoptosis. However molecular mechanism underlying apoptosis as a result of thyroid hormone signaling is poorly understood. The antiapoptotic role of Senescence Marker Protein-30 (SMP30) has been characterized in response to varieties of stimuli as well as in knock out model. Our earlier data suggest that thyroid hormone 3, 3'5 Triiodo L Thyronine (T(3)), represses SMP30 in rat liver. METHODOLOGY/PRINCIPAL FINDINGS: In highly metastatic MCF-7, human breast cancer cell line T3 treatment repressed SMP30 expression leading to enhanced apoptosis. Analysis by flow cytometry and other techniques revealed that overexpression and silencing of SMP30 in MCF-7 resulted in decelerated and accelerated apoptosis respectively. In order to identify the cis-acting elements involved in this regulation, we have analyzed hormone responsiveness of transiently transfected hSMP30 promoter deletion reporter vectors in MCF-7 cells. As opposed to the expected epigenetic outcome, thyroid hormone down regulated hSMP30 promoter activity despite enhanced recruitment of acetylated H3 on thyroid response elements (TREs). From the stand point of established epigenetic concept we have categorised these two TREs as negative response elements. Our attempt of siRNA mediated silencing of TRß, reduced the fold of repression of SMP30 gene expression. In presence of thyroid hormone, Trichostatin- A (TSA), which is a Histone deacetylase (HDAC) inhibitor further inhibited SMP30 promoter activity. The above findings are in support of categorisation of both the thyroid response element as negative response elements as usually TSA should have reversed the repressions. CONCLUSION: This is the first report of novel mechanistic insights into the remarkable downregulation of SMP30 gene expression by thyroid hormone which in turn induces apoptosis in MCF-7 human breast cancer cells. We believe that our study represents a good ground for future effort to develop new therapeutic approaches to challenge the progression of breast cancer.

Alterations in expression of senescence marker protein-30 gene by 3,3',5-triiodo-L-thyronine (T3).

Thyroid hormone (T3) is essential for normal development, differentiation, and metabolic balance of the body. A toxic dose of T(3) in animals increases the basal metabolic rate and reactive oxygen species production, resulting more oxidative stress through Ca(2+) influx to cytoplasm. Senescence Marker Protein-30 (SMP30) is preferentially expressed in the liver and protects cells against various injuries by enhancement of Ca(2+) efflux to either extra cellular space or intraorganellar spaces through membrane Ca(2+) pump activity. In this paper we report an alteration in the level of SMP30 gene expression using RT-PCR and western blot analysis in T(3) treated female Wistar rats. The results indicate that there is an induction of SMP30 expression during early hours of T(3 )treatment and it declines in severe hyperthyroidism. Therefore, we speculate that SMP30 is regulated by T(3) and might play a protective role in hyperthyroidism.