• Issue front cover thumbnail

      Volume 41, Issue 4

      December 2016,   pages  563-805

    • The Nobel Prize for understanding autophagy, a cellular mechanism of waste disposal that keeps us healthy

      MEGHA BANSAL GHANSHYAM SWARUP

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      The Nobel Prize in Physiology or Medicine, 2016, was awarded to Prof Yoshinori Ohsumi from TokyoInstitute of Technology, Yokohoma, Japan, for his work that helped in understanding the molecularmechanisms of autophagy, a process used by most eukaryotic cells to degrade a portion of cytoplasmincluding damaged organelles, large protein complexes and aggregated proteins in lysosomes. This processof autophagy (self-eating) maintains cellular homeostasis and helps the cell and the organism to surviveduring periods of stress, such as starvation, by recycling the cellular components to generate amino acidsand nutrients needed for producing energy. Autophagy and ubiquitin-proteasome system are the two majorprotein degradation systems in the cell.The lysosome was identified by Christian de Duve in the 1950s as a membrane bound organelle in thecell that contains degradative enzymes such as proteases, lipases, acid phosphatases, etc. (de Duve, 2005).The term autophagy was coined by Christian de Duve in 1963. Autophagy generally occurs at low level, butit increases under conditions such as stress and differentiation/remodelling of tissues. Autophagy wasprimarily studied by electron microscopy for decades because no molecular markers were available for itsmolecular analysis.

    • Triphala, a formulation of traditional Ayurvedic medicine, shows protective effect against X-radiation in HeLa cells

      YUKI TAKAUJI KENSUKE MIKI JUMA MITA MOHAMMAD NAZIR HOSSAIN MASATAKE YAMAUCHI MITOMU KIOI DAI AYUSAWA MICHIHIKO FUJII

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      Ayurveda is a holistic medical system of traditional medicine, and Triphala is one of the most popular formulations inAyurveda. Triphala is composed of three kinds of herb, Terminalia chebula, Terminalia bellirica, and Emblicaofficinalis. Since Triphala is shown to exhibit a protective activity against ionizing radiation in mice, we investigatedits activity in HeLa cells. We found that Triphala showed the protective effects against X-radiation and bleomycin,both of which generate DNA strand breaks, in HeLa cells. Further, Triphala efficiently eliminated reactive oxygenspecies (ROS) in HeLa cells. Thus, the antioxidant activity of Triphala would likely play a role in its protective actionsagainst X-radiation and bleomycin because both agents damage DNA through the generation of ROS. Theseobservations suggested that the radioprotective activity of Triphala can be, at least partly, studied with the cellscultured in vitro. The simple bioassay system with human cultured cells would facilitate the understanding of themolecular basis for the beneficial effects of Triphala.

    • Engineering and introduction of de novo disulphide bridges in organophosphorus hydrolase enzyme for thermostability improvement

      GHOLAMREZA FARNOOSH KHOSRO KHAJEH ALI MOHAMMAD LATIFI HOSSEIN AGHAMOLLAEI

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      The organophosphorus hydrolase (OPH) has been used to degrade organophosphorus chemicals, as one of the mostfrequently used decontamination methods. Under chemical and thermal denaturing conditions, the enzyme has beenshown to unfold. To utilize this enzyme in various applications, the thermal stability is of importance. The engineeringof de novo disulphide bridges has been explored as a means to increase the thermal stability of enzymes in the rationalmethod of protein engineering. In this study, Disulphide by Design software, homology modelling and moleculardynamics simulations were used to select appropriate amino acid pairs for the introduction of disulphide bridge toimprove protein thermostability. The thermostability of the wild-type and three selected mutant enzymes wereevaluated by half-life, ΔG inactivation (ΔGi) and structural studies (fluorescence and far-UV CD analysis). Dataanalysis showed that half-life of A204C/T234C and T128C/E153C mutants were increased up to 4 and 24 min,respectively; however, for the G74C/A78C mutant, the half-life was decreased up to 9 min. For the T128C/E124Cmutant, both thermal stability and Catalytic efficiency (kcat) were also increased. The half-life and ΔGi results werecorrelated to the obtained information from structural studies by circular dichroism (CD) spectrometry and extrinsicfluorescence experiments; as rigidity increased in A204C/T2234C and T128C/E153C mutants, half-life and ΔGi alsoincreased. For G74C/A78C mutant, these parameters decreased due to its higher flexibility. The results weresubmitted a strong evidence for the possibility to improve the thermostability of OPH enzyme by introducing adisulphide bridge after bioinformatics design, even though this design would not be always successful.

    • Characterization of a DUF820 family protein Alr3200 of the cyanobacterium Anabaena sp. strain PCC7120

      PRASHANTH S RAGHAVAN GAGAN D GUPTA HEMA RAJARAM VINAY KUMAR

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      The hypothetical protein ‘Alr3200’ of Anabaena sp. strain PCC7120 is highly conserved among cyanobacterialspecies. It is a member of the DUF820 (Domain of Unknown Function) protein family, and is predicted to have aDNase domain. Biochemical analysis revealed a Mg(II)-dependent DNase activity for Alr3200 with a specific activityof 8.62×104 Kunitz Units (KU) mg−1 protein. Circular dichroism analysis predicted Alr3200 to have ~40% β-strandsand ~9% α-helical structures. Anabaena PCC7120 inherently expressed Alr3200 at very low levels, and its overexpressionhad no significant effect on growth of Anabaena under control conditions. However, Analr3200+, therecombinant Anabaena strain overexpressing Alr3200, exhibited zero survival upon exposure to 6 kGy of γ-radiation,which is the LD50 for wild type Anabaena PCC7120 as well as the vector control recombinant strain, AnpAM.Comparative analysis of the two recombinant Anabaena strains suggested that it is not the accumulated Alr3200 perse, but its possible interactions with the radiation-induced unidentified DNA repair proteins of Anabaena, whichhampers DNA repair resulting in radiosensitivity.

    • Quaternary ammonium salt N-(dodecyloxycarboxymethyl)-N,N,N-trimethyl ammonium chloride induced alterations in Saccharomyces cerevisiae physiology

      EWA OBŁĄK AGATA PIECUCH EWA MACIASZCZYK-DZIUBIŃSKA DONATA WAWRZYCKA

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      We investigated the influence of the quaternary ammonium salt (QAS) called IM (N-(dodecyloxycarboxymethyl)-N,N,N-trimethyl ammonium chloride) on yeast cells of the parental strain and the IM-resistant mutant (EO25 IMR)growth. The phenotype of this mutant was pleiotropic. The IMR mutant exhibited resistance to ethanol, osmotic shockand oxidative stress, as well as increased sensitivity to UV. Moreover, it was noted that mutant EO25 appears to havean increased resistance to clotrimazole, ketoconazole, fluconazole, nystatin and cycloheximide. It also toleratedgrowth in the presence of crystal violet, DTT and metals (selenium, tin, arsenic). It was shown that the presence ofIM decreased ergosterol level in mutant plasma membrane and increased its unsaturation. These results indicatechanges in the cell lipid composition. Western blot analysis showed the induction of Pma1 level by IM. RT-PCRrevealed an increased PMA1 expression after IM treatment.

    • Genetic and biochemical evidences reveal novel insights into the mechanism underlying Saccharomyces cerevisiae Sae2-mediated abrogation of DNA replication stress

      INDRAJEET GHODKE K MUNIYAPPA

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      In Saccharomyces cerevisiae, the Mre11-Rad50-Xrs2 (MRX) protein complex plays pivotal roles in double-strandbreak (DSB) repair, replication stress and telomere length maintenance. Another protein linked to DSB repair is Sae2,which regulates MRX persistence at DSBs. However, very little is known about its role in DNA replication stress andrepair. Here, we reveal a crucial role for Sae2 in DNA replication stress. We show that different mutant alleles of SAE2cause hypersensitivity to genotoxic agents, and when combined with Δmre11 or nuclease-defective mre11 mutantalleles, the double mutants are considerably more sensitive suggesting that the sae2 mutations synergize with mre11mutations. Biochemical studies demonstrate that Sae2 exists as a dimer in solution, associates preferentially withsingle-stranded and branched DNA structures, exhibits structure-specific endonuclease activity and cleaves thesesubstrates from the 5′ end. Furthermore, we show that the nuclease activity is indeed intrinsic to Sae2. Interestingly,sae2G270D protein possesses DNA-binding activity, but lacks detectable nuclease activity. Altogether, our data suggesta direct role for Sae2 nuclease activity in processing of the DNA structures that arise during replication and DNAdamage and provide insights into the mechanism underlying Mre11-Sae2-mediated abrogation of replication stress-relateddefects in S. cerevisiae.

    • Metabolic alterations derived from absence of Two-Pore Channel 1 at cardiac level

      VANESSA GARCÍA-RÚA SANDRA FEIJÓO-BANDÍN MARÍA GARCÍA-VENCE ALANA ARAGÓN-HERRERA SUSANA B BRAVO DIEGO RODRÍGUEZ-PENAS ANA MOSQUERA-LEAL PAMELA V LEAR JOHN PARRINGTON JANA ALONSO ESTHER ROSELLÓ-LLETÍ MANUEL PORTOLÉS MIGUEL RIVERA JOSÉ RAMÓN GONZÁLEZ-JUANATEY FRANCISCA LAGO

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      Two-pore channels (TPCs or TPCNs) are novel voltage-gated ion channels that have been postulated to act as Ca2+ and/orNa+ channels expressed exclusively in acidic organelles such as endosomes and lysosomes. TPCNs participate in theregulation of diverse biological processes and recently have been proposed to be involved in the pathophysiology ofmetabolic disorders such as obesity, fatty liver disease and type 2 diabetes mellitus. Due to the importance of thesepathologies in the development of cardiovascular diseases, we aimed to study the possible role of two-pore channel 1(TPCN1) in the regulation of cardiac metabolism. To explore the cardiac function of TPCN1, we developed proteomicapproaches as 2-DE-MALDI-MS and LC-MALDI-MS in the cardiac left ventricle of TPCN1 KO and WT mice, and foundalterations in several proteins implicated in glucose and fatty acid metabolism in TPCN1 KO vs. WT mice. The resultsconfirmed the altered expression of HFABP, a key fatty acid transport protein, and of enolase and PGK1, the key enzymes inthe glycolytic process. Finally, in vitro experiments performed in neonatal rat cardiomyocytes, in which TPCN1 was silencedusing siRNAs, confirmed that the downregulation of TPCN1 gene expression increased 2-deoxy-D-[3H]-glucose uptake andGLUT4 mobilization into cell peripherals in cardiac cells. Our results are the first to suggest a potential role for TPCNs incardiac metabolism regulation.

    • Selective serotonin reuptake inhibitor sertraline inhibits voltage-dependent K+ channels in rabbit coronary arterial smooth muscle cells

      HAN SOL KIM HONGLIANG LI HYE WON KIM SUNG EUN SHIN IL-WHAN CHOI AMY L FIRTH HYOWEON BANG YOUNG MIN BAE WON SUN PARK

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      We examined the effects of the selective serotonin reuptake inhibitor (SSRI) sertraline on voltage-dependent K+ (Kv)channels in freshly isolated rabbit coronary arterial smooth muscle cells using the voltage-clamp technique. Sertralinedecreased the Kv channel current in a dose-dependent manner, with an IC50 value of 0.18 μM and a slope value (Hillcoefficient) of 0.61. Although the application of 1 μM sertraline did not affect the steady-state activation curves,sertraline caused a significant, negative shift in the inactivation curves. Pretreatment with another SSRI, paroxetine,had no significant effect on Kv currents and did not alter the inhibitory effects of sertraline on Kv currents. From theseresults, we concluded that sertraline dose-dependently inhibited Kv currents independently of serotonin reuptakeinhibition by shifting inactivation curves to a more negative potential.

    • Expressions of visual pigments and synaptic proteins in neonatal chick retina exposed to light of variable photoperiods

      KUMAR ABHIRAM JHA TAPAS C NAG SHASHI WADHWA TARA SANKAR ROY

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      Light causes damage to the retina, which is one of the supposed factors for age-related macular degeneration inhuman. Some animal species show drastic retinal changes when exposed to intense light (e.g. albino rats). Althoughbirds have a pigmented retina, few reports indicated its susceptibility to light damage. To know how light influences acone-dominated retina (as is the case with human), we examined the effects of moderate light intensity on the retina ofwhite Leghorn chicks (Gallus g. domesticus). The newly hatched chicks were initially acclimatized at 500 lux for 7days in 12 h light: 12 h dark cycles (12L:12D). From posthatch day (PH) 8 until PH 30, they were exposed to 2000 luxat 12L:12D, 18L:6D (prolonged light) and 24L:0D (constant light) conditions. The retinas were processed fortransmission electron microscopy and the level of expressions of rhodopsin, S- and L/M cone opsins, and synapticproteins (Synaptophysin and PSD-95) were determined by immunohistochemistry and Western blotting. Rearing in24L:0D condition caused disorganization of photoreceptor outer segments. Consequently, there were significantlydecreased expressions of opsins and synaptic proteins, compared to those seen in 12L:12D and 18L:6D conditions.Also, there were ultrastructural changes in outer and inner plexiform layer (OPL, IPL) of the retinas exposed to24L:0D condition. Our data indicate that the cone-dominated chick retina is affected in constant light condition, withchanges (decreased) in opsin levels. Also, photoreceptor alterations lead to an overall decrease in synaptic proteinexpressions in OPL and IPL and death of degenerated axonal processes in IPL.

    • Long non-coding RNA HOTAIR regulates proliferation and invasion via activating Notch signalling pathway in retinoblastoma

      CHANGXIA DONG SHAOYI LIU YONGBIN LV CHUNPING ZHANG HEYING GAO LIXIA TAN HONG WANG

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      Retinoblastoma is the most frequently occurring tumour in the eyes in early childhood. Novel targets that areimportant for the diagnosis or treatment of retinoblastoma could be valuable in increasing the survival rate of patientsaffected by this disease. Long non-coding RNAs (lncRNAs) are a recently discovered type of RNAs with no proteincodingfunction; yet it has become increasingly clear that lncRNAs are responsible for important gene regulatoryfunctions in various diseases. In this study, the expression of lncRNA HOTAIR was measured by qRT-PCR, andHOTAIR expression was found to be significantly upregulated in human retinoblastomas tissues as compared with thatin paracancerous tissues. Knockdown of HOTAIR restricted the proliferation and invasion of the more invasiveretinoblastoma Y79 cells, and led to G0/G1 arrest, possibly through inhibiting phospho-RB1, RB1 and CCNE.Furthermore, we found that the Notch signalling pathway was activated abnormally in retinoblastoma cell lines, whileknockdown of HOTAIR attenuated the endogenous Notch signalling pathway in vitro and in vivo. In addition,knockdown of HOTAIR could inhibit the tumour progression in a xenograft model of retinoblastoma. In summary,our findings indicate that HOTAIR may play important roles in retinoblastoma progression via Notch pathway.HOTAIR has the potential to enhance the development of novel targeted diagnostic and therapeutic approaches forretinoblastoma.

    • Reserpine requires the D2-type receptor, dop-3, and the exoribonuclease, eri-1, to extend the lifespan in C. elegans

      KOPAL SAHARIA RANJEET KUMAR KULDEEP GUPTA SHRILEKHA MISHRA JAMUNA R SUBRAMANIAM

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      Lifespan extension is an all systems encompassing event. Involvement of reduced insulin/IGF1 signalling is wellworked out, first in the model organism Caenorhbaditis elegans followed by other systems including humans. But therole of neuronal component in lifespan extension is not well understood due to the refractory nature of neurons tosmall RNA interference (sRNAi) in C. elegans. Earlier, we have demonstrated that an antihypertensive drug,reserpine, extends lifespan through modulation of neurotransmitter release, especially, acetylcholine, in C. elegans.Intriguingly, the reserpine mediated lifespan extension (RMLE) does not happen through the known longevitypathways. Here, we report that the D2-type dopamine receptor (DOP-3), which acts through the inhibitory Gproteincoupled (Gαi) pathway mediated signalling is partly required for RMLE. In the dop-3 loss of function mutantRMLE is shortened. DOP-3 acts through Gαo (goa-1). One of the downstream targets of G protein signalling is thetranscription factor, jun-1. MRP-1, an ATP binding cassette transporter, belonging to the multidrug resistance proteinfamily is one of the genes turned on by JUN-1. RMLE is shortened in dop-3→goa-1→jun1→mrp-1 loss of functionmutants, elucidating the contribution of dop-3 signalling. The dop-3 receptor system is known to inhibit acetylcholinerelease. This suggests dopamine receptor, dop-3 could be contributing to the modulation of acetylcholine release byreserpine. ERI-1 is a 3′-5′ exoribonuclease, one of the negative regulators of sRNAi, whose loss of function makesneurons amenable to siRNA. In the absence of eri-1, RMLE is shortened. In the dop-3 loss-of-function background,lack of eri-1 completely abolishes RMLE. This suggests that dop-3 and eri-1 act in independent parallel pathways forRMLE and these two pathways are essential and sufficient for the longevity enhancement by reserpine in C. elegans.

    • Ayurvedic Amalaki Rasayana promotes improved stress tolerance and thus has anti-aging effects in Drosophila melanogaster

      VIBHA DWIVEDI SUBHASH C LAKHOTIA

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      Amalaki Rasayana (AR) is a common Ayurvedic herbal formulation of Phyllanthus emblica fruits and some otheringredients, and is used for general good health and healthy aging. We reported it to improve life history traits and tosuppress neurodegeneration as well as induced apoptosis in Drosophila. The present study examines responses ofDrosophila reared on AR-supplemented food to crowding, thermal or oxidative stresses. Wild-type larvae/flies rearedon AR-supplemented food survived the various cell stresses much better than those reared on control food. AR-fedmutant park13 or DJ-1βDelta93 (Parkinson’s disease model) larvae/flies, however, showed only partial or no protection,respectively, against paraquat-induced oxidative stress, indicating essentiality of DJ-1β for AR-mediated oxidativestress tolerance. AR feeding reduced the accumulation of reactive oxygen species (ROS) and lipid peroxidation evenin aged (35-day-old) wild-type flies while enhancing superoxide dismutase (SOD) activity. We show that while Hsp70or Hsp83 expression under normal or stress conditions was not affected by AR feeding, Hsp27 levels were elevated inAR-fed wild-type control as well as heat-shocked larvae. Therefore, besides the known anti-oxidant activity ofPhyllanthus emblica fruits, dietary AR also enhances cellular levels of Hsp27. Our in vivo study on a model organismshows that AR feeding significantly improves tolerance to a variety of cell stresses through reduced ROS and lipidperoxidation on the one hand, and enhanced SOD activity and Hsp27 on the other. The resulting better homeostasisimproves life span and quality of organism’s life.

    • A lateralized functional auditory network is involved in anuran sexual selection

      FEI XUE GUANGZHAN FANG XIZI YUE ERMI ZHAO STEVEN E BRAUTH YEZHONG TANG

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      Right ear advantage (REA) exists in many land vertebrates in which the right ear and left hemisphere preferentiallyprocess conspecific acoustic stimuli such as those related to sexual selection. Although ecological and neural mechanismsfor sexual selection have been widely studied, the brain networks involved are still poorly understood. In this study weused multi-channel electroencephalographic data in combination with Granger causal connectivity analysis to demonstrate,for the first time, that auditory neural network interconnecting the left and right midbrain and forebrain functionasymmetrically in the Emei music frog (Babina daunchina), an anuran species which exhibits REA. The results showedthe network was lateralized. Ascending connections between the mesencephalon and telencephalon were stronger in theleft side while descending ones were stronger in the right, which matched with the REA in this species and implied thatinhibition from the forebrainmay induce REA partly. Connections from the telencephalon to ipsilateral mesencephalon inresponse to white noise were the highest in the non-reproductive stage while those to advertisement calls were the highestin reproductive stage, implying the attention resources and living strategy shift when entered the reproductive season.Finally, these connection changes were sexually dimorphic, revealing sex differences in reproductive roles.

    • Functional analyses of Populus euphratica brassinosteroid biosynthesis enzyme genes DWF4 (PeDWF4) and CPD (PeCPD) in the regulation of growth and development of Arabidopsis thaliana

      JIANPING SI YAN SUN LU WANG YING QIN CHONGYING WANG XINYU WANG

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      DWF4 and CPD are key brassinosteroids (BRs) biosynthesis enzyme genes. To explore the function of Populuseuphratica DWF4 (PeDWF4) and CPD (PeCPD), Arabidopsis thaliana transgenic lines (TLs) expressing PeDWF4,PeCPD or PeDWF4 plus PeCPD, namely PeDWF4-TL, PeCPD-TL and PeCP/DW-TL, were characterized. Comparedwith wild type (WT), the changes of both PeDWF4-TL and PeCPD-TL in plant heights, silique and hypocotylslengths and seed yields were similar, but in bolting time and stem diameters, they were opposite. PeCP/DW-TL wasmore in plant heights and the lengths of primary root, silique, and fruit stalk, but less in silique numbers and seedyields than either PeDWF4-TL or PeCPD-TL. PeDWF4 and PeCPD specially expressed in PeDWF4-TL or PeCPDTL,and the transcription level of PeDWF4 was higher than that of PeCPD. In PeCP/DW-TL, their expressions wereall relatively reduced. Additionally, the expression of PeDWF4 and PeCPD differentially made the expression levelsof AtDWF4, AtCPD, AtBR6OX2, AtFLC, AtTCP1 and AtGA5 change in the TLs. The total BRs contents werePeDWF4-TL > PeCP/DW-TL > WT > PeCPD-TL. These results imply that PeDWF4 is functionally not exactly thesame as PeCPD and there may be a synergistic and antagonistic effects in physiology between both of them in theregulation of plant growth and development.

    • PluriPred: AWeb server for predicting proteins involved in pluripotent network

      SUKHEN DAS MANDAL SUDIPTO SAHA

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      Pluripotency is a unique property of stem cells that allows them to differentiate into all types of adult cells or maintainthe self-renewal property. PluriPred predicts whether a protein is involved in pluripotency from primary proteinsequence using manually curated pluripotent proteins as training datasets. Machine learning techniques (MLTs) suchas Support Vector Machine (SVM), Naïve Base (NB), Random Forest (RF), and sequence alignment techniqueBLAST were used in our study. The combination of SVM and PSI-BLAST was our proposed best model, whichobtained a sensitivity of 77.40%, specificity of 79.72%, accuracy of 79.2%, and area under the ROC curve was 0.82using 5-fold cross-validation. Furthermore, PluriPred gives the confidence of the prediction from training dataset’sSVM score distribution and p-value from BLAST. We validated our proposed model with the other existing highthroughputstudies using blind/independent datasets. Using PluriPred, 233 novel core and 323 novel extended corepluripotent proteins from mouse proteome, and 167 novel core and 385 extended core pluripotent proteins fromhuman proteome, were predicted with high confidence. The Web application of PluriPred is available from bicresources.jcbose.ac.in/ssaha4/pluripred/. Many pluripotent genes/proteins take part in protein-protein networks associatedwith stem cell, cancer, and developmental biology, and we believe that PluriPred will help in these research.

    • Brain-derived neurotrophic factor and early-life stress: Multifaceted interplay

      NATALYA P BONDAR TATIANA I MERKULOVA

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      The brain-derived neurotrophic factor (BDNF) is a key regulator of neural development and plasticity. Longtermchanges in the BDNF pathway are associated with childhood adversity and adult depression symptoms.Initially, stress-induced decreases in the BDNF pathway were found in some studies, but subsequent reportsindicated the relationship between stress and BDNF to be much more complex, and the concept wassignificantly revised. In the present mini-review, we focus on the structure and regulation of the Bbnf geneas well as on the stress–BDNF interactions under early-life adverse conditions.

    • Heterochromatin and the molecular mechanisms of ‘parent-of-origin’ effects in animals

      PRIM B SINGH

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      Twenty five years ago it was proposed that conserved components of constitutive heterochromatin assemble heterochromatinlikecomplexes in euchromatin and this could provide a general mechanism for regulating heritable (cell-to-cell) changesin gene expressibility. As a special case, differences in the assembly of heterochromatin-like complexes on homologouschromosomes might also regulate the parent-of-origin-dependent gene expression observed in placental mammals. Here,the progress made in the intervening period with emphasis on the role of heterochromatin and heterochromatin-likecomplexes in parent-of-origin effects in animals is reviewed.

    • Recent excitements in protein NMR: Large proteins and biologically relevant dynamics

      SAI CHAITANYA CHILIVERI MANDAR V DESHMUKH

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      The advent of Transverse Relaxation Optimized SpectroscopY (TROSY) and perdeuteration allowed biomolecularNMR spectroscopists to overcome the size limitation barrier (~20 kDa) in de novo structure determination of proteins.The utility of these techniques was immediately demonstrated on large proteins and protein complexes (e.g. GroELGroES,ClpP protease, Hsp90-p53, 20S proteasome, etc.). Further, recent methodological developments such asResidual Dipolar Couplings and Paramagnetic Relaxation Enhancement allowed accurate measurement of long-rangestructural restraints. Additionally, Carr-Purcell-Meiboom-Gill (CPMG), rotating frame relaxation experiments (R1ρ)and saturation transfer experiments (CEST and DEST) created never-before accessibility to the μs–ms timescaledynamic parameters that led to the deeper understanding of biological processes. Meanwhile, the excitement in thefield continued with a series of developments in the fast data acquisition methods allowing rapid structural studies onless stable proteins. This review aims to discuss important developments in the field of biomolecular NMRspectroscopy in the recent past, i.e., in the post TROSY era. These developments not only gave access to the structuralstudies of large protein assemblies, but also revolutionized tools in the arsenal of today’s biomolecular NMR and pointto a bright future of biomolecular NMR spectroscopy.

    • The autoregulatory loop: A common mechanism of regulation of key sex determining genes in insects

      SURESH KUMAR SAWANTH GAJULA GOPINATH NAGRAJ SAMBRANI KALLARE P ARUNKUMAR

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