Biology subjects

Mukundan, S.

Publications and source records attributed to Mukundan, S..

7 recordsLinked to original sources

bioRxiv2025

Overhang Optimizer for Golden Gate Assembly (OOGGA)

Golden Gate cloning is a popular method for achieving ordered assembly of DNA fragments using complementary four-base pair overhangs. The success of this method relies on the ability of DNA ligases to ligate the different fragments. This ability is affected by the relative efficiency and accuracy with which the ligase recognizes and joins overhanging regions. In this study, we report a dynamic programming approach, called Overhang Optimizer for Golden Gate Assembly (OOGGA), that optimizes these overhangs for their accuracy and efficiency. OOGGA provides the theoretically optimal fragments for Golden Gate assembly, provided a DNA sequence, a length range of the fragments and/or the number of required fragments. As expected, OOGGA outperforms NEBridge SplitSet(R) in both fidelity and efficiency metrics in a set of 3 example DNA sequences. OOGGA also provides support of degenerate nucleotide codes, options to exclude/include motifs for predictions and weights to bias the program for efficiency and fidelity.

bioinformatics↗

bioRxiv2024

Light and temperature-dependent developmental role of Auxin Binding Protein 1 (ABP1) in Arabidopsis thaliana

Auxin Binding Protein 1 (ABP1) is a small glycoprotein of about 22 kDa that has long been debated as the auxin receptor, and has been put into question for its unclear functions. Despite its conservancy during land plant evolution, its precise role in plant development is still elusive. Historically, it has been implicated in various rapid responses such as membrane polarization, calcium fluxes, TMK1-based cell-surface signalling, auxin canalization, etc. A relatively recent observation questioning the role of ABP1 in plant development led us to explore its probable functions if any. In the current study, we reinvestigated the plausible function of ABP1 using its CRISPR-based loss-of-function mutants, namely abp1-C1 and abp1-C2. Here we show that, ABP1 acts as a positive regulator for primary root elongation under red and secondary root elongation under blue light in seedlings at 22 {degrees}C. Under red light at 18 {degrees}C, it has a negative effect on hypocotyl growth inhibition. Furthermore, it is involved in flowering time control at 18 {degrees}C irrespective of the photoperiod. We show that the transcript levels of Phytochrome B (phyB) and GIGANTEA (GI) are altered in the mutants of ABP1 under red light and low temperature (18 {degrees}C) regimes. Further, ABP1 show a pronounced role in tolerance to dehydration induced due to low temperature (18 {degrees}C), which correlates with an increase in endogenous abscisic acid (ABA), salicylic acid (SA), and a decrease in jasmonic acid (JA) content in leaves. The functional roles of ABP1 under red light, low temperature and dehydration tolerance in Arabidopsis thaliana once again frames it to be an important regulator under adverse and varied conditions that the plant can experience, and thus opened up new avenues for further studies.

plant biology↗

bioRxiv2023

High-affinity biomolecular interactions are modulated by low-affinity binders

Molecular interactions play a central role in all biological processes. The strength of these interactions is often characterized by their dissociation constants (KD). The high affinity interactions (KD [≤] 10-8 M) are crucial for the proper execution of cellular processes and are thus extensively investigated. Detailed molecular and biochemical analyses of such high affinity interactions have lent considerable support to the concept of binary on/off switches in different biological contexts. However, such studies have typically discounted the presence of low-affinity binders (KD > 10-5 M) in the cellular environment. In this study, we have assessed the potential influence of such low affinity binders on high affinity interactions. By employing Gillespie stochastic simulations as well as continuous methods, we demonstrate that the presence of low-affinity binders can indeed alter the kinetics and the steady state of high-affinity interactions. We refer to this effect as herd regulation and have evaluated its possible impact in two different contexts including sex determination in Drosophila melanogaster and in signalling systems that employ molecular thresholds. Lastly, we suggest an in vitro experimental strategy to validate herd regulation. Based on these analyses, we propose that low-affinity binders are likely prevalent in different biological contexts where the outcomes are contingent upon threshold value determinants and thus potentially impact their homeostatic regulation.

systems biology↗

bioRxiv2022

HLA-II-dependent neuroimmune changes in Group A Streptococcal Necrotizing Fasciitis

IntroductionStreptococcus pyogenes (Group A Streptococcus, GAS) bacteria cause a spectrum of human diseases ranging from self-limiting pharyngitis and mild uncomplicated skin infections (impetigo, erysipelas, cellulitis) to highly morbid and rapidly invasive life-threatening infections such as streptococcal toxic shock syndrome and necrotizing fasciitis (NF). HLA-Class II allelic polymorphisms are linked with differential outcomes and severity of GAS infections. The dysregulated immune response and peripheral cytokine storm elicited due to invasive GAS infections increase the risk for toxic shock and multiple organ failure in genetically susceptible individuals. We hypothesized that while the host immune mediators regulate the immune responses against peripheral GAS infections, these interactions may simultaneously trigger neuropathology and, in some cases, induce persistent alterations in the glial phenotypes. Here we studied the consequences of peripheral GAS skin infection on the brain in an HLA-II transgenic mouse model of GAS NF with and without treatment with an antibiotic, clindamycin (CLN). MethodsMice expressing the human HLA-II DR3 (DR3) or the HLA-II DR4 (DR4) allele were divided into three groups: i) uninfected controls, ii) subcutaneously infected with a clinical GAS strain isolated from a patient with GAS NF, and iii) GAS infected with CLN treatment (10mg/kg/5 days, intraperitoneal). The groups were monitored for 15 days post-infection. Skin GAS burden and lesion area, splenic and hippocampal mRNA levels of inflammatory markers, and immunohistochemical changes in hippocampal GFAP and Iba-1 immunoreactivity were assessed. ResultsSkin GAS burden and hippocampal mRNA levels of inflammatory markers S100A8/A9, IL-1{beta}, IL-33, inflammasome-related caspase-1 (Casp1), and NLRP6 were elevated in infected DR3 but not DR4 mice. The levels of these markers were significantly reduced following CLN treatment in DR3 mice. Although GAS was not detectable in the brain, astrocyte and microglia activation were evident from increased GFAP and Iba-1 mRNA levels respectively, in DR3 and DR4 mice. However, CLN treatment significantly reduced GFAP immunoreactivity in DR3 mice and not DR4 mice. ConclusionOur data suggest a skin-brain axis during GAS NF demonstrating that peripherally induced pathological conditions regulate neuroimmune and gliotic events, and CLN may attenuate peripheral infection and subsequent neuroimmune changes in an HLA-II-dependent manner.

neuroscience↗

bioRxiv2021

Metallothionein 3-zinc axis suppresses caspase-11 inflammasome activation and impairs antibacterial immunity

Non-canonical inflammasome activation by mouse caspase-11 (or human CASPASE- 4/5) is crucial for the clearance of certain gram-negative bacterial infections, but can lead to severe inflammatory damage. Factors that promote non-canonical inflammasome activation are well recognized, but less is known about the mechanisms underlying its negative regulation. Herein, we identify that the caspase-11 inflammasome in mouse and human macrophages (M{phi}) is negatively controlled by the zinc (Zn2+) regulating protein, metallothionein 3 (MT3). Upon challenge with intracellular lipopolysaccharide (iLPS), M{phi} increased MT3 expression that curtailed the activation of caspase-11 and its downstream targets caspase-1 and interleukin (IL)-1{beta}. Mechanistically, MT3 increased intramacrophage Zn2+ to downmodulate the TRIF-IRF3-STAT1 axis that is prerequisite for caspase-11 effector function. MT3 suppressed activation of the caspase-11 inflammasome, while caspase-11 and MT3 synergized in impairing antibacterial immunity. The present study identifies an important yin-yang relationship between the non-canonical inflammasome and MT3 in controlling inflammation and immunity to gram- negative bacteria. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=149 HEIGHT=200 SRC="FIGDIR/small/454033v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@15070f1org.highwire.dtl.DTLVardef@27174forg.highwire.dtl.DTLVardef@6b48beorg.highwire.dtl.DTLVardef@174b850_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2019

Does Sex Modify an Association of Electrophysiological Substrate with Sudden Cardiac Death? The Atherosclerosis Risk in Communities (ARIC) Study

BackgroundSex is a well-recognized risk factor for sudden cardiac death (SCD). Sex differences in electrophysiological (EP) substrate of SCD are known. However, it remains unknown whether sex can modify an association of EP substrate with SCD.\n\nMethodsParticipants from the Atherosclerosis Risk in Communities study with analyzable ECGs (n=14,725; age, 54.2{+/-}5.8 yrs; 55% female, 74% white) were included. EP substrate was characterized by traditional 12-lead ECG (heart rate, QRS, QTc, Cornell voltage), spatial ventricular gradient (SVG) and sum absolute QRST integral (SAI QRST) metrics. Two competing outcomes were adjudicated SCD and nonSCD. Interaction of ECG metrics with sex was studied in Cox proportional hazards and Fine-Gray competing risk models. Relative hazard ratio (RHR) and relative sub-hazard ratio (RSHR) with a 95% confidence interval for SCD and nonSCD risk for women relative to men were calculated. Model 1 was adjusted for prevalent cardiovascular disease (CVD) and risk factors. Time-updated model 2 was additionally adjusted for incident non-fatal CVD.\n\nResultsOver a median follow-up of 24.4 years, there were 530 SCDs (incidence 1.72 (1.58-1.88)/1000 person-years) and 2,178 nonSCDs (incidence 7.09; (6.80-7.39)/ 1000 person-years). Women experienced a greater than men risk of SCD associated with Cornell voltage (RHR 1.18(1.06-1.32); P=0.003), SAI QRST (RHR 1.16(1.04-1.30); P=0.007), area SVG magnitude (RHR 1.24(1.05-1.45); P=0.009), and peak SVG magnitude (RHR 1.22(1.04-1.44); P=0.018), independently from incident CVD. Greater risk of SCD for women than men associated with QRS duration (RHR 1.24(1.07-1.44); P=0.004) and QTc (RSHR 1.15(1.02-1.30); P=0.025) was explained by incident CVD. Furthermore, women had greater odds of SCD associated with heart rate (RSHR 1.19(1.01-1.40); P=0.036), independently of incident CVD.\n\nConclusionsSex modifies an association of EP substrate with SCD. In women, global EP substrate is associated with up to 27% greater risk of SCD than in men. Development of sex-specific risk scores of SCD is necessary. Further studies of mechanisms behind sex differences in EP substrate of SCD are warranted.

epidemiology↗

bioRxiv2019

Dynamic Predictive Accuracy of Electrocardiographic Biomarkers of Sudden Cardiac Death within a Survival Framework: The Atherosclerosis Risk in Communities (ARIC) study

BackgroundThe risk of sudden cardiac death (SCD) is known to be dynamic. However, an accuracy of a dynamic SCD prediction, and \"expiration date\" of ECG biomarkers is unknown. Our goal was to measure dynamic predictive accuracy of ECG biomarkers of SCD and competing outcomes.\n\nMethodsAtherosclerosis Risk In Community study participants with analyzable digital ECGs were included (n=15,768; 55% female, 73% white, age 54.2{+/-}5.8 y). ECGs of 5 follow-up visits were analyzed. Global electrical heterogeneity (GEH) and traditional ECG metrics were measured. Adjudicated SCD served as the primary outcome; non-sudden cardiac death served as competing outcome. Time-dependent area under the (receiver operating characteristic) curve (AUC) analysis was performed to assess prediction accuracy of a continuous biomarker in a period of 3,6,9 months, and 1,2,3,5,10, and 15 years, using survival analysis framework.\n\nResultsOver a median 24.4 y follow-up, there were 581 SCDs (incidence 1.77 (95%CI 1.63-1.92)/1,000 person-years), and 838 nonSCDs [2.55 (95%CI 2.39-2.73)]. Resting heart rate was the strongest (AUC 0.930) short-term (3-month) non-specific SCD predictor, whereas spatial peak QRS-T angle predicted specifically SCD 15 years after ECG recording (AUC 0.719). QRS duration (AUC 0.885) and QTc (AUC 0.711) short-term predicted advanced structural heart disease better than SCD. \"Expiration date\" for most ECG biomarkers was two years after ECG recording. GEH significantly improved reclassification of SCD risk beyond age, sex, race, diabetes, hypertension, coronary heart disease and stroke.\n\nConclusionShort-term predictors of SCD, nonSCD, and biomarkers of long-term SCD risk differed, reflecting differences in transient vs. persistent SCD substrates.

epidemiology↗