Biology subjects

Tripathi, V.

Publications and source records attributed to Tripathi, V..

28 records · Page 2Linked to original sources

bioRxiv2021

AtERF60 negatively regulates ABR1 to modulate salt, drought, and basal resistance in Arabidopsis

APETALA2 (AP2)/ERF family transcription factors (TFs) contribute an important function against various external cues. Our study reports AtERF60, an AP2/ERF TF, which plays a key role in regulating the ABR1 gene, leading to an altered basal resistance response in Arabidopsis. AtERF60 is induced in response to drought, salt, abscisic acid (ABA), salicylic acid (SA), and bacterial pathogen PstDC3000 infection. AtERF60 interacts with DEHYDRATION RESPONSE ELEMENTS and GCC box, indicating its ability to regulate multiple responses. The overexpressing lines of AtERF60 demonstrated increased resistance to PstDC3000 infection, whereas erf60 mutant lines showed increased susceptibility. Complementation of the erf60 mutant background exhibits no significant difference towards PstDC3000 infection compared with the WT Col-0. Microarray and qRT-PCR analysis of overexpression and mutant lines indicated that AtERF60 regulates stress-inducible genes. The induction of these differentially expressed transcripts was significantly increased in erf60 mutant lines, whereas it was reversed in wild-type lines when AtERF60 was complemented. ABR1 was one of the differentially expressed transcripts, and we discovered that AtERF60 interacts with the DRE cis-elements in the ABR1 promoter. Further, qRT-PCR expression analysis after infection with PstDC3000 in AtERF60-OX, erf60 and its complementation background suggest that the mutation in AtERF60 upregulates ABR1 activity, leading to the enhanced susceptibility towards PstDC3000. Conversely, AtERF60 overexpression suppresses ABR1 activity, strengthening the basal resistance responses of Arabidopsis.

plant biology↗

bioRxiv2021

A cost-effective and efficient approach for generating and assembling reagents for conducting real-time PCR

Real-time PCR is a widely used technique for quantification of gene expression. However, commercially available kits for real-time PCR are very expensive. The ongoing coronavirus pandemic has severely hampered the economy in a number of developing countries, resulting in a reduction in available research funding. The fallout of this will result in limiting educational institutes and small enterprises from using cutting edge biological techniques such as real-time PCR. Here, we report a cost-effective approach for preparing and assembling cDNA synthesis and real-time PCR mastermixes with similar efficiencies as commercially available kits. Our results thus demonstrate an alternative to commercially available kits.

molecular biology↗

bioRxiv2021

Weak Task Synchronization of Default Mode Network in Task Based Paradigms

Default mode network (DMN) are the areas of the brain that get deactivated upon the presentation of externally focused, attention demanding tasks. Interestingly, these areas are consistently deactivated irrespective of the nature of the task. Recent evidence shows that the behavior of the DMN may be more nuanced than what was earlier believed. We wanted to investigate the dynamics of DMN further and utilized novel statistical approaches to study the same. We analysed four publicly available datasets (total 223 subjects) across seven tasks related to different cognitive modalities and found out that there is a large amount of variability in the presumed task-negative network across subjects, trials and regions. We designed a method called Inter Trial Temporal Synchronization Analysis (IT-TSA) and Inter Subject TSA (IS-TSA) to analyse variability across trials and subjects respectively. We found out that all task-negative regions and even weak task-positive regions have low synchronization across trials and subjects. We hypothesize that the DMN variability might be more due to neural differences rather than structural or HRF variations. We also investigated the use of non GLM technique like the Inter Subject Correlation (ISC) and found that it may only be suitable to study tightly synchronized task-positive regions. Our study challenges the understanding of DMN as a task-negative region and advances the findings that DMN acts as an active component associated with a unique set of self-referential, autobiographical processes which are deactivated differentially and non linearly across trials and subjects in the presence of extrinsic processes. HighlightsO_LIDefault Mode Network is weakly synchronized across trials and subjects in task paradigms. C_LIO_LIInter Subject Correlation is not suited to detect task-based activations in DMN. C_LIO_LISynchronization Analysis reveals temporal structure in fMRI data. C_LI

neuroscience↗

bioRxiv2021

Loss of Nuclear DNA ligase III Can Revert PARP Inhibitor Resistance in BRCA1-deficient Cells by Increasing DNA Replication Stress

Inhibitors of poly(ADP-ribose) (PAR) polymerase (PARPi) have entered the clinic for the treatment of homologous recombination (HR)-deficient cancers. Despite the success of this approach, preclinical and clinical research with PARPi has revealed multiple resistance mechanisms, highlighting the need for identification of novel functional biomarkers and combination treatment strategies. Functional genetic screens performed in cells and organoids that acquired resistance to PARPi by loss of 53BP1 identified loss of LIG3 as an enhancer of PARPi toxicity in BRCA1-deficient cells. Enhancement of PARPi toxicity by LIG3 depletion is dependent on BRCA1 deficiency but independent of the loss of 53BP1 pathway. Mechanistically, we show that LIG3 loss promotes formation of MRE11-mediated post-replicative ssDNA gaps in BRCA1-deficient and BRCA1/53BP1 double-deficient cells exposed to PARPi, leading to an accumulation of chromosomal abnormalities. LIG3 depletion also enhances efficacy of PARPi against BRCA1-deficient mammary tumors in mice, suggesting LIG3 as a potential therapeutic target.

cancer biology↗

bioRxiv2021

Evaluating Wayfinding Designs in Healthcare Settings through EEG Data and Virtual Response Testing

Wayfinding difficulties in healthcare facilities have been shown to increase anxiety among patients and visitors and reduce staff operational efficiency. Wayfinding-oriented interior design features have proven beneficial, but the evaluation of their performance is hindered by the unique nature healthcare facilities and the expense of testing different navigational aids. This study implemented a virtual-reality testing platform to evaluate the effects of different signage and interior hospital design conditions during navigational tasks; evaluated through behavioral responses and mobile EEG. The results indicated that using color to highlight destinations and increase the contrast of wayfinding information yielded significant benefits when combined with wayfinding-oriented environmental affordances. Neural dynamics from the occipital cortex showed beta-band desynchronization with enhanced color condition and additional theta-band desynchronization with enhanced environmental affordance. This multimodal testing platform has the potential to establish a robust body of evidence for future wayfinding design strategies.

neuroscience↗

bioRxiv2020

A Humanized Antibody against LRG1 that Inhibits Angiogenesis and Reduces Retinal Vascular Leakage

Pathological angiogenesis contributes to morbidity in a number of diseases including cancer, diabetic retinopathy and the neovascular form of age-related macular degeneration, leading to significant efforts to develop effective anti-angiogenic therapeutics for these conditions. The field is dominated by inhibitors of vascular endothelial growth factor (VEGF), yet angiogenesis can also be driven and modified by other factors. We have previously demonstrated that leucine-rich alpha-2-glycoprotein 1 (LRG1) contributes to abnormal vessel growth by activating a TGF{beta} switch. Here we report the development and characterisation of a function-blocking fully humanised IgG4 and its Fab fragment, that bind to LRG1 with high affinity and specificity and inhibit vascular leakage in the mouse model of laser-induced choroidal neovascularisation. In summary, we have developed a therapeutic antibody that targets a VEGF-independent signalling axis, which may be effective in a number of conditions either as monotherapy or in combination with other vascular targeted therapies.

pathology↗

bioRxiv2020

A prevalent and culturable microbiota links ecological balance to clinical stability of the human lung after transplantation

There is accumulating evidence that the lower airway microbiota impacts lung health. However, the link between microbial community composition and lung homeostasis remains elusive. We combined amplicon sequencing and culturomics to characterize the viable bacterial community in 234 longitudinal bronchoalveolar lavage samples from 64 lung transplant recipients and established links to viral loads, host gene expression, lung function, and transplant health. We find that the lung microbiota post-transplant can be categorized into four distinct compositional states, pneumotypes. The predominant balanced pneumotype was characterized by a diverse bacterial community with moderate viral loads, and host gene expression profiles suggesting immune tolerance. The other three pneumotypes were characterized by being either microbiota-depleted, or dominated by potential pathogens, and were linked to increased immune activity, lower respiratory function, and increased risks of infection and rejection. Collectively, our findings establish a link between the lung microbial ecosytem, human lung function, and clinical stability post-transplant. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=191 SRC="FIGDIR/small/106211v1_ufig1.gif" ALT="Figure 1"> View larger version (57K): org.highwire.dtl.DTLVardef@1df2be3org.highwire.dtl.DTLVardef@ba839dorg.highwire.dtl.DTLVardef@116f657org.highwire.dtl.DTLVardef@86d8ac_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2020

Anthrax Toxin as a Molecular Platform to Target Nociceptive Neurons and Modulate Pain

Bacterial toxins are able to act on neurons to modulate signaling and function. Here, we find that nociceptive sensory neurons that mediate pain are enriched in the receptor for anthrax toxins, ANTXR2. Anthrax Edema Toxin (ET) induced cAMP and PKA signaling in Nav1.8+ nociceptive neurons and modulated pain in vivo. Peripherally administered ET mediated mechanical allodynia in naive mice and during B. anthracis infection. Intrathecally administered ET produced analgesic effects, potently blocking pain-like behaviors in multiple mouse models of inflammatory and chronic neuropathic pain. Nociceptor-specific ablation of ANTXR2 attenuated ET-induced signaling and analgesia. Modified anthrax toxin successfully delivered exogenous protein cargo into nociceptive neurons, illustrating utility of the anthrax toxin system as a molecular platform to target pain. ET further induced signaling in human iPSC-derived sensory neurons. Our findings highlight novel interactions between a bacterial toxin and nociceptors that may be utilized for developing new pain therapeutics. SUMMARYANTXR2 expression on nociceptive neurons allows selective targeting and modulation of pain by native and engineered anthrax toxins.

neuroscience↗

bioRxiv2019

The viral SUMO-targeted Ubiquitin Ligase ICP0 is phosphorylated and activated by host kinase Chk2

When the Herpes Simplex virus (HSV) genome enters the nucleus for replication and transcription, phase-segregated nuclear protein bodies called PML Nuclear Bodies (PML NBs) colocalize with the genome and repress it. HSV encodes a SUMO-targeted Ubiquitin ligase ICP0 that degrades PML NBs to alleviate the repression. The molecular mechanism used by ICP0 to target PML NBs is unclear. For reasons unknown, the growth of HSV is dependent on the ATM/Chk2 pathway. Here we identify a bonafide SUMO-Interacting motif in ICP0 (SLS4) that is essential and sufficient to target SUMOylated proteins in PML NBs like PML and Sp100. Phosphorylation of SLS4 creates new salt-bridges between SUMO and SLS4, increases the SUMO/SLS4 affinity and switches ICP0 into a potent STUbL. We also report that ICP0 exploits the kinase Chk2 to phosphorylate SLS4 and enhance its STUbL activity. Our results uncover how a viral STUbL counters antiviral response by exploiting an unprecedented mechanism involving three post-translational modifications; ubiquitination, SUMOylation, and phosphorylation.

biochemistry↗

bioRxiv2019

Phosphorylation by CK2 increases the SUMO-dependent activity of Cytomegalovirus transactivator IE2

Viral factors manipulate the host post-translational modification (PTM) machinery for replication. Distinctly, phosphorylation and SUMOylation can regulate the activity of human cytomegalovirus (HCMV) protein IE2. However, the molecular mechanism of this process is unknown. Taking a structural, biochemical and cellular approach, we uncover a cross-talk of phosphorylation and SUMOylation exploited by IE2. A scan for the SUMO Interacting Motifs (SIMs) revealed two SIMs in IE2. A real-time SUMOylation assay indicated that the N-terminal SIM (IE2-SIM1) enhanced IE2 SUMOylation up to 4-fold. Kinetic analysis and structural studies proved that IE2 is a SUMO cis-E3 ligase. Two putative CK2 sites adjacent to IE2-SIM1 are phosphorylated in-vitro and in cellular conditions. Phosphorylation drastically increased the IE2/SUMO affinity, IE2-SUMOylation and cis-E3 activity of IE2. Additional salt-bridges between the phosphoserines and SUMO account for the higher IE2/SUMO affinity. Phosphorylation also enhances the SUMO-dependent transactivation activity and auto-repression activity of IE2. Together, our findings highlight a novel mechanism where SUMOylation and phosphorylation of the viral cis-E3 ligase and transactivator protein IE2, works in tandem to enable transcriptional regulation of viral genes.\n\nAuthor summaryThe host protein SUMO is a crucial regulator of cellular processes. Conjugation of other proteins to SUMO by a process called SUMOylation, can change the proteins function or localization and regulate downstream cellular events. The SUMO pathway is exploited by viruses to transcribe viral genes and replicate the viral genome. IE2 is an essential gene of human Cytomegalovirus (HCMV), which acts as a transactivator and helps to transcribe other viral proteins required for viral genome replication and viral assembly. SUMOylation of IE2 is necessary for its function. Here, we have uncovered that IE2 functions as a cis-SUMO-E3 ligase, where a SUMO-Interacting Motif (SIM) in IE2 enhances its SUMOylation. Interestingly, phosphorylation of the SIM in IE2 augments its cis-E3 activity to further increase SUMOylation. Moreover, SIM phosphorylation also enhances the interaction between IE2 and SUMOylated binding partners. Thus, we uncover an exciting process, where phosphorylation enhances both covalent and non-covalent interaction of a protein (IE2) and SUMO. We also observe that the cross-talk of phosphorylation and SUMOylation has significant effects on the transactivation function of IE2. Overall, we discover how a viral protein IE2 exploits crosstalk between SUMOylation and Phosphorylation to enhance its activity and in turn, ensure efficient viral replication.

biochemistry↗