Dissecting the Ca2+ dependence of DesA1 function in Mycobacterium tuberculosis.

Mycobacterium tuberculosis (M. tb) has a complex cell wall, composed largely of mycolic acids, that are crucial to its structural maintenance. The M. tb desaturase A1 (DesA1) is an essential Ca2+-binding protein that catalyses a key step in mycolic acid biosynthesis. To investigate the structural and functional significance of Ca2+ binding, we introduced mutations at key residues in its Ca2+-binding ßγ-crystallin motif to generate DesA1F303A, E304Q, and F303A-E304Q. Complementation of a conditional ΔdesA1 strain of Mycobacterium smegmatis, with the Ca2+ non-binders F303A or F303A-E304Q, failed to rescue its growth phenotype; these complements also exhibited enhanced cell wall permeability. Our findings highlight the criticality of Ca2+ in DesA1 function, and its implicit role in the maintenance of mycobacterial cellular integrity.

Exome sequencing and functional analyses revealed CETN1 variants leads to impaired cell division and male fertility.

Human spermatogenesis requires an orchestrated expression of numerous genes in various germ cell subtypes. Therefore, the genetic landscape of male infertility is highly complex. Known genetic factors alone account for at least 15% of male infertility. However, ~40% of infertile men remain undiagnosed and are classified as idiopathic infertile men. We performed exome sequencing in 47 idiopathic infertile men (discovery cohort), followed by replication study (40 variants in 33 genes) in 844 infertile men and 709 controls using Sequenom MassARRAY® based genotyping. We report 17 variants in twelve genes that comprise both previously reported (DNAH8, DNAH17, FISP2 and SPEF2) and novel candidate genes (BRDT, CETN1, CATSPERD, GMCL1, SPATA6, TSSK4, TSKS and ZNF318) for male infertility. The latter have a strong biological nexus to human spermatogenesis and their respective mouse knockouts are concordant with human phenotypes. One candidate gene CETN1, identified in this study, was sequenced in another independent cohort of 840 infertile and 689 fertile men. Further, CETN1 variants were functionally characterized using biophysical and cell biology approaches. We demonstrate that CETN1 variant- p.Met72Thr leads to multipolar cells, fragmented nuclei during mitosis leading to cell death and show significantly perturbed ciliary disassembly dynamics. Whereas CETN1-5' UTR variant; rs367716858 leads to loss of a methylation site and increased reporter gene expression in vitro. We report a total of eight novel candidate genes identified by exome sequencing, which may have diagnostic relevance and can contribute to improved diagnostic workup and clinical management of male infertility.

Secretagogin is a Ca2+-dependent stress-responsive chaperone that may also play a role in aggregation-based proteinopathies.

Secretagogin (SCGN) is a three-domain hexa-EF-hand Ca2+-binding protein that plays a regulatory role in the release of several hormones. SCGN is expressed largely in pancreatic ß-cells, certain parts of the brain, and also in neuroendocrine tissues. The expression of SCGN is altered in several diseases, such as diabetes, cancers, and neurodegenerative disorders; however, the precise associations that closely link SCGN expression to such pathophysiologies are not known. In this work, we report that SCGN is an early responder to cellular stress, and SCGN expression is temporally upregulated by oxidative stress and heat shock. We show the overexpression of SCGN efficiently prevents cells from heat shock and oxidative damage. We further demonstrate that in the presence of Ca2+, SCGN efficiently prevents the aggregation of a broad range of model proteins in vitro. Small-angle X-ray scattering (BioSAXS) studies further reveal that Ca2+ induces the conversion of a closed compact apo-SCGN conformation into an open extended holo-SCGN conformation via multistate intermediates, consistent with the augmentation of chaperone activity of SCGN. Furthermore, isothermal titration calorimetry establishes that Ca2+ enables SCGN to bind α-synuclein and insulin, two target proteins of SCGN. Altogether, our data not only demonstrate that SCGN is a Ca2+-dependent generic molecular chaperone involved in protein homeostasis with broad substrate specificity but also elucidate the origin of its altered expression in several cancers. We describe a plausible mechanism of how perturbations in Ca2+ homeostasis and/or deregulated SCGN expression would hasten the process of protein misfolding, which is a feature of many aggregation-based proteinopathies.

Polymer modified magnetic-luminescent nanocomposites for combined optical imaging and magnetic fluid hyperthermia in cancer therapy: analysis of Mn2+ doping for enhanced heating effect, hemocompatibility and biocompatibility.

Magnetic MnxFe3-xO4 nanoparticles and polymer coated magnetic-luminescent MnxFe3-xO4@(Y,Dy/Eu)VO4 nanocomposites were prepared to study their comparative heat generation efficiency and biocompatibilities. Cubic crystalline phases were obtained for the nanoparticles and cubic-tetragonal biphasic phases were observed for the nanocomposites. The successful doping of Mn2+ was also confirmed by inductively coupled plasma optical emission spectroscopy. The compositions and the surface modification chemistry were confirmed by infrared spectroscopy. The formation of near-spherical and cubic/cuboid nanoparticles was observed from electron microscopy. Comparative analysis of induction heating efficiencies and magnetization values of the synthesized materials was performed for the samples. The samples showed an efficient heating effect under the influence of alternating magnetic field strengths - 3.05 × 106 kA m-1 s-1 and 4.58 × 106 kA m-1 s-1. A higher Mn2+ content was found to possess higher magnetization and perform better in heat generation. The nanocomposites give brilliant color emission on excitation using ultraviolet wavelengths - 300 and 315 nm. Their hydrodynamic radii and zeta potential values indicate good stability of the dispersions. Hemocompatibility studies were carried out to ascertain the effect on red blood cells. The materials were also found to exhibit excellent biocompatibility towards HeLa cell lines. This article will provide a new insight into the use of MnxFe3-xO4 based nanocomposites for magnetic fluid hyperthermia in cancer therapy.

Extracellular secretagogin is internalized into the cells through endocytosis.

Secretagogin (SCGN) is a calcium-sensor protein with a regulatory role in glucose metabolism and the secretion of several peptide hormones. Many, but not all, functions of SCGN can be explained by its intracellular manifestation. Despite early data on SCGN secretion, the secretory mechanism, biological fate, physiological implications and trans-cellular signalling of extracellular SCGN remain unknown. We here report that extracellular SCGN is readily internalized into the C2C12 cells in an energy-dependent manner. Using endocytosis inhibitors, we demonstrate that SCGN internalizes via clathrin-mediated endocytosis, following which, SCGN localizes largely in the cytosol. Exogenous SCGN treatment induces a global proteomic reprogramming in C2C12 cells. Gene ontology search suggests that SCGN-induced proteomic reprogramming favours protein synthesis and cellular growth. We thus validated the cell proliferative action of SCGN using C2C12, HepG2 and NIH-3T3 cell lines. Based on the data, we propose that circulatory SCGN is internalized into the target cells and modulates the expression of cell growth-related proteins. The work suggests that extracellular SCGN is a functional protein, which communicates with specific cell types and directly modulates cell proliferation.

Conformational scanning of individual EF-hand motifs of calcium sensor protein centrin-1.

Centrin-1, a Ca2+ sensor protein of the centrin family is a crucial player for cell division in eukaryotes and plays a key role in the microtubule organising centre. Despite being regarded as a calcium sensor with a matched structure to calmodulin/troponin C, the protein undergoes mild changes in conformation and binds Ca2+ with moderate affinity. We present an in-depth analysis of the Ca2+ sensing by individual EF-hand motifs of centrin-1 and address unsolved questions of the rationales for moderate affinity and conformational transitions of the protein. Employing the more sensitive approach of Trp scanning of individual EF-hand motif, we have undertaken an exhaustive investigation of Ca2+ binding to individual EF-hand motifs, named EF1 to EF4. All four EF-hand motifs of centrin-1 are structural as all of them bind both Ca2+ and Mg2+. EF1 and EF4 are the most flexible sites as they undergo drastic conformational changes following Ca2+ binding, whereas EF3 responds to Ca2+ minimally. On the other hand, EF2 moves towards the protein surface upon binding Ca2+. The independent filling mode of Ca2+ to EF-hand motifs and lack of intermotif communication explain the lack of cooperativity of binding, thus constraining centrin-1 to a moderate affinity binding protein. Thus, centrin-1 is distinct from other calcium sensors such as calmodulin.

Identification of VEGFA-centric temporal hypoxia-responsive dynamic cardiopulmonary network biomarkers.

AIMS: Hypoxia, a pathophysiological condition, is profound in several cardiopulmonary diseases (CPD). Every individual's lethality to a hypoxia state differs in terms of hypoxia exposure time, dosage units and dependent on the individual's genetic makeup. Most of the proposed markers for CPD were generally aim to distinguish disease samples from normal samples. Although, as per the 2018 GOLD guidelines, clinically useful biomarkers for several cardio pulmonary disease patients in stable condition have yet to be identified. We attempt to address these key issues through the identification of Dynamic Network Biomarkers (DNB) to detect hypoxia induced early warning signals of CPD before the catastrophic deterioration. MATERIALS AND METHODS: The human microvascular endothelial tissues microarray datasets (GSE11341) of lung and cardiac expose to hypoxia (1% O2) for 3, 24 and 48 h were retrieved from the public repository. The time dependent differentially expressed genes were subjected to tissue specificity and promoter analysis to filtrate the noise levels in the networks and to dissect the tissue specific hypoxia induced genes. These filtered out genes were used to construct the dynamic segmentation networks. The hypoxia induced dynamic differentially expressed genes were validated in the lung and heart tissues of male rats. These rats were exposed to hypobaric hypoxia (simulated altitude of 25,000 or PO2 - 282 mm of Hg) progressively for 3, 24 and 48 h. KEY FINDINGS: To identify the temporal key genes regulated in hypoxia, we ranked the dominant genes based on their consolidated topological features from tissue specific networks, time dependent networks and dynamic networks. Overall topological ranking described VEGFA as a single node dynamic hub and strongly communicated with tissue specific genes to carry forward their tissue specific information. We named this type of VEGFAcentric dynamic networks as "V-DNBs". As a proof of principle, our methodology helped us to identify the V-DNBs specific for lung and cardiac tissues namely V-DNBL and V-DNBC respectively. SIGNIFICANCE: Our experimental studies identified VEGFA, SLC2A3, ADM and ENO2 as the minimum and sufficient candidates of V-DNBL. The dynamic expression patterns could be readily exploited to capture the pre disease state of hypoxia induced pulmonary vascular remodelling. Whereas in V-DNBC the minimum and sufficient candidates are VEGFA, SCL2A3, ADM, NDRG1, ENO2 and BHLHE40. The time dependent single node expansion indicates V-DNBC could also be the pre disease state pathological hallmark for hypoxia-associated cardiovascular remodelling. The network cross-talk and expression pattern between V-DNBL and V-DNBC are completely distinct. On the other hand, the great clinical advantage of V-DNBs for pre disease predictions, a set of samples during the healthy condition should suffice. Future clinical studies might further shed light on the predictive power of V-DNBs as prognostic and diagnostic biomarkers for CPD.

HAHmiR.DB: a server platform for high-altitude human miRNA-gene coregulatory networks and associated regulatory circuits.

Around 140 million people live in high-altitude (HA) conditions! and even a larger number visit such places for tourism, adventure-seeking or sports training. Rapid ascent to HA can cause severe damage to the body organs and may lead to many fatal disorders. During induction to HA, human body undergoes various physiological, biochemical, hematological and molecular changes to adapt to the extreme environmental conditions. Several literature references hint that gene-expression-regulation and regulatory molecules like miRNAs and transcription factors (TFs) control adaptive responses during HA stress. These biomolecules are known to interact in a complex combinatorial manner to fine-tune the gene expression and help in controlling the molecular responses during this stress and ultimately help in acclimatization. High-Altitude Human miRNA Database (HAHmiR.DB) is a unique, comprehensive and curated collection of miRNAs that have been experimentally validated to be associated with HA stress, their level of expression in different altitudes, fold change, experiment duration, biomarker association, disease and drug association, tissue-specific expression level, Gene Ontology (GO) and Kyoto Encyclopaedia of Gene and Genomes (KEGG) pathway associations. As a server platform, it also uniquely constructs and analyses interactive miRNA-TF-gene coregulatory networks and extracts regulatory circuits/feed-forward loops (FFLs). These regulatory circuits help to offer mechanistic insights into complex regulatory mechanisms during HA stress. The server can also build these regulatory networks between two and more miRNAs of the database and also identify the regulatory circuits from this network. Hence, HAHmiR.DB is the first-of-its-kind database in HA research, which is a reliable platform to explore, compare, analyse and retrieve miRNAs associated with HA stress, their coregulatory networks and FFL regulatory-circuits. HAHmiR.DB is freely accessible at http://www.hahmirdb.in.

Google trend analysis of climatic zone based Indian severe seasonal sensitive population.

BACKGROUND: Our earlier Google Trend (GT) Analytics study reported that the worldwide human population severely subject to four seasonal (sensitive) comorbid lifestyle diseases (SCLD) such as asthma, obesity, hypertension and fibrosis. The human population subject to seasonal variability in these four diseases activity referred as "severe seasonal sensitive population". In India, the estimated burden of these four seasonal diseases is more than 350 million as on the year 2018. It is a growing crisis for India with a projected disease burden of 500 million in the year 2025. This study was aimed to decipher the genuine SCLD seasonal trends in the entire Indian population using GT and validate these trends in Indian climatic zones. METHODS: GT is used to study the temporal trends in web search using weekly Relative Search Volume (RSV) for the period 2004 to 2017. The relative search volume (RSV) of the four-severe seasonal comorbid diseases namely Asthma, Hypertension, Obesity and Fibrosis were collected with and without obesity as the reference. The RSV were collected using the GT selection options as (i) Whole India (ii) Jammu and Kashmir (Cold zone) (iii) Rajasthan (Hot and Dry zone) (iii) West Bengal (Hot and Humid zone) and (iv) Uttar Pradesh state (Composite zone). The time series analysis was carried out to find seasonal patterns, comorbidity, trends and periodicity in the entire India and four of its states (zones). RESULTS: Our analysis of entire India (2004-2017) revealed high significant seasonal patterns and comorbidity in all the four diseases of SCLD. The positive tau values indicated strong positive seasonal trends in the SCLD throughout the period (Table). The auto correlation analysis revealed that these diseases were subjected to 3, 4 and 6 months period seasonal variations. Similar seasonal patterns and trends were also observed in all the four Indian temperature zones. Overall study indicated that SCLD seasonal search patterns and trends are highly conserved in India even in drastic Indian climatic zones. CONCLUSIONS: The clinical outcome arise out of these observations could be of immense significance in handling the major chronic life style diseases asthma, hypertension, obesity and fibrosis. The possible strong comorbid relationship among asthma, hypertension, obesity and fibrosis may be useful to segregate Indian seasonal sensitive population. In disease activity-based chronotherapy, the search interest of segment of the population with access to Internet may be used as an indicator for public health sectors in the early detection of SCLD from a specific country or a region. As this disease population could be highly subject to the adverse effect of seasons in addition to life style and other environmental factors. Our study necessitates that these Indian populations need special attention from the Indian health care sectors.

Secretagogin Regulates Insulin Signaling by Direct Insulin Binding.

Secretagogin (SCGN) is a ß-cell enriched, secretory/cytosolic Ca2+-binding protein with unknown secretory regulation and functions. Recent findings suggest that SCGN deficiency correlates with compromised insulin response and diabetes. However, the (patho)physiological SCGN-insulin nexus remains unexplored. We here report that SCGN is an insulin-interacting protein. The protein-protein interaction between SCGN and insulin regulates insulin stability and increases insulin potency in vitro and in vivo. Mutagenesis studies suggest an indispensable role for N-terminal domain of SCGN in modulating insulin stability and function. SCGN supplementation in diabetogenic-diet-fed mice preserves physiological insulin responsiveness while relieving obesity and cardiovascular risk. SCGN-insulin interaction mediated alleviation of hyperinsulinemia by increased insulin internalization, which translates to reduced body fat and hepatic lipid accumulation, emerges as a plausible mechanism for the preservation of insulin responsiveness. These findings establish SCGN as a functional insulin-binding protein (InsBP) with therapeutic potential against diabetes.

Secretagogin Binding Prevents α-Synuclein Fibrillation.

Secretagogin (SCGN) is a secreted calcium sensor that has emerged as a potential multifunctional protein of neuroendocrine cells. A significantly reduced level of expression of SCGN has been reported in the hippocampus of a mouse model of Alzheimer's disease (AD) and in Parkinson's patients, although the biochemical implications and mechanistic underpinnings of the altered SCGN expression in neurodegenerative diseases remain unknown. We have pursued the interaction of SCGN with α-synuclein that we discovered in impartial pull-down analyses to decode the SCGN interactome. SCGN physically binds α-synuclein and rescues it from detrimental fibrillation. Correspondingly, it is observed that a significant reduction in the cytotoxicity of α-synuclein fibrils is caused by SCGN. We map these antifibrillar attributes to the central region and C-terminal domain of SCGN, while the N-terminal domain is not essential for this activity. On the basis of these results, a broader neuroprotective function of SCGN by proficient chaperone action is proposed. An intriguing correlation of this interaction with a reduced level of expression of SCGN in neurodegenerative diseases shall inspire further studies of the physiological role of SCGN in precluding pathological protein aggregation.

ßγ-Crystallination Endows a Novel Bacterial Glycoside Hydrolase 64 with Ca2+-Dependent Activity Modulation.

The prokaryotic ßγ-crystallins are a large group of uncharacterized domains with Ca2+-binding motifs. We have observed that a vast number of these domains are found appended to other domains, in particular, the carbohydrate-active enzyme (CAZy) domains. To elucidate the functional significance of these prospective Ca2+ sensors in bacteria and this widespread domain association, we have studied one typical example from Clostridium beijerinckii, a bacterium known for its ability to produce acetone, butanol, and ethanol through fermentation of several carbohydrates. This novel glycoside hydrolase of family 64 (GH64), which we named glucanallin, is composed of a ßγ-crystallin domain, a GH64 domain, and a carbohydrate-binding module 56 (CBM56). The substrates of GH64, ß-1,3-glucans, are the targets for industrial biofuel production due to their plenitude. We have examined the Ca2+-binding properties of this protein, assayed its enzymatic activity, and analyzed the structural features of the ß-1,3-glucanase domain through its high-resolution crystal structure. The reaction products resulting from the enzyme reaction of glucanallin reinforce the mixed nature of GH64 enzymes, in contrast to the prevailing notion of them being an exotype. Upon disabling Ca2+ binding and comparing different domain combinations, we demonstrate that the ßγ-crystallin domain in glucanallin acts as a Ca2+ sensor and enhances the glycolytic activity of glucanallin through Ca2+ binding. We also compare the structural peculiarities of this new member of the GH64 family to two previously studied members.IMPORTANCE We have biochemically and structurally characterized a novel glucanase from the less studied GH64 family in a bacterium significant for fermentation of carbohydrates into biofuels. This enzyme displays a peculiar property of being distally modulated by Ca2+ via assistance from a neighboring ßγ-crystallin domain, likely through changes in the domain interface. In addition, this enzyme is found to be optimized for functioning in an acidic environment, which is in line with the possibility of its involvement in biofuel production. Multiple occurrences of a similar domain architecture suggest that such a "ßγ-crystallination"-mediated Ca2+ sensitivity may be widespread among bacterial proteins.

Abundant Perithecial Protein (APP) from Neurospora is a primitive functional analog of ocular crystallins.

The eye arose during the Cambrian explosion from pre-existing proteins that would have been recruited for the formation of the specialized components of this organ, such as the transparent lens. Proteins suitable for the role of lens crystallins would need to possess unusual physical properties and the study of such earliest analogs of ocular crystallins would add to our understanding of the nature of recruitment of proteins as lens/corneal crystallins. We show that the Abundant Perithecial Protein (APP) of the fungi Neurospora and Sordaria fulfils the criteria for an early crystallin analog. The perithecia in these fungal species are phototropic, and APP accumulates at a high concentration in the neck of the pitcher-shaped perithecium. Spores are formed at the base of the perithecium, and light contributes to their maturation. The hydrodynamic properties of APP appear to exclude dimer formation or aggregation at high protein concentrations. APP is also deficient in Ca2+ binding, a property seen in its close homolog, the calcium-binding cell adhesion molecule (DdCAD-1) from Dictyostelium discoidum. Comparable to crystallins, APP occurs in high concentrations and seems to have dispensed with Ca2+ binding in exchange for greater stability. These crystallin-like attributes of APP lead us to demonstrate that it is a primitive form of ocular crystallins.

Veiled Potential of Secretagogin in Diabetes: Correlation or Coincidence?

Secretagogin (SCGN) is a calcium sensor protein enriched in neuroendocrine cells in general and pancreatic ß-cells in particular. SCGN regulates insulin secretion through several Ca2+-dependent interactions. Recent studies implicate SCGN in the ß-cell physiology and extracellular insulin function, making it an intriguing candidate in diabetes research. Here, we propose a conjoining theme of diversified SCGN function in diabetes pathology. In our opinion, SCGN is an attractive therapeutic candidate ascribed by its role in ß-cell maintenance and neuronal functions and in the efficacy of insulin. To scrutinize the therapeutic prospects of SCGN, we abridge putative diabetes-related properties of SCGN and put forth strategies to determine the precise role of SCGN in the pathogenesis/preclusion of diabetes.

Secretagogin Purification and Quality Control Strategies for Biophysical and Cell Biological Studies.

Secretagogin (SCGN) has recently gained attention due to its modulatory effect on insulin/CRH secretion and function. However, a large pool of speculated SCGN functions remains unexplored. A major deficiency is the lack of knowledge about the biological functions of extracellular SCGN. We here describe convenient methods for the scalable production of His-tagged and untagged mouse SCGN. The protocol is optimized to remove endotoxins, and thus the protein is suited for biological applications such as cell culture treatment or animal injections. We also outline expedient methods to check the purity of SCGN preparation for biological applications.

Tryptophan Scanning Mutagenesis of EF-Hand Motifs.

Ca2+ regulation in living systems occurs via specific structural alterations, subtle or drastic, in the Ca2+-binding domains of sensor proteins. Sensor proteins perform designated nonredundant roles within the dense network of Ca2+-binding proteins. A detailed understanding of the structural changes in calcium sensor proteins due to Ca2+ spikes that vary spatially, temporally, and in magnitude would provide better insights into the mechanism of Ca2+ sensing. This chapter describes a method to study various stages during apo to the holo transition of Ca2+-binding proteins by Trp-mediated scanning of individual EF-hand motifs. We describe the applicability of this procedure to caldendrin, which is a neuronal Ca2+-binding protein and to integrin-binding protein. Tryptophan mutants of full-length caldendrin were designed to reveal local structural changes in each EF-hand of the protein. This method, referred to as "EF-hand scanning tryptophan mutagenesis," not only allows the identification of canonical and noncanonical EF-hands using very low concentrations of protein but also enables visualization of the hierarchical filling of Ca2+ into the canonical EF-hands.

Analysis of Hypoxiamir-Gene Regulatory Network Identifies Critical MiRNAs Influencing Cell-Cycle Regulation Under Hypoxic Conditions.

BACKGROUND: Hypoxia is a pathophysiological condition which arises due to low oxygen concentration in conditions like cardiovascular diseases, inflammation, ascent to higher altitude, malignancies, deep sea diving, prenatal birth, etc. A number of microRNAs (miRNAs), Transcription Factors (TFs) and genes have been studied separately for their role in hypoxic adaptation and controlling cell-cycle progression and apoptosis during this stress. OBJECTIVE: We hypothesize that miRNAs and TFs may act in conjunction to regulate a multitude of genes and play a crucial and combinatorial role during hypoxia-stress-responses and associated cellcycle control mechanisms. METHOD: We collected a comprehensive and non-redundant list of human hypoxia-responsive miRNAs (also known as hypoxiamiRs). Their experimentally validated gene-targets were retrieved from various databases and a comprehensive hypoxiamiR-gene regulatory network was built. RESULTS: Functional characterization and pathway enrichment of genes identified phospho-proteins as enriched nodes. The phospho-proteins which were localized both in the nucleus and cytoplasm and could potentially play important role as signaling molecules were selected; and further pathway enrichment revealed that most of them were involved in NFkB signaling. Topological analysis identified several critical hypoxiamiRs and network perturbations confirmed their importance in the network. Feed Forward Loops (FFLs) were identified in the subnetwork of enriched genes, miRNAs and TFs. Statistically significant FFLs consisted of four miRNAs (hsa-miR-182-5p, hsa- miR-146b-5p, hsa-miR-96, hsa-miR-20a) and three TFs (SMAD4, FOXO1, HIF1A) both regulating two genes (NFkB1A and CDKN1A). CONCLUSION: Detailed BioCarta pathway analysis identified that these miRNAs and TFs together play a critical and combinatorial role in regulating cell-cycle under hypoxia, by controlling mechanisms that activate cell-cycle checkpoint protein, CDKN1A. These modules work synergistically to regulate cell-proliferation, cell-growth, cell-differentiation and apoptosis during hypoxia. A detailed mechanistic molecular model of how these co-regulatory FFLs may regulate the cell-cycle transitions during hypoxic stress conditions is also put forth. These biomolecules may play a crucial and deterministic role in deciding the fate of the cell under hypoxic-stress.

High altitude mediated skeletal muscle atrophy: Protective role of curcumin.

Chronic hypobaric hypoxia induced muscle atrophy results in decreased physical performance at high altitude. Curcumin has been shown to have muscle sparing effects under cachectic conditions. However, the protective effects of curcumin under chronic hypobaric hypoxia have not been studied till now. Therefore, the present study aims at evaluating the effects of curcumin administration on muscle atrophy under chronic hypobaric hypoxia. Male Sprague Dawley rats were divided into four groups: Control (C)-normoxia exposed, Control Treated (CT)-normoxia exposed and administered with curcumin at a dose of 100 mg/kg body weight for 14 days, Hypoxia (H)-exposed to hypobaric hypoxia for 14 days and Hypoxia Treated (HT)-exposed to hypobaric hypoxia and administered with curcumin for 14 days. Oxidative stress, muscle protein degradation, proteolytic pathways, myosin heavy chain (MHC), CPK activity and muscle histology were performed in gastrocnemius muscle samples of the exposed rats. In addition, fatigue time on treadmill running was also evaluated to observe the effects of curcumin administration on physical performance of the rats. As previously shown, hypobaric hypoxia increased muscle protein degradation via upregulated calpain and ubiquitin-proteolytic pathways. An enhanced oxidative stress has been linked to upregulation of these pathways under hypoxic conditions. Curcumin administration resulted in reduced oxidative stress as well as reduced activity of the proteolytic pathways in HT group as compared to H group thereby resulting in reduced muscle protein degradation under hypobaric hypoxia. Histology of rat muscle revealed an increased number of muscle fibres in HT as compared to H group. Thus, increased number of muscle fibres and decreased muscle proteolysis following curcumin administration, lead to enhanced muscle mass under hypobaric hypoxia resulting in improved physical performance of the rats.

Novel NR5A1 Pathogenic Variants Cause Phenotypic Heterogeneity in 46,XY Disorders of Sex Development.

Steroidogenic factor 1 (NR5A1/SF1) is a key transcription factor that is known to regulate the development of adrenal glands and gonads and is also involved in steroidogenesis. Several pathogenic NR5A1 variants have been reported to cause 46,XY disorders of sex development (DSD), with varying clinical phenotypes ranging from hypospadias to complete gonadal dysgenesis. Most often, the primary cause of DSD is due to variants in gene(s) related to gonadal development or the steroidogenic pathway. In the present study, we have analyzed 64 cases of 46,XY DSD for pathogenic NR5A1 variants. We report a total of 3 pathogenic variants of which 2 were novel (p.Gly22Ser and p.Ser143Asn) and 1 was already known (p.Ser32Asn). Functional studies have revealed that the 2 mutations p.Gly22Ser and p.Ser32Asn could significantly affect DNA binding and transactivation abilities. Further, these mutant proteins showed nuclear localization with aggregate formation. The third mutation, p.Ser143Asn, showed unspeckled nuclear localization and normal DNA binding, but the ability of transcriptional activation was significantly reduced. In conclusion, we recommend screening for NR5A1 pathogenic variants in individuals with features of 46,XY DSD for better diagnosis and management.

Chronic lifestyle diseases display seasonal sensitive comorbid trend in human population evidence from Google Trends.

Seasonal and human physiological changes are important factors in the development of many diseases. But, the study of genuine seasonal impact on these diseases is difficult to measure due to many other environment and lifestyle factors which directly affect these diseases. However, several clinical studies have been conducted in different parts of the world, and it has clearly indicated that certain groups of population are highly subjected to seasonal changes, and their maladaptation can possibly lead to several disorders/diseases. Thus, it is crucial to study the significant seasonal sensitive diseases spread across the human population. To narrow down these disorders/diseases, the study hypothesized that high altitude (HA) associated diseases and disorders are of the strong variants of seasonal physiologic changes. It is because, HA is the only geographical condition for which humans can develop very efficient physiological adaptation mechanism called acclimatization. To study this hypothesis, PubMed was used to collect the HA associated symptoms and disorders. Disease Ontology based semantic similarity network (DSN) and disease-drug networks were constructed to narrow down the benchmark diseases and disorders of HA. The DSN which was further subjected to different community structure analysis uncovered the highly associated or possible comorbid diseases of HA. The predicted 12 lifestyle diseases were assumed to be "seasonal (sensitive) comorbid lifestyle diseases (SCLD)". A time series analyses on Google Search data of the world from 2004-2016 was conducted to investigate whether the 12 lifestyle diseases have seasonal patterns. Because, the trends were sensitive to the term used as benchmark; the temporal relationships among the 12 disease search volumes and their temporal sequences similarity by dynamic time warping analyses was used to predict the comorbid diseases. Among the 12 lifestyle diseases, the study provides an indirect evidence in the existence of severe seasonal comorbidity among hypertension, obesity, asthma and fibrosis diseases, which is widespread in the world population. Thus, the present study has successfully addressed this issue by predicting the SCLD, and indirectly verified them among the world population using Google Search Trend. Furthermore, based on the SCLD seasonal trend, the study also classified them as severe, moderate, and mild. Interestingly, seasonal trends of the severe seasonal comorbid diseases displayed an inverse pattern between USA (Northern hemisphere) and New Zealand (Southern hemisphere). Further, knowledge in the so called "seasonal sensitive populations" physiological response to seasonal triggers such as winter, summer, spring, and autumn become crucial to modulate disease incidence, disease course, or clinical prevention.