bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genesis of Antibiotic Resistance (AR) XV: Genetic (Modification) Roulette impairs response to antibiotics – An Insidious AR Pandemic Phenomenon.
Authors: Kannan Subburaj , Alvarez Angelica , Hernandez Guadalupe , Hernandez Martin , Garza Jasmin , Carrizales Tania , Carrizales Diana , Crabtree Cynthia , Subburaj Prabavathy 
Published: November 2023

Molecular plant breeding techniques utilize antibiotic resistance genes (ARG) are used as a selection marker in the gene‐transfer construct for genetic modification (GM). It is known that the naked viral DNA promoter that regulate ARG, are hotspots for recombination with host DNA inducing AR via horizontal gene transfer (HGT). For example the npt II gene confers resistance to Neomycin, Kanamycin and other antibiotics, while aad‐3 gene confers resistance to antibiotics, Streptomycin and Spectinomycin (Goldstein et al., 2005). Clinical response to antibiotic therapy performed by Indiana State University and University of California San Diego found that hospitals observed decreased effectiveness of antibiotic drugs. Accordingly, the National Summary Data presented in Antibiotic Threats in U.S., by Friedan of CDC&P predicts that at least 2,049,442 illness and 23, 000 deaths due to AR. Clostridium difficile, Carbapenum resistant enterobacteriaceae, drug resistant Neisseria gonorrhoeae, Salmonella serotype typhi, multi‐drug resistant M. tuberculosis pose imminent threat to the public health. Genetic Roulette implies that increase in an untreatable infectious diseases is a multi factorial event inclusive of persistence of antibiotic r(resistance) genes, and resistome, (Davies and Davies 2010). Neither the Frieden's report on AR threat level (http://www.cdc.gov/drugresistance/pdf/ar‐threats‐2013‐508.pdf) from CDC & P, Atlanta, GA, United States) nor TATFAR report ( http://www.cdc.gov/drugresistance/pdf/tatfar‐progress_report_2014.pdf) nor National Action Plan For Combating Antibiotic‐Resistant Bacteria, (https://www.whitehouse.gov/sites/default/files/docs/national_action_plan_for_combating_antibotic‐resistant_bacteria.pdf) addresses this concern in light looming AR pandemic. Morbidity and mortality rate due to AR induced by nptII, aad‐3 and other antibiotics will be presented in EB2016.Support or Funding InformationProfessional development funds provided by SWTJC to S. Kannan for 2014–2015
Genesis Of Antibiotic Resistance VIII: “Auto Medication”, an universal practice That Contravenes Hardy‐ Weinberg ‐ Castle Principle (HWCP), Escalates as a Serious Risk Factor In Mitigating AR Pandemic.§
Authors: Kannan Subburaj , Carrizales Diana , Carrizales Tania , Alvarez Angelica , Jouregui Suzzette 
Published: November 2023

“Auto ‐ Medication” is a universal practice especially in low‐income, un insured, lack adequate health care(rural setting) facility, lack literacy on antibiotic therapy and antibiotic resistance. Based on the population dynamics, poverty level, health care provider per 1000 people, India has been chosen for this case study. A retrospective systematic review for auto‐medication of antibiotics and morbidity vs. mortality data on antibiotic resistance cases during 2013, and 2014 has been completed using the annual report of public health foundation of India. In essence, the public health report is void of the surveillance data on i. reported cases of antibiotic resistance among the emergency department visit, ii. Morbidity vs. mortality rate due to antibiotic resistance among the hospital acquired infections, and iii. Morbidity vs. mortality rate due to Clostridium diffcile infections. An effort to perform the country wide (India) data base analysis on the total number of cases in terms of morbidity vs. mortality rate due to antibiotic resistance is inconclusive. The reason for this inconsistency is due to the fact that there is no reporting mechanism on total number of cases due to antibiotic resistance to local authorities, state and national registry. Statistical analysis on incidence of antibiotic resistance among population of 24 states is incomplete. CDC (Frieden's) report on AR and United States presidential executive strategy to mitigate AR pandemic fail to account the role of immigrants from regions where the prevalence of unregulated antibiotic use with potential role in AR pandemic. Though “Auto Medication”, is a universal practice, global migratory pattern of the population from regions like India, and Mexico contravenes Hardy‐ Weinberg ‐ Castle Principle (HWCP), could pose as a serious risk factor for AR Pandemic.Support or Funding InformationSupported by Professional Development Funds.
Lactobacillus plantarum Improves Metabolic Outcomes and Alters the Colonic Immune State in Mice Fed a High Fat Diet
Authors: Heeney Dustin , Barouei Javad , Hsieh Yu‐hsin , Martinic Alice , Mishchuk Darya , Kieffer Dorothy , Slupsky Carolyn , Martin Roy , Marco Maria 
Published: November 2023

Obesity, type 2 diabetes mellitus, and metabolic syndrome are associated with a dysbiotic microbiome and unregulated immune activation. Thus, identifying dietary strategies enriching for members of the microbiota that correct for these deficiencies in the indigenous microbiota is useful for managing and preventing those chronic conditions. This approach holds particular promise for consumption of strains of Lactobacillus species commonly consumed in fermented foods. Herein, we investigated the effects of regular consumption of Lactobacillus plantarum (LP) on both treatment, and prevention of diet‐induced obesity. Study 1 Treatment: Male C57BL/6J mice received a high‐fat (HF) diet (45.9% energy from fat) for 9 weeks and then were maintained on either the HF diet or HF diet supplemented with 109 cells LP, feeding every other day (n=10/grp), for another 6 weeks. Study 2 Prevention: Male C57BL/6J mice were fed a HF diet with or without feeding 109 cells LP every other day, for nine weeks (n=10/grp). For both studies, ingestion of LP resulted in higher levels of Lactobacillus in the digestive tract but did not result in global alterations to the intestinal microbiome. In study 1, supplementation with LP significantly increased colonic production of the T‐cell proliferative factor IL‐2 (p = 0.010) and the lymphoid organ maintenance signal TNF‐beta (p = 0.0298). In study 2, supplementation with LP significantly increased colonic production of the T‐reg promoting cytokine IL‐27 (p = 0.0325). In both studies IL‐23, a driver of antibacterial and mucosal barrier maintenance, was significantly upregulated in LP supplemented mice compared to HF diet controls (p = 0.0056, 0.0457, for study 1 and 2 respectively). In Study 2, LP‐fed mice were partially protected from weight gain compared with the HF diet controls (p = 0.0310) and showed improved oral glucose tolerance. The reduction in weight gain was not associated with significant changes in food intake. Results from our study provide novel evidence of the capabilities of LP to inform the maintenance of the colonic immune system and that dietary supplementation with this strain can be beneficial even while maintaining a HF diet.Support or Funding InformationAmerican Diabetes Association
Mechanism and Interactions of Human Pyruvate Dehydrogenase Complex with its Kinase1
Authors: Guevara Elena L , Nemeria Natalia S , Yang Luying , Jordan Frank 
Published: November 2023

The human pyruvate dehydrogenase complex (PDC) consist of three principal catalytic components for its mission: E1, E2, and E3. The core of the complex is a strong subcomplex between E2 and an E3‐binding protein (E3BP). The PDC is subject to regulation at E1 by serine phosphorylation by four kinases (PDK1–4), and inactivation reversed by the action of two phosphatases (PDP1 and 2). Specifically, the regulation of PDC via phosphorylation of E1 by PDK1 is the focus of these studies, as PDK1 has been strongly implicated as an overexpressed culprit in multiple cancer cells; hence inhibition of the interaction of PDK1 with the E1 or E2•E3BP, could provide novel targets for the design of anticancer agents. Since the PDKs reside on (have a stronger binding to) the E2•E3BP core of the entire complex, specific inhibition of the PDK1‐E2•E3BP interaction provides a possible new approach to treatment of cancer. Inactivation of E1 via phosphorylation by PDK1‐4 was initially studied to determine PDK interaction and therefore activation by E2•E3BP or its C‐terminally truncated proteins. The studies suggest that the individual PDK isoforms require different degrees of activation: PDK2 appears to be the most active and not highly selective whereas PDK4 is the least active never fully inactivating E1. Recently, the interaction loci between PDK1 (PDK2) and the E2•E3BP core was defined by the Jordan group by two complementary methods, HD‐exchange MS (HDX‐MS) and NMR1. Interrogation of the sites on E2•E3BP identified to interact with PDK1 (PDK2) was accomplished by site‐directed mutagenesis studies in order to validate the identified ‘hot spots’. To date, there is evidence of the variants on the L1L2S region of E2 effecting inactivation of E1 via phosphorylation by PDK1. Further studies will be performed to identify the protein‐protein interaction of these L1L2S variants with PDK1 and PDK2 using HDX‐MS, isothermal titration calorimetry (ITC) and fluorescence titrations. These measurements will confirm whether the ‘hot spots’ interrogated indeed are important for the interaction of E2•E3BP with the PDK1, and whether it is specific to PDK1 by comparing such measurements with PDK2. These experiments will identify ‘real’ hot spots, against which rational drug design could be undertaken.Support or Funding InformationThis research was supported by the National Institutes of Health under Ruth L. Kirschstein National Research Service Award GM113601‐01A1 from the National Institute of General Medical Sciences.
Covalent inhibition of ERK docking interactions
Authors: Kaoud Tamer Saad , Johnson William H. , Piserchio Andrea , Ebelt Nancy D , Warthaka Mangalika , Cano Micael , Sammons Rachel , Wang Qiantao , Ren Pengyu , Ghose Ranajeet , Dalby Kevin N. 
Published: November 2023

The extracellular‐regulated kinase (ERK) signaling pathway has been implicated in the regulation of several cellular functions, including cell differentiation, proliferation, and inflammatory responses. Upon stimulation, ERKs have been reported to phosphorylate a large number of substrates localized either in the cytoplasm or the nucleus. Taking advantage of our understanding of ERK docking interactions, we tried to design/discover substrate‐selective inhibitors that target the protein‐binding site of ERK. Here we report the discovery of the first irreversible ERK inhibitor that targets its protein‐docking site. Protein NMR, mass spectrometry, mutagenesis and molecular docking studies showed that the compound forms a covalent bond with a conserved cysteine residue C‐159. Extensive biochemical studies have been employed to estimate its kinetic parameters and its kinase‐selectivity profile. Finally, the new ERK inhibitor was shown to inhibit both ERK phosphorylation and its ability to phosphorylate downstream substrates (e.g. p90RSK and Elk‐1) in HEK293 cells. The inhibitor showed dose‐dependent inhibitory effects on HEK293 and melanoma A375 cell proliferation and viability. Targeting ERK in cells was confirmed using a chemical‐genetic approach where the C159A mutant was used to rescue the effects of this compound on ERK signaling. The discovered compound represents a new class of ERK inhibitor that uniqely labels its protein binding site. Further development may result in a pharmacologically useful inhibitor.Support or Funding InformationThis research has been supported by the following grants from the National Institutes of Health: GM059802 (to K.N.D.),GM084278 (to R.G.), and G12 MD007603 (towards partial support of the NMR facilities at The City College of New York). R.G. and K.D. are members of the New York Structural Biology Center, NYSTAR facility. T.S.K. acknowledges Cancer Prevention and Research Institute of Texas Postdoctoral Training Award (RP140108).
Phosphorylation of PKA Regulatory Subunit RIα by PKG: a Mechanism for “Desensitizing” the Type I PKA Holoenzyme
Authors: Haushalter Kristofer J. , Casteel Darren E. , Stefan Eduard , Taylor Susan S. 
Published: November 2023

The catalytic subunit of protein kinase A (PKA‐C) is a critical signaling protein in eukaryotes, requiring a high degree of regulation to control its phosphorylation activity under both homeostatic and stress conditions in the cell. The inhibitory mechanism for the four functionally non‐redundant regulatory subunits of PKA (RIα, RIβ, RIIα and RIIβ) has been well characterized with regard to their ability to keep PKA‐C in an inactive “holoenzyme” complex until activation with cAMP. However, recent evidence suggests that unique features within Type I PKA holoenzymes may be alternatively regulated via non‐canonical signaling mechanisms, such as regulatory subunit phosphorylation and oxidation. Previous work demonstrated that serine 101 of the RIα regulatory subunit can be phosphorylated by cGMP‐dependent protein kinase (PKG) in vitro, whereupon phospho‐RIα no longer inhibited PKA‐C at normal (1:1) stoichiometric ratios. In this study, we validated this phosphorylation mechanism under in vivo conditions to determine whether this phosphorylation event could lead to a physiologically relevant “desensitization” of PKA‐C in the context of cellular stress. Here, we show that PKG is indeed able to incorporate phosphate onto S101 of RIα under both in vitro and in vivo conditions, and moreover that phosphorylation does not occur upon RIα where Ser101 is substituted with Ala (i.e. S101A). The PKG site is not accessible in the holoenzyme prior to activation by cAMP which thus creates an ordered pathway for crosstalk between PKA and PKG. Using a luciferase protein complementation assay, we show that the phospho‐mimetic mutation (i.e. S101E) decreases holoenzyme formation in cells, and this is accompanied by increased downstream activation of PKA‐C. Because PKG signaling can be activated by ROS (i.e. nitric oxide) in cells, ongoing experiments are aimed toward determining if this phosphorylation mechanism may serve as an alternate pathway for “priming” the PKA holoenzyme for activation within the context of oxidative stress. Taken together, these findings support the proposed mechanism for Type I holoenzyme “desensitization” as a potential in vivo pathway for establishing signaling crosstalk between PKA and PKG signaling.Support or Funding InformationSupport for this work was funded by NIH GM34921 to sst.
Tracking Dynamics of Vascular Development in Time and Space using Sox18 Dimer split‐Fluorescent Protein Biosensors
Authors: Giles Nichole , McCann Alex , Moustaqil Mehdi , Overman Jeroen , Fontaine Franck , Francois Mathias , Sierecki Emma , Gambin Yann 
Published: November 2023

The temporal and milieu‐specific control of cell fate is dependent on the fine‐tuning of the activity of transcription factors. One way this is achieved is by varying the composition of specific protein complexes that binds cooperatively at DNA enhancer regions. SOX18 is a key transcription factor that is transiently expressed in nascent endothelial cells during embryogenesis and adult neovascularization. Mutations of human SOX18 cause Hypotrichosis‐Lymphedema‐Telangietasia and the protein is overexpressed in gastric and pancreatic cancer cell lines. Recently, we showed that SOX18 could act as a heterodimer and more importantly demonstrated that SOX18 can homodimerize in vitro. This is further supported by the identification of DNA binding sites specific to the homodimer and subsets of gene under the control of the homodimer only. These observations suggest that SOX18 heterodimer and homodimer may play different roles during vascular development.Here, we developed a SOX18 dimer split‐fluorescent protein (FP) biosensor to track the dynamics of SOX18 dimerization and its influence on different gene expression during vascular development in zebrafish embryo.However, the major obstacle to the general use of these biosensors, especially in living organisms, is the amount of optimisation required, as the complementation depends on multiple parameters: the nature of the fluorescent protein, the relative position of the two parts, the length and flexibility of the linker regions… To shorten the process, the initial screening and optimization of split‐FP biosensors with emission ranging from blue (BFP) to red (mCherry) was realized using a cell‐free expression platform.Once optimised, the SOX18 dimer biosensor was expressed in an endothelial cell line (HUVEC) for validation and imaging optimization. The sensor was then adapted to a zebrafish embryogenesis model where expression is under the control of endothelial specific fli1 promoter.The study will be extended to the creation of reporters for different heterodimers of SOX18 to identify the partners driving the different steps of vasculature development.Support or Funding InformationEMBL Australia Single Molecule Centre Sydney Node
Super‐resolution Imaging of the HER Family of Receptor Tyrosine Kinases
Authors: van Lengerich Bettina , Puchner Elias , Huang Bo , Jura Natalia 
Published: November 2023

The human epidermal growth factor receptor (HER/ErbB) family of receptor tyrosine kinases encompasses transmembrane signaling proteins important for cellular growth, differentiation, and survival. The four members of this family (EGFR, HER2, HER4 and the HER3 pseudokinase receptor) are able to bind to numerous different growth factor ligands, leading to receptor homo and hetero‐oligomerization and subsequent activation. This combinatorial potential gives rise to diverse signaling outputs, the extent of which is still not fully understood due to the lack of precise knowledge of how HER receptors interact with each other upon ligand binding. We seek to understand HER receptor interactions at the membrane by directly imaging the receptor molecules at a single molecule level. We use a technique that can accurately count absolute numbers of molecules at the membrane by employing a correction for fluorophore blinking in conjunction with stochastic optical reconstruction microscopy (STORM). This reveals the spatial organization and extent of oligomerization of receptors with sub‐diffraction limit resolution. We then probe how this organization and oligomerization evolve as a function of growth factor ligand stimulation. We also examine the effects of mutations that disrupt key interfaces in active HER receptor dimers, as well as of activating HER receptor mutations that are prevalent in cancer. Finally, we determine the role that over‐expression of a partner co‐receptor has on the extent of receptor oligomerization, and how ligand stimulation influences these interactions. This work aims to contribute to the fundamental understanding of the activation mechanism of HER receptors and to help advance development of new therapeutics targeting aberrant cross‐talk among HER receptors in human disease.Support or Funding InformationThis work is funded by an American Cancer Society post‐doctoral fellowship to BvL.
Phenotype analysis and gene expression profiling in a yeast model of Cystinosis
Authors: Walnofer April E. , Vitiello Seasson 
Published: November 2023

CTNS is a gene that encodes the protein Cystinosin, a lysosomal cystine transporter. When CTNS is nonfunctional, it results in the disease cystinosis, which has a hallmark sign of cystine accumulation in the lysosome and cell death. CTNS has an orthologous gene in Saccharomyces cerevisiae, ERS1. When ERS1 is deleted, ers1‐Δ cells do not accumulate cystine. It has been shown that sixteen modifier genes in S. cerevisiae are differentially expressed between parental ERS1+ and ers1‐Δ, which may contribute to compensation of the lack of cystine accumulation. Gene expression profiling and phenotype analysis of ERS1 has been done based on characterization of the sixteen modifier genes. This includes looking at respiration, redox regulation, genetic interactions of modifier genes with ERS1, and sulfur homeostasis.Support or Funding InformationInstitutional Development Award (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health [P20GM103443] and the Cystinosis Research Foundation.
Three‐Dimensional Trunk Kinematics of Humans and Chimpanzees: New Insights on Early Hominin Bipedalism
Authors: Thompson Nathan E , Demes Brigitte , O'Neill Matthew C 
Published: November 2023

The human trunk is involved in a variety of locomotor tasks that promote stability and efficiency during locomotion. The trunk and upper body must create a stable base for the head, yet must also move enough to cancel out angular momentum of the swinging lower limbs and to help shift the center of mass (CoM) laterally towards the stance‐side foot. These abilities are likely achieved in part by the derived morphology of the human trunk. Humans possess a tall and narrow trunk compared to the more squat trunk morphology of our closest living relatives (great apes).Because of some similarities between early hominins and chimpanzees in trunk morphology (thoracic shape in Australopithecus) and lower limb features that may affect trunk motion (potentially wide step widths in Ardipithecus), the role of the upper body during locomotion in our early ancestors remains unclear. Here, we investigated three‐dimensional trunk motion in chimpanzees and humans in order to determine the effects of overall trunk morphology on upper body stability during bipedal locomotion.The trunk was divided into three anatomical segments (thorax, lumbar region, and pelvis). In the transverse plane, all chimpanzee trunk segments rotated in the same direction at the same time (in phase), whereas in humans, the thorax rotated out of phase with the pelvis. However, relative to the pelvis, both humans and chimpanzees achieved the same amount of thoracic (17.1° and 19.1° respectively, n.s.) and lumbar rotations (6.1° and 6.5°, n.s.). In the frontal plane, chimpanzees laterally flexed (listed) all three trunk segments (13–22°) towards the support side limb, whereas in humans only the thorax listed (3–4°) towards the support side. The greater thoracic listing in chimpanzees may be necessary to compensate for their wider step width, absent a valgus knee. Indeed, in chimpanzees, the angular excursion of the thorax (beyond that which is due to list of the pelvis alone) is correlated with increased lateral displacement of the CoM (P<0.01), which is in turn correlated with increased mediolateral mechanical work (P<0.001).The results of this research suggest that some important aspects of bipedal locomotion were already present in the trunks of the last common ancestor (LCA) of humans and chimpanzees. Regardless of the precise reconstruction of the trunk, the LCA was likely already capable of using its thorax to compensate for some degree of rotation of the pelvis. However, if early hominins utilized wide step widths (as has been suggested for Ardipithecus ramidus), there would have likely been more active listing of the thorax in order to shift the CoM over the support side foot. This, in turn, would have likely been accompanied by an increase in mediolateral mechanical work.Support or Funding InformationThis research was funded by the NSF BSC 0935321 and The Leakey Foundation.