Biology subjects

Pal, U.

Publications and source records attributed to Pal, U..

At least 19 recordsLinked to original sources

bioRxiv2026

Develop a durable, memory-driven, CspZ-targeting Lyme disease vaccine by rationale adjuvant selection

Rational adjuvant selection is a systematic approach based on adjuvant-mediated immunomodulation to identify safe vaccine regimens that enhance protective immunity. Transmitted through ticks and caused by the bacterium Borrelia burgdorferi (Bb), Lyme disease (LD) is the most common vector-borne disease in the Northern hemisphere. There are no effective vaccines, making it suitable for testing the concept of rational adjuvant selection. Here, we formulated our previously developed and effective LD vaccine antigen, CspZ-YAC187S, with different adjuvants suitable for human use; we analyzed the immune response by transcriptomics and tested the vaccine efficacy after Bb infection. We identified Alum-CpG and Alum-Gal to elicit the highest titers of CspZ-YAC187S-dependent protective antibodies and robust levels of protection but through distinct mechanisms of immunomodulation. We demonstrated that immunization with Alum-CpG formulated CspZ-YAC187S provided up to nine months of protective bactericidal antibody titers, as well as recall-memory response to prevent LD after natural infection. Immunity was linked to elevated levels of IgG1 memory cells in the vaccine-triggered immune responses. This work thus identified a durable, memory immunity-driven LD vaccine, ultimately paving the road to understanding the mechanisms of rationale adjuvant selection for vaccine development.

immunology↗

bioRxiv2025

A positive feedback loop of MPK3-PIN1A trafficking-auxin flux trio governs dual gravitropic and wounding response in rice

The auxin flow in plants plays a pivotal role in gravitropic organ movement and tissue regeneration following injury. However, the regulatory aspects of these phenomena are not entirely understood. In this study, we found that hyperactivity of MAP Kinase 3 (MPK3) led to downregulation of PINs and reduction in tissue auxin content, impairing gravitropic response of rice and accelerating tissue senescence following mechanical wounding. The MPK3 was found to phosphorylate PIN1A majorly at its Ser351 residue, mutation of which into its phospho-mimic variant enhanced latters endosomal trafficking and consequently improved auxin flux. Further, overexpression of the phospho-null variant of PIN1A lowered gravitropic response of rice, thus causing wider tiller angle similar to the pin1a knockout lines. Additionally, upon wounding, their tissue senescence was faster in comparison to the overexpression lines of wild-type and phospho-mimic variants of PIN1A, which also displayed improved resistance to wounding by Bipolaris oryzae. The PIN1A trafficking-mediated auxin flux also regulated MPK3 activity via a positive feedback loop. Together, these results uncover a novel feedback loop encompassing "MPK3-PIN1A trafficking-auxin flux" trio that regulates dual gravitropic and wounding response in rice.

plant biology↗

bioRxiv2024

Dengue Protease-Mediated Activation and Polarization of Macrophages

Controlled activation of macrophages to achieve desired phenotypes in vivo and in vitro could lead to possible treatments for several inflammatory and proliferative diseases. This investigation uniquely established that dengue protease enhances the polarization and activation of human and murine macrophages following extracellular exposure. It was observed that macrophages have a robust response to extracellularly administered purified dengue NS2B/NS3 protease; it stimulated the macrophages in classical M1 directions and enhanced both the mitogen-activated protein kinases (MAPK) and c-Jun N-terminal kinases (JNK). The cells exhibited an increase in the intracellular levels of proinflammatory cytokines such as IL-2, IL-6, IL-12, and IFN-{gamma} and an elevation of phosphorylated versions of Akt, P38 MARK, SAPK, and ERK. In tandem, this investigation also established that the heightened responses of stimulated cells were associated with an increase in the cellular reactive oxygen species (ROS) level and the nuclear translocation of the NF-{kappa}B (P65) protein. The cell viability assay showed that NS2B/NS3 protease exerts no major toxic effect on macrophages after 24 hours of treatment, even at a dosage (20 g/ml) which was four times higher than the effective dose (5 g/ml). Remarkably, we also observed that the native form of the viral protease, which drives its enzyme activities, had no bearing on the antigenic qualities of the enzyme. Thus, our study highlighted the efficacy of dengue viral NS2B/NS3 protease as a non-toxic ex vivo macrophage activating/polarizing agent and may serve a vital role in macrophage-based cell therapy in the near future. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=101 SRC="FIGDIR/small/624958v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@d29cd1org.highwire.dtl.DTLVardef@1357e96org.highwire.dtl.DTLVardef@109cc52org.highwire.dtl.DTLVardef@45536b_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2024

Mechanistic insights into structure-based design of a Lyme disease subunit vaccine

The quality of protective immunity plays a critical role in modulating vaccine efficacy, with native antigens often not able to trigger sufficiently strong immune responses for pathogen killing. This warrants creation of structure-based vaccine design, leveraging high-resolution antigen structures for mutagenesis to improve protein stability and efficient immunization strategies. Here, we investigated the mechanisms underlying structure-based vaccine design using CspZ-YA, a vaccine antigen from Borrelia burgdorferi, the bacteria causing Lyme disease (LD), the most common vector-borne disease in the Northern Hemisphere. Compared to wild-type CspZ-YA, we found CspZ-YAI183Y and CspZ-YAC187S required lower immunization frequency to protect mice from LD-associated manifestations and bacterial colonization. We observed indistinguishable human and mouse antigenicity between wild-type and mutant CspZ-YA proteins after native infection or active immunization. This supports our newly generated, high-resolution structures of CspZ-YAI183Y and CspZ-YAC187S, showing no altered surface epitopes after mutagenesis. However, CspZ-YAI183Y and CspZ-YAC187S favored the interactions between helices H and I, consistent with their elevated thermostability. Such findings are further strengthened by increasing ability of protective CspZ-YA monoclonal antibodies in binding to CspZ-YA at a physiological temperature (37{degrees}C). Overall, this study demonstrated enhanced intramolecular interactions improved long-term stability of antigens while maintaining protective epitopes, providing a mechanism for structure-based vaccine design. These findings can ultimately be extended to other vaccine antigens against newly emerging pathogens for the improvement of protective immunity.

immunology↗

bioRxiv2024

Duplex PCR assay to determine sex and mating status of Ixodes scapularis (Acari: Ixodidae), vector of the Lyme disease pathogen

Ticks are a major health threat to humans and other animals, through direct damage, toxicoses, and transmission of pathogens. An estimated half a million people are treated annually in the United States of America for Lyme disease, a disease caused by the bite of a black-legged tick (Ixodes scapularis Say) infected with the bacterial pathogen Borrelia burgdorferi. This tick species also transmits another six human-disease causing pathogens, for which vaccines are currently unavailable. While I. scapularis are sexually dimorphic at the adult life stage, the DNA sequence differences between male and female I. scapularis that could be used as a sex-specific marker have not yet been established. We determine the sex-specific DNA sequences for I. scapularis (male heterogametic system with XY), using whole-genome resequencing and restriction site-associated DNA sequencing. Then we identify a male-specific marker that we use as the foundation of a molecular sex identification method (duplex PCR) to differentiate the sex of an I. scapularis tick. In addition, we provide evidence that this molecular sexing method can establish the mating status of adult females that have been mated and inseminated with male-determining sperm. Our molecular tool allows the characterization of mating and sex-specific biology across development for I. scapularis, a major pathogen vector, which is crucial for a better understanding of their biology and controlling tick populations.

genomics↗

bioRxiv2024

Complement therapeutic Factor H-IgG proteins as pre-exposure prophylaxes against Lyme borreliae infections

Lyme disease (LD) is the most common vector-borne disease in the northern hemisphere and is caused by the bacteria Borrelia burgdorferi sensu lato (also known as Lyme borreliae) with no effective prevention available. Lyme borreliae evade complement killing, a critical arm of host immune defense, by producing outer surface proteins that bind to a host complement inhibitor, factor H (FH). These outer surface proteins include CspA and CspZ, which bind to the 6th and 7th short consensus repeats of FH (SCR(6-7)), and the OspE family of proteins (OspE), which bind to the 19th and 20th SCR (SCR19-20). In this study, we produced two chimeric proteins, FH-Fc, containing the Fc region of immunoglobulin G (Fc) with SCR(6-7) or SCR(19-20). We found that both FH-Fc constructs killed B. burgdorferi in the presence of complement and reduced bacterial colonization and LD-associated joint inflammation in vivo. While SCR(6-7)-Fc displayed Lyme borreliae species-specific bacterial killing, SCR(19-20)-Fc versatilely eradicated all tested bacterial species/strains. This correlated with SCR(6-7)-Fc binding to select variants of CspA and CspZ, but SCR(19-20)-Fc binding to all tested OspE variants. Overall, we demonstrated the concept of using FH-Fc constructs to kill Lyme borreliae and defined underlying mechanisms, highlighting the potential of FH-Fc as a pre-exposure prophylaxis against LD infection. AUTHOR SUMMARYTransmitted by ticks, Lyme disease (LD) is the most common vector-borne disease in North America and has experienced an expanded geographical range and increasing number of cases in recent years. No effective prevention is currently available. The causative agent of LD, Borrelia burgdorferi sensu lato (Bbsl), is a complex containing a variety of species. To escape from killing by complement, one of the mammalian host defense mechanisms, Bbsl produces outer surface proteins that bind to a complement inhibitor, factor H (FH). These FH-binding proteins (i.e., CspA, CspZ, and OspE) evade complement by recruiting FH to the bacterial surface. Here we produced two FH-Fc fusion proteins, which combine human immunoglobulin Fc with the human FH domains that bind to Bbsl FH-binding proteins. We found that FH-Fc constructs kill Bbsl in vitro and prevent colonization and LD manifestations in murine models, correlating with these FH-Fc constructs ability to bind to CspA, CspZ, and OspE from respective Bbsl species. These results suggest the possibility of using FH-Fc as a prevention against LD.

microbiology↗

bioRxiv2024

Phosphorylation of KRP3, a KIP-related protein by MPK3 modulates rice tiller and seed number by fine-tuning cell division

During the cell cycle process, multiple inhibitors function as checkpoint regulators ensuring an impeccable duplication of genetic material. Kip-Related Proteins (KRPs) are a group of plant-specific cell cycle inhibitors and are known to regulate plant architecture and yield by differentially regulating cell division. KRPs, though functionally conserved, are dynamic in their amino acid composition. Interestingly, some KRPs are specific to dicots, and some are for monocots. In this study, we have identified that the presence of KRP3 and KRP6 is specific to the Poaceae family of monocots. An in-depth study of KRP3 showed a strict regulation of its expression in actively dividing cells during the G1-S phase progression of cell division. Our study identified KRP3 as a phosphorylation target of MPK3. The phosphorylation enhances KRP3 protein stability, which leads to strong inhibition of cell proliferation. By generating knock-out lines of krp3, mpk3 and double knock-out krp3mpk3, our study demonstrated that the MPK3-KRP3 module regulates rice root and shoot development as well as tiller and seed numbers. It was observed that KRP3 regulate the rate of cell division in a dose-dependent manner. The study, in a nutshell, establishes the role of KRP3 as an important regulator of rice vigor and yield.

plant biology↗

bioRxiv2024

Flame retardant tetrabromobisphenol A (TBBPA) disrupts histone acetylation during zebrafish maternal-to-zygotic transition

3,3,5.5-Tetrabromobisphenol A (TBBPA) is a widely used brominated flame-retardant utilized in the production of electronic devices and plastic paints. The objective of this study is to use zebrafish as a model and determine the effects of TBBPA exposure on early embryogenesis. We initiated TBBPA exposures (0, 10, 20 and 40M) at 0.75 h post fertilization (hpf) and monitored early developmental events such as cleavage, blastula and epiboly that encompass maternal-to-zygotic transition (MZT) and zygotic genome activation (ZGA). Our data revealed that TBBPA exposures induced onset of developmental delays by 3 hpf (blastula). By 5.5 hpf (epiboly), TBBPA-exposed (10-20 M) embryos showed concentration-dependent developmental lag by up to 3 stages or 100% mortality at 40 M. Embryos exposed to sublethal TBBPA concentrations from 0.75-6 hpf and raised in clean water to 120 hpf showed altered larval photomotor response (LPR), suggesting a compromised developmental health. To examine the genetic basis of TBBPA-induced delays, we conducted mRNA-sequencing on embryos exposed to 0 or 40 M TBBPA from 0.75 hpf to 2, 3.5 or 4.5 hpf. Read count data showed that while TBBPA exposures had no overall impacts on maternal or maternal-zygotic genes, collective read counts for zygotically activated genes were lower in TBBPA treatment at 4.5 hpf compared to time-matched controls, suggesting that TBBPA delays ZGA. Gene ontology assessments for both time- and stage-matched differentially expressed genes revealed TBBPA-induced inhibition of chromatin assembly- a process regulated by histone modifications. Since acetylation is the primary histone modification system operant during early ZGA, we immunostained embryos with an H3K27Ac antibody and demonstrated reduced acetylation in TBBPA-exposed embryos. Leveraging in silico molecular docking studies and in vitro assays, we also showed that TBBPA potentially binds to P300- a protein that catalyzes acetylation- and inhibits P300 activity. Finally, we co-exposed embryos to 20 M TBBPA and 50 M n-(4-chloro-3-trifluoromethyl-phenyl)-2-ethoxy-6-pentadecyl-benzamide (CTPB) -a histone acetyltransferase activator that promotes histone acetylation- and showed that TBBPA-CTPB co or pre-exposures significantly reversed TBBPA-only developmental delays, suggesting that TBBPA-induced phenotypes are indeed driven by repression of histone acetylation. Collectively, our work demonstrates that TBBPA disrupts ZGA and early developmental morphology, potentially by inhibiting histone acetylation. Future studies will focus on mechanisms of TBBPA-induced chromatin modifications.

pharmacology and toxicology↗

bioRxiv2023

Tick hemocytes have pleiotropic roles in microbial infection and arthropod fitness

Uncovering the complexity of systems in non-model organisms is critical for understanding arthropod immunology. Prior efforts have mostly focused on Dipteran insects, which only account for a subset of existing arthropod species in nature. Here, we describe immune cells or hemocytes from the clinically relevant tick Ixodes scapularis using bulk and single cell RNA sequencing combined with depletion via clodronate liposomes, RNA interference, Clustered Regularly Interspaced Short Palindromic Repeats activation (CRISPRa) and RNA-fluorescence in situ hybridization (FISH). We observe molecular alterations in hemocytes upon tick infestation of mammals and infection with either the Lyme disease spirochete Borrelia burgdorferi or the rickettsial agent Anaplasma phagocytophilum. We predict distinct hemocyte lineages and reveal clusters exhibiting defined signatures for immunity, metabolism, and proliferation during hematophagy. Furthermore, we perform a mechanistic characterization of two I. scapularis hemocyte markers: hemocytin and astakine. Depletion of phagocytic hemocytes affects hemocytin and astakine levels, which impacts blood feeding and molting behavior of ticks. Hemocytin specifically affects the c-Jun N-terminal kinase (JNK) signaling pathway, whereas astakine alters hemocyte proliferation in I. scapularis. Altogether, we uncover the heterogeneity and pleiotropic roles of hemocytes in ticks and provide a valuable resource for comparative biology in arthropods.

immunology↗

bioRxiv2022

Application of Temperature-Responsive HIS-Tag Fluorophores to Differential Scanning Fluorimetry Screening of Small Molecule Libraries

Differential scanning fluorimetry is a rapid and economical biophysical technique used to monitor perturbations to protein structure during a thermal gradient most often by detecting protein unfolding events through an environment-sensitive fluorophore. By employing an NTA-complexed fluorophore that is sensitive to nearby structural changes in 6xHIS-tagged protein, a robust and sensitive differential scanning fluorimetry (DSF) assay is established with the specificity of an affinity tag-based system. We developed, optimized, and miniaturized this histidine-tag DSF assay (HIS-DSF) into a 1536-well high-throughput biophysical platform using the Borrelial high temperature requirement A protease (BbHtrA) as a proof of concept for the workflow. A production run of the BbHtrA HIS-DSF assay showed a tight negative control group distribution of Tm values with an average coefficient of variation of 0.51% and median coefficient of variation of compound Tm of 0.26%. The HIS-DSF platform will provide an additional assay platform for future drug discovery campaigns with applications in buffer screening and optimization, target engagement screening, and other biophysical assay efforts.

biochemistry↗

bioRxiv2022

Co-infection of Ehrlichia with B. burgdorferi drives emergency myelopoiesis and promotes Lyme arthritis

Lyme disease is caused by the extracellular pathogen Borrelia burgdorferi (Bb), transmitted by the Ixodes scapularis tick. Approximately one-third of infected individuals develop arthritis of weight-bearing joints, though it is unclear why some patients develop arthritis and severe systemic disease while others do not. C57BL/6 (B6) mice are susceptible to Bb infection but do not develop arthritis, providing an in vivo model to evaluate mechanisms regulating development of Lyme arthritis. We demonstrate here that co-infection of B6 mice with the tick-borne pathogens Bb and Ehrlichia muris (Em) induced significant arthritis. Although co-infection did not impact bacterial burden or growth of either pathogen, the resultant Lyme arthritis in co-infected mice correlated with significant hematologic disturbances. Whereas single Bb infection elicited no overt hematologic changes, Em infection resulted in thrombocytopenia, lymphopenia, monocytosis, and granulocytosis, which was consistently observed in mice co-infected with both Bb and Em. Hematologic changes correlated with profound changes to the hematopoietic stem and progenitor cell (HSPC) populations in Em-infected mice. Most notable were dramatic reductions in populations of HSPCs committed to myeloid-biased differentiation. Co-infection resulted in persistent hematologic changes and bone marrow inflammation. Our data demonstrate for the first time that B6 mice, resistant to developing Lyme arthritis, exhibit severe joint pathology in the presence of a second pathogen, correlating with persistent emergency myelopoiesis. Our data support the conclusion that pathogen burden is not sufficient for disease and specific inflammatory signals and cells regulate the development of Lyme arthritis. ImportanceTick-borne illnesses, historically relegated to specific geographic areas, are increasing in prevalence and distribution. Borrelia burgdorferi causes Lyme disease, the most common tick-borne illness in North America, characterized by debilitating arthritis, carditis, and neurologic complications. It remains unclear why certain infected individuals develop severe disease while others are only mildly symptomatic. Human monocytic ehrlichiosis (HME) is another tick-borne disease that often results in profound illness and severe hematological disturbances. We show here that co-infection of B6 mice, resistant to Lyme arthritis, with Borrelia burgdorferi and Ehrlichia muris, used to model HME, results in the development of severe arthritis and emergency myelopoiesis. Our work suggests that immune activation driven by co-infection contributes to the development of Lyme arthritis.

immunology↗

bioRxiv2022

Function and evolution of aquaporin IsAQP1 in the Lyme disease vector Ixodes scapularis

Ticks are important vectors of pathogenic viruses, bacteria, and protozoans to humans, wildlife, and domestic animals. Due to their life cycle, ticks face significant challenges related to water homeostasis. When blood-feeding they must excrete water and ions, but when off-host (for stretches lasting several months) they must conserve water to avoid desiccation. Aquaporins (AQPs), a family of membrane-bound water channels, are key players in osmoregulation in many animals but remain poorly characterized in ticks. Here, we bioinformatically identified AQP-like genes from the deer tick Ixodes scapularis and used phylogenetic approaches to map the evolution of the aquaporin gene family in arthropods. We find that most arachnid AQP-like sequences (including those of I. scapularis) form a monophyletic group clustered within aquaglycerolporins (GLPs) from bacteria to vertebrates. This gene family is absent from insects, revealing divergent evolutionary paths for AQPs in different hematophagous arthropods. We next sequenced the full-length cDNA of I. scapularis aquaporin 1 (IsAQP1) and expressed it heterologously in Xenopus oocytes to functionally characterize its permeability to water and solutes. We additionally examined IsAQP1 expression across different life stages and adult female tissues. We found IsAQP1 to be an efficient water channel with salivary gland-specific expression, implying its function may be closely tied to osmoregulation during blood-feeding. Its functional properties were unique: unlike most GLPs, IsAQP1 has low glycerol permeability, and unlike most AQPs, it is insensitive to mercury. Together, our results suggest IsAQP1 plays an important role in tick water balance during blood feeding and may hold promise as a target for novel vector control efforts. Graphical TOC/Abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY eTOCIsAQP1, an aquaporin found in the black legged tick (Ixodes scapularis), is a functional water channel with low permeability to glycerol. In a phylogenetic analysis of 176 sequences from 48 arachnid species, most tick and other arachnid AQP-like genes formed a monophyletic clade nested within aquaglyceroporins (GLPs). IsAQP1 shows salivary-gland specific expression in adult females--with highest expression in unfed females--suggesting a role in water balance during blood feeding.

genetics↗

bioRxiv2022

Dynamics of Borrelia Burgdorferi Invasion and Intravasation in a Tissue Engineered Dermal Microvessel Model

Lyme disease is a tick-borne disease prevalent in North America, Europe, and Asia. Dissemination of vector-borne pathogens, such as Borrelia burgdorferi (Bb), results in infection of distant tissues and is the main contributor to poor outcomes. Despite the accumulated knowledge from epidemiological, in vitro, and in animal studies, the understanding of dissemination remains incomplete with several important knowledge gaps, especially related to invasion and intravasation at the site of a tick bite, which cannot be readily studied in animal models or humans. To elucidate the mechanistic details of these processes we developed a tissue-engineered human dermal microvessel model. Fluorescently-labeled Bb (B31 strain) were injected into the extracellular matrix (ECM) of the model to mimic tick inoculation. High resolution, confocal imaging was performed to visualize Bb migration in the ECM and intravasation into circulation. From analysis of migration paths we found no evidence to support adhesin-mediated interactions between Bb and components of the ECM or basement membrane, suggesting that collagen fibers serve as inert obstacles to migration. Transendothelial migration occurred at cell-cell junctions and was relatively fast, consistent with Bb swimming in ECM. In addition, we found that Bb alone can induce endothelium activation, resulting in increased immune cell adhesion but no changes in global or local permeability. Together these results provide new insight into the minimum requirements for dissemination of Bb at the site of a tick bite, and highlight how tissue-engineered models are complementary to animal models in visualizing dynamic processes associated with vector-borne pathogens. Significance StatementUsing a tissue-engineered human dermal microvessel model we reveal new insight into the invasion and intravasation of Borrelia burgdorferi (Bb), a causative agent of Lyme disease in North America, following a tick bite. These results show how tissue-engineered models enable imaging of dynamic processes that are challenging in animal models or human subjects.

bioengineering↗

bioRxiv2022

The G-protein-coupled estrogen receptor, a gene co-expressed with ERα in breast tumors, is regulated by estrogen-ERα signalling in ERα positive breast cancer cells.

PurposeThe purpose of this study was to assess the relationship between GPER, a seven transmembrane G-protein coupled estrogen receptor, and ER in breast tumors, and to make inroads into the mechanistic basis and clinical significance. MethodsTCGA-BRCA data was mined to examine the relationship between GPER and ER expression. GPER mRNA, and protein expression were analyzed in ER-positive or -negative breast tumors from two cohorts using immunohistochemistry, western blotting, or RT-qPCR. The Kaplan-Meier Plotter was employed for survival analysis. The influence of estrogen in vivo was studied by examining GPER expression levels in estrus or diestrus mouse mammary tissues, and the impact of 17{beta}-estradiol (E2) administration in juvenile or adult mice. The effect of E2, or propylpyrazoletriol (PPT, an ER agonist) stimulation on GPER expression was studied in MCF-7 and T47D cells, with or without tamoxifen or ER knockdown. ER-binding to the GPER locus was explored by analysing ChIP-seq data (ERP000380), in silico prediction of estrogen response elements, and chromatin immunoprecipitation assay. ResultsClinical data revealed significant positive association between GPER and ER expression in breast tumors. The median GPER expression in ER-positive tumors was significantly higher than ER-negative tumors. High GPER expression was significantly associated with longer overall survival of patients with ER-positive tumors. In vivo experiments showed a positive effect of E2 on GPER expression. E2 induced GPER expression in MCF-7 and T47D cells; an effect mimicked by PPT. Tamoxifen or ER-knockdown blocked the induction of GPER. Estrogen-mediated induction was associated with increased ER occupancy in the upstream region of GPER. ConclusionGPER expression is positively associated with ER in breast tumors, and a transcriptional target of the estrogen-ER signalling axis. More in-depth studies are required to establish the significance of GPER-ER co-expression, and their interplay in breast tumor development, progression, and treatment.

cancer biology↗

bioRxiv2022

CD36 homologs determine microbial resistance to the Lyme disease spirochete

Pattern recognition receptors sense pathogens in arthropods and mammals through distinct immune processes. Whether these molecules share a similar function and recognize the same microbe in evolutionarily distant species remain ill-defined. Here, we establish that the CD36 superfamily is required for Borrelia burgdorferi resistance in both the arthropod vector and humans. Using the blacklegged tick Ixodes scapularis and an electronic health record-linked biobank, we demonstrate that CD36 members elicit immunity to the Lyme disease spirochete. In ticks, the CD36-like protein Croquemort recognizes lipids and initiates the immune deficiency and jun N-terminal kinase pathways against B. burgdorferi. In humans, exome sequencing and clinical information reveal that individuals with CD36 loss-of-function variants have increased prevalence of Lyme disease. Altogether, we discovered a conserved mechanism of anti-bacterial immunity. One Sentence SummaryLipid receptors belonging to the CD36 superfamily exhibit a shared immune function in both ticks and humans.

immunology↗

bioRxiv2021

The Lyme Disease agent co-opts adiponectin receptor-mediated signaling in its arthropod vector

Adiponectin-mediated pathways contribute to mammalian homeostasis; however, little is known about adiponectin and adiponectin receptor signaling in arthropods. In this study, we demonstrate that Ixodes scapularis ticks have an adiponectin receptor-like protein (ISARL) but lack adiponectin - suggesting activation by alternative pathways. ISARL expression is significantly upregulated in the tick gut after Borrelia burgdorferi infection suggesting that ISARL-signaling may be co-opted by the Lyme disease agent. Consistent with this, RNA interference (RNAi)-mediated silencing of ISARL significantly reduced the B. burgdorferi burden in the tick. RNA-seq-based transcriptomics and RNAi assays demonstrate that ISARL-mediated phospholipid metabolism by phosphatidylserine synthase I is associated with B. burgdorferi survival. Furthermore, the tick complement C1q-like protein 3 interacts with ISARL, and B. burgdorferi facilitates this process. This study identifies a new tick metabolic pathway that is connected to the life cycle of the Lyme disease spirochete. Significance StatementAdiponectin binds to adiponectin receptors and participates in glucose and lipid metabolism in mammals. In this study, we found that ticks have an adiponectin receptor-like protein but lack adiponectin. Importantly, we demonstrated that the Lyme disease agent, Borrelia burgdorferi, takes advantage of the adiponectin receptor signaling pathway to establish infection in its arthropod vector, Ixodes scapularis. Our study sheds light on the understanding of Borrelia-tick interactions and provides insights into how a human infectious disease agent may evolve to manipulate host metabolism for its own benefits. Understanding this pathway may lead to new ways to interfere with the Borrelia life cycle, and this mechanism may be applicable to additional microbes that are transmitted by ticks, mosquitoes or other arthropods.

microbiology↗

bioRxiv2021

Essential Loop Dynamics Modulates Catalytic Activity in α-Chymotrypsin

Conformational dynamics of macromolecules including enzymes are essential for their function. The present work reports the role of essential dynamics in alpha-chymotrypsin (CHT) which correlates with its catalytic activity. Detailed optical spectroscopy and classical molecular dynamics (MD) simulation were used to study thermal stability, catalytic activity and dynamical flexibility of the enzyme. The study of the enzyme kinetics reveals an optimum catalytic efficiency at 308K. Polarization gated fluorescence anisotropy with 8-anilino-1-napthelene sulfonate (ANS) have indicated increasing flexibility of the enzyme with an increase in temperature. Examination of the structure of CHT reveal the presence of five loop regions (LRs) around the catalytic S1 pocket. MD simulations have indicated that flexibility increases concurrently with temperature which decreases beyond optimum temperature. Principal component analysis (PCA) of the eigenvectors manifests essential dynamics and gatekeeping role of the five LRs surrounding the catalytic pocket which controls the enzyme activity.

biophysics↗

bioRxiv2021

A Combined Spectroscopic and Molecular Modeling Study on Structure-Function-Dynamics under Chemical Modification: Alpha-Chymotrypsin with Formalin Preservative

Enzyme conformations can be altered via modification of its amino acid residues, side chains and large-scale domain modifications, which are closely linked to its function. Herein, we have addressed the role of residue modification in catalytic activity and molecular recognition of an enzyme alpha-chymotrypsin (CHT) in presence of covalent cross-linker formalin. Optical spectroscopy studies exhibit reduced catalytic activity of the enzyme with increased formalin concentration. Polarization gated anisotropy studies of a fluorophore 8-anilino-1- napthelenesulfonic acid (ANS) in CHT show a dip rise pattern in presence of formalin which is consistent with the generation of multiple ANS binding sites in the enzyme owing to modifications of its local amino acid residues. Molecular docking study on minimal local residue modifications in CHT reveals formation of a stable enzyme-substrate complex even with the serine-histidine cross-linked enzyme which prohibits product formation giving rise to reduced catalytic activity.

biophysics↗