bluuebunny/crossref_metadata_2025 huggingface dataset:
A New Curricular Schema for Allopathic Medical Schools in the United States
Authors: Taylor Melissa
Published: June 2021

Medical education in the United States has been changing dramatically in the last few decades. Medical schools are moving away from a traditional, didactic, and discipline‐based curriculum, to a more integrative, active, and student‐centered curriculum. Medical schools have revised their curricula for many reasons, and one of the more common reasons is to meet current Liaison Committee on Medical Education (LCME) standards. Additionally, medical schools have integrated their curricula for the same, as well as other, reasons. There is no one singular way that a medical school can revise and integrate its curriculum. This study examines the types of curricula offered at the 150 allopathic medical schools in the United States. A search of medical curricula, as per the medical schools' websites, was conducted. This study utilized content analysis to tabulate frequencies of common key words from the medical schools' websites. General key words, such as “integration”, “organ systems”, “discipline”, and “block”, were reviewed first. A more specific list of key words, such as “horizontal integration”, “integration of basic and clinical science”, and “organ systems‐based blocks” was then generated. A table was created with these key words and tallied for all the medical schools studied. Twenty‐five different classifications of medical curricula were found, including thirteen different ways to integrate a curriculum. The most common form of integration was an “integration of basic and clinical sciences” through “topic‐based courses.” These courses are not the traditional discipline‐specific courses, but also are not fully based around organ systems units. This study shows that a wide variety of curricular schema can be utilized to organize a modern‐day medical curriculum.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Challenge of Brains: A Ludic and Amusing Game Useful to Review Neuroanatomy for Undergraduate Students
Authors: Rueff‐Barroso Carlos R. , Veiga Luana Cordeiro , Ronconi Karoline Souza , Cerqueira Gilberto Santos , Fazan Valéria Paula Sassoli
Published: June 2021

Teaching of human gross anatomy must be always under reflections, adaptations and modifications that should follow social changes in behaviors. The youth of the 21st century has changed rapidly and is no longer comparable with the last century students. The task of teaching these young students also needs to undergo conceptual and practical changes in order follow the need for more interactive, dynamic and attractive ways to study anatomy. We aimed to create, develop and test a board game that provides a full review of the neuroanatomy subject for physical therapy students, in a way to stimulate studying and help to sediment subject's knowledge, also dealing with difficulties and doubts. We created a board game and 354 cards with questions and answers that include the entire neuroanatomy contend of the semester. Extra cards with bonus or penalties to players were also included. The game was played by teams that were separated by colors. After participating, students answered to a quiz related to their impressions about the game. The quiz showed that: the game was useful for self‐assessment (90.6%), colors made the activity more fun (100%), the game was fun to play (96.9%), students would like to have more classes like this (90.6%), the game increased the stimulus to study for the test (96.9%), students found the method effective as a review class (96.9%), students felt motivated to study neuroanatomy after playing the game (87,5%), and the game stimulated interaction between the students (90.6%) and student/professor (93.8%). The adjectives used more often for the game were amusing (14.3%), dynamic (13.2%), interesting (13.2%) and stimulant (11%). We observed that 50% of the students increased their grades after playing the game. The activity proved to be very effective in stimulating and deepening the students' neuroanatomy knowledge, with a large percentage of approval and improvement of students' grades.

Support or Funding Information

CNPq

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Research Trends as Seen Through the Ten Year Publication History of the Journal Anatomical Sciences Education
Authors: Thompson andrew R. , Husmann Polly R.
Published: June 2021

2018 marks the ten year anniversary of Anatomical Sciences Education (ASE), which has become one of the premier journals in the field of scientific educational research. ASE provides a home for educational research in the areas of gross anatomy, embryology, histology, and neuroanatomy. While these sub‐disciplines of anatomy have been taught for centuries, methods of their instruction have transformed drastically over the past several decades. These changes have spurred an increased interest in anatomy education research, which was a driving factor in the creation of ASE. Thus, ASE provides a window into trends involving both pedagogy in the field of anatomy and development of anatomy education research as a distinct field. The goal of this paper is to evaluate these areas by analyzing publications over the ten year history of ASE. Results show that while all sub‐disciplines are represented, gross anatomy has dominated with over 85% of the publications focusing on this area. Perhaps not surprisingly, approximately 70% of the research involves pre‐clinical medical students, a trend that has remained consistent over the past ten years. There has, however, been a statistically significant (p<0.05) change in the type of data used in these projects. Earlier research was largely descriptive in nature and utilized either quantitative or qualitative data whereas mixed method approaches were more common in recent years. These and several other areas related to anatomy education research will be explored and discussed in the context of the broader field of educational research.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Creating Instruction to Optimize Learning of the Anatomical Sciences
Authors: Lisk Kristina , Agur Anne , Woods Nicole N.
Published: June 2021

Integration of basic and clinical science knowledge is increasingly being recognized as important for practice in the health professions. The concept of ‘cognitive integration’ places emphasis on role of basic science in providing critical connections to clinical signs and symptoms while accounting for the fact that clinicians may not explicitly articulate their use of basic science knowledge in clinical reasoning. In this study, we aimed to extend previous work on cognitive integration using new learning materials teaching musculoskeletal pathologies with allied health students. In addition, we aimed to further our understanding of cognitive integration and conceptual coherence by using a diagnostic justification task to investigate the impact of integrated basic science instruction on novices' diagnostic reasoning process. Participants (N = 43) were randomly assigned to a integrated basic science (BaSci) or clinical science (CS) training group. The BaSci group was taught the clinical features along with the underlying causal mechanisms of four musculoskeletal pathologies while the CS group was taught only the clinical features. To equalize the learning time between the two conditions, the CS group was taught epidemiology and potential treatment options for each of the pathologies. Participants completed a diagnostic accuracy and memory test immediately after learning and one‐week later. A diagnostic justification test was also completed one‐week after initial learning. Novices who learned the integrated causal mechanisms had superior diagnostic accuracy (p<0.01) and a better understanding of the relative importance of key clinical features (p<0.01). Although participants from both groups identified correct features on the justification test, those in the BaSci group identified key diagnostic features rather than features that were common across disease categories. Participants in the BaSci group also outperformed the CS group on the basic memory test; however, this difference was no longer evident one‐week later. This study demonstrates the positive impact of integrating basic anatomical education and clinical science instruction on students' diagnostic reasoning ability in addition to diagnostic accuracy. These findings further our understanding of conceptual coherence by providing explicit evidence of the advantage learners have when basic science knowledge is cognitively integrated.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
A Comparison Of Learning with Classical Lecture and Demonstration Methods Versus Individually and Collaboratively Prepared Concept Maps
Authors: ANAND MAHINDRA KUMAR
Published: June 2021

Introduction

The use of concept maps as teaching‐learning tool facilitates knowledge integration and development of meaningful learning. It was developed by Novak and Gowin and is based on Ausubel's theory of meaningful learning. Present study was undertaken to compare the learning with concept maps and classical lecture‐demonstration methods in gross anatomy and to obtain feedback of students and faculty regarding the same.

Methods

A randomized cross sectional prospective study was conducted on 150 medical students of 1st year. Students were divided into 3 groups A, B and C. Group A studied “gross anatomy of foot” by developing concept maps in groups.. Group C learnt through lecture and demonstration method. Group B studied with individually prepared concept maps. Pre and posttest were conducted for all groups with pre validated questionnaire. Feedback was obtained from both students and faculty through a 5 point Likert scale. Anova and Student's T test was applied for statistical analysis.

Result

There was significant improvement in knowledge in group A and B students in comparison to group C students. This improvement was more marked in regards to high cognitive type assessment questions. 90% students found that development of concept maps helped them to understand topic better but it was time consuming.

Discussion and Conclusion

Concept mapping is a better learning tool in comparison to classical lecture and demonstration method. Preparation of concept maps in groups help in development of better understanding and learning of the subject than learning with classical lecture‐demonstration method and individually prepared concept maps.

Support or Funding Information

NIL

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Assessing if an Physical and Interactive Anatomy and Physiology Model Enhances and Enriches Student Learning
Authors: Llewelyn KaCee , Seed Brian Vaughn
Published: June 2021

The purpose of this study is to test a teaching model that utilizes multiple styles and forms of learning, and determining if this physical and interactive model increases learning and comprehensive understanding of the muscular and skeletal systems.

Many human anatomy and physiology teaching models on the market today fail to capture how the muscular and skeletal systems work in conjunction with one another. Students have resorted to simply learning the different muscle groups and bone prominences. They fail to understand origin and insertion points and what this means for the human body on a functional level. The model that we have designed allows students to see bone prominences, origin and insertion points, ligaments, tendons, deep muscles, and superficial muscles. It will also show the students the nerves that innervate these muscles. This physical model will allow students to experience hands on learning as they move the model itself, visual experience as they see the muscles move, and auditory experience as the educator teaches the material using the model. This model will be tested to see if students gain a comprehensive understanding on how the human body moves.

It will be tested in various classrooms to determine if it will aide educators in using many forms and styles of learning simultaneously, so that each student can benefit with their own style. It will be used to teach the muscular and skeletal system to participants between the ages of eleven and twenty‐two; ranging from middle school to entry level undergraduate. The participants will be given a survey which will be administered before and after the model is used to determine the model's ability to teach the muscular and skeletal systems, functional movement, overall anatomy comprehension, participants motivation and desire to learn more. This model will be tested in January 2018, and results from the surveys will be analyzed to assess whether the model was successful in aiding comprehension and increasing enthusiasm for anatomy and biomedical learning.

Support or Funding Information

Biomedical Learning and Student Training, Part of the Diversity Consortium of the National Institute of Health

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Examination of foramen magnum position in relationship to the degree of bipedalism in mammals
Authors: Abati Jena Michelle , Hechtel Laura
Published: June 2021

A more inferior and anterior positioning of the foramen magnum has been found in bipedal mammals, such as humans while quadrupedal mammals are known to have a more superior and posterior positioning. However, it is not known if the degree to which this positioning varies is related to the degree of bipedal stance taken by quadrupeds. In this study, it was predicted that mammal species that can assume a more bipedal stance and often lift their head and trunk in order to more effectively feed, forage, or protect themselves, are likely to have a more inferior and anterior positioning of the foramen magnum when compared to completely quadrupedal mammalian species. In this study, foramen magnum position was compared in various mammalian skulls provided by the Cornell University Museum of the Vertebrates. Measurements of the foramen magnum in relationship to other cranial structures were taken including, the nuchal crest to the superior edge and inferior edge of the foramen magnum, the nuchal crest to the base of the occipital condyles, and the superior edge of the palatine to the inferior edge of the foramen magnum. Results indicate a significant difference in foramen magnum positioning between completely quadrupedal mammals (e.g. Canis latrans), and those mammals capable of taking a bipedal stance on occasion (e.g. Ursus americanus and Sciurus caroliensis). Therefore, the positioning of the foramen magnum is most likely a continuum and reflects the degree to which a mammal is likely to utilize a bipedal stance for feeding or protection from predation.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Signals from the Gut Influence Mouse Behavior: Examination of Mechanisms
Authors: Peterson Diana Coomes
Published: June 2021

The intestinal microflora can have significant influences on brain activity. Gut microbiota release neurotransmitters that are hypothesized to send signals via the vagal nerve back to the brain. However, such afferent interactions have never been proven. The current experiments utilized optogenetic vectors (AAV‐CaMKIIa‐eNpHR3.0‐EYFP) placed within the jejunum to label vagal nerves. These vectors induce the insertion of light sensitive chloride channels into neuronal membranes that when stimulated with light deactivate the neuron. By placing these vectors in the jejunum, the experiment selectively deactivated only the select fibers that run from the jejunum to the brain. Control animals were injected with saline.

In all animals, normal animal behavior was recorded for anxiety measures (elevated plus maze), open field behaviors, and forced swim tests. Each test was performed with both light deactivation of the vagus, and no‐light conditions. Results indicated that mouse behavior was not affected for the control animals nor for animals in the no‐light vector conditions. However, the animal behavior was dramatically altered for each animal in the optogenetic light deactivation condition. In each behavioral test, animals showed a freezing behavior for the extent of time that the vagus was deactivated. Within the forced swim experiments, frozen animals did not keep their head above water and tended to sink to the bottom of the tank (requiring multiple rescues during each session).

The results support the hypothesis that the jejunum can influence brain activity via vagal nerve efferents. I conclude that optogenetic deactivation of the vagal afferents of the jejunum has a direct influence on animal behavior and thus brain circuits. Therefore, this pathway provides an optimal mechanism by which future nutritional therapies can be devised for a variety of brain pathologies.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
A Comparison of Epinephrine, Esmolol, and the Combination of Both in Reperfusion Injury After Murine Myocardial Ischemia
Authors: Blaskowsky Justin , Eckle Tobias
Published: June 2021

Current research suggests that epinephrine, when used during cardiac arrest, pulseless electrical activity, or ventricular fibrillation, may contribute to reperfusion injury. Esmolol, a short acting cardioselective beta‐blocker, has recently been shown to help mitigate this effect. The objective of this study is to demonstrate the possible cardio‐protective effects esmolol has on murine myocardium. The study was done using 24 C57/BL6 six‐month‐old mice randomized into four experimental groups. Ischemia was induced using the hanging weight system described by Eckle et. al. The mice were exposed to ischemia for one hour and subsequently reperfused with either saline, epinephrine, esmolol, or a combination of epinephrine and esmolol for two hours. After the reperfusion period, blood was drawn to measure troponin levels, and staining with Evans Blue (EB) was performed on the murine hearts. Sectioning and subsequent staining with triphenyltetrazolium chloride (TTC) was done to provide contrast against the EB stain. Infarct size was defined as the ratio of ischemic viable versus non‐viable tissue. The infarct size for each heart was calculated with ImageJ, and later MATLAB was used to provide an automated analysis of infarct area. A full murine heart model was then generated using MATLAB to visually demonstrate differences in infarct size.

Preliminary results have indicated a statistically significant reduction of infarct size in the esmolol group when compared to epinephrine, and ongoing analysis shows similar results. The future goals of this study are to provide additional evidence of esmolol as a cardio‐protective drug, and to demonstrate a simple computerized way to identify myocardial infarct area in an animal model.

Support or Funding Information

Research was supported through the University of Colorado Anschutz Medical Campus and the Master of Science in Modern Human Anatomy Program.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Midline Episiotomy May Pose Less Risk of Nerve Damage Than Mediolateral Episiotomy: A Cadaveric Anatomical Study
Authors: Patel Akash , Hung Jun , Castro Monica , Plochocki Jeffrey H. , Hall Margaret I.
Published: June 2021

Episiotomy, the surgical incision of the perineum to assist during vaginal delivery, can be classified into two main approaches, midline and mediolateral. While use of surgical episiotomies is in a decline, clinicians throughout the world continue to commonly utilize episiotomy. However, current literature provides insufficient anatomic evidence as to which of the two approaches is preferable. Many clinicians utilize a mediolateral approach, arguing that this incision ensures the safety of the external anal sphincter and therefore will not endanger maternal fecal continence postpartum. Other clinicians prefer a midline approach, arguing that incising the perineal body will endanger fewer anatomical structures. In this study, we utilize human cadaveric dissection to identify and understand which anatomical structures are damaged through the use of the two episiotomy approaches to justify a preferential episiotomy procedure. We performed a midline and one of four mediolateral episiotomy incisions using blunt scissors on 15 female cadavers used for medical education at Midwestern University. The posterior minor labia in the midline provided a zero‐degree axis from which angles of incision were calculated to either 60, 45, 30, or 20 degrees from midline using digital calipers and a protractor. After incision, dissection revealed structure damage. All dissections revealed that mediolateral episiotomy damaged the ipsilateral bulb of the vestibule and the overlying bulbospongiosus muscle. The superficial perineal nerve was damaged in 20% of the cadavers, and the anterior labial nerve was damaged in 13% of the cadavers. However, we suspect that during birth when the region is engorged with blood and the vaginal orifice pulled inferiorly and anteriorly, mediolateral episiotomy could result in nerve damage more often than in cadaveric dissection. Based on these results, we suggest that a midline approach might be preferable. Possible denervation to perineal structure may outweigh the potential risks to the external anal sphincter, which would likely heal within weeks postpartum with its nerve supply intact.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Characterization of the roles of connexin 32 in zebrafish vascular development
Authors: Chen Yi‐Ting
Published: June 2021

The process of vascular development in vertebrates are mainly divided into vasculogenesis and angiogenesis. We previously identified the transcription factors Islet2 (isl2) and Nr2f1b required for the growth of intersegmental vessels (ISV) and caudal vein plexus (CVP) in zebrafish. Microarray analysis showed Connexin 32 is regulated by Isl2/Nr2f1b. Gap junction protein Connexins32 (Cx32) has been shown functions on the blood vessels in injury mice and enhances angiogenesis in cell‐based study, however, there is no reference on the vascular development in animal study. In addition, signal pathways related to cx32 and angiogenesis have not yet been clarified. In this study, we first showed cx32 mRNA is expressed is expressed in developing vessels from 18S to 30 hpf stages, suggesting its roles in vasculature. Knockdown of cx32 by morpholino injection causes vascular defects in ISV and CVP, suggesting the role of cx32 in vascular growth. TUNEL assay showed that vascular defects do not caused by cell death, but likely due to the impairment of endothelial cell proliferation and migration. Consistent with vascular growth defects, loss of cx32 affects the expression of the vascular markers flt4, mrc1, flk, stabilin, and ephrinb2. Furthermore, the overexpression of cx32 does not impair the vascular growth, but rescue the cx32 morphant, suggesting the specificity of morpholino knockdown. Finally, we examine the interaction between cx32 and multiple signals and observed that cx32 is regulated by VEGF signals for ISV growth and BMP signals for CVP patterning.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Using Electric Cell‐substrate Impedance Sensing (ECIS) Technology as a Tool for Mathematical Analysis of the Effects of Glucose on Bone‐like Osteosarcoma Cells (UMR 106‐01 BSP)
Authors: Choi Jeannie , Hernandez Bryan , Joseph Eugene
Published: June 2021

Mathematical modeling has been useful in studies of the progression of type II diabetes. However, not many studies have been done using electric cell‐substrate impedance sensing (ECIS) technology to validate mathematical modeling in bone cells exposed to high concentrations of glucose. In this study we hypothesized that ECIS technology can be used as a predictive tool to evaluate the effects of glucose on cell membrane impedance in UMR cell cultures by mathematical analysis through predictive mathematical modeling. Briefly, UMR cells were plated at 2.5 × 105 cells/mL in eight‐well (8W) ECIS array plates. Designated wells were treated with different concentrations of glucose (10, 5, and 3 mM). Cell cultures were incubated in a humidified chamber at 37° C with infusion of 5% CO2. Membrane resistance, time of attachment and rate of spreading of the cells were monitored and measured in vitro, using the ECIS machine. Based on the data generated by the ECIS machine, a mathematical formula was derived to predict the resistance of the UMR cell membrane for any concentration of glucose used. The formula was tested based on the generated data. F test ANOVA was employed to compare variances between all experimental trials; all values in the results are expressed as means ±SD. The results show that the highest concentration of glucose caused a decreased time of attachment, rate of spreading and an increased cell membrane resistance in the UMR cells. The following formulas were derived based on the ECIS data: dR/dt=kR0ekt and R[G]=R0e+sG (where dR/dt=rate of spreading, k=rate constant, R0=initial resistance, t=time, [G]=glucose concentration, and s=rate constant (1/[glucose]). The formulas were found to be of predictive value. The results suggest that the ECIS technology is a potentially useful tool in the mathematical analysis of physiological phenomena in bone‐like cells in vitro.

Support or Funding Information

This work was supported by the Ryckman Endowment Student Research Fund of the Biology Department at La Sierra University in Riverside, CA.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Calcium and lipid exchange machinery in Sertoli cells
Authors: Adams Arlo , Yoo David , Vogl A. Wayne
Published: June 2021

Sperm production (spermatogenesis) takes place in the seminiferous tubules of the testis. Specialized junction complexes termed ‘ectoplasmic specializations' (ESs) in the seminiferous epithelium have several key functions. ESs between Sertoli cells disassemble during the movement of early spermatogenic cells towards the apex of the epithelium and those ESs between a Sertoli cell and late stage spermatids disassemble during sperm release. There is evidence that structures called ‘Tubulobulbar Complexes' (TBCs) are endocytic structures responsible for the internalization and eventual degradation of junctions associated with ESs, allowing for early spermatocyte translocation from the basal epithelium and release of late spermatids from the apical epithelium. One key feature of both ESs and TBCs is their extensive association with peripheral ER. ESs are tripartite in structure consisting of Sertoli cell plasma membrane (PM), a layer of actin bundles, and an overlying leaflet of ER. TBCs are double membrane invaginations that appear as ESs are beginning to break down. They are long, endocytic in‐growths, with a swelled bulb region that contains an extensive ER‐PM contact site. TBC bulbs eventually ‘bud’, fuse, and become endosomes. Little is understood about the function of the ER at both ESs and TBCs, though it is well established that ER contact sites in other cells function in both calcium signalling and lipid exchange. If ER in ESs and ER‐TBC contacts function in calcium and lipid exchange, then known calcium signalling machinery and lipid exchange proteins should be localized to these structures. Using confocal imaging, we localized the CACNA1H voltage gated calcium channel to the PM of apical ESs, and the VAPA integral ER protein, which is associated with lipid synthesis and exchange machinery, to the apical ER as well as the ER in basal regions known to contain ESs. This data adds further weight to the hypothesis that calcium exchange is integral to regulation of ES and TBC structure, as well as begins our exploration into the possible role of lipid exchange at these sites.

Support or Funding Information

This work was supported by the Natural Sciences and Engineering Research Council (NSERC) of Canada (155397‐2013 to A. Wayne Vogl)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Upregulation Of Eicosanoid Signalling In Lung Following Fipronil And Endotoxin Interaction
Authors: Sethi Ram Saran , Pandit Arif Ahmad , Gandham Ravi Kumar , Verma Ramneek , Singh Baljit
Published: June 2021

It is estimated that nearly 1.8 billion people worldwide use pesticides to minimize the crop damage in efforts to increase food production to feed the growing world population. Exposures to lower concentartion of various pesticides such as fipronil and phenylpyrazole can result in fatal poisonings as well as airway inflammation. Inhalation of endotoxins, which are frequently present in the agricultural buildings, also induces lung inflammation. Animals exposed to both fipronil and endotoxin may demonstrate enhanced lung inflammation. There, however, are limited data available on the molecular mechanisms underlying the toxicity induced by fipronil alone or in combination with endotoxin. Hence, we employed a comprehensive microarray approach in a mouse model to identify the molecular mechanism involved in the lung inflammation by exposing the mice to fipronil (1/20th of LD50/d orally for 90 d) followed by intranasal E. coli LPS (80μg/mouse) or normal saline (N=10 each). Fipronil with or without LPS significantly (p<0.05) increased the total histopathological score suggesting lung damage. Ingenuity Pathway Analysis (IPA) software analysis predicted eicosanoid signalling among the top 5 canonical pathways involved in airway inflammation. Eicosanoid signalling plays important role in pulmonary inflammation and immune response via cytokine production, cell proliferation, antibody formation and antigen presentation. Fipronil alone caused 3‐fold increase in Phospholipase A2G5 (PLA2G5), which hydrolyses the membrane phospholipids to generate various eicosanoids. PLA2G5 showed 1.63 and 4.34 fold increase following exposure to endotoxin alone or in combination with fipronil, respectively. RT‐qPCR and immunohistochemistry analysis validated the microarray data for the PLA2G5. The data taken together indicate up‐regulation of functionally relevant PLA2G5 during fipronil or/and endotoxin induced lung inflammation, which may also play a role in lung inflammation associated with fipronil exposure.

Support or Funding Information

GADVASU, Ludhiana

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Non‐Canonical DNA in the c‐Myc Hotspot Region
Authors: Long Kimberly , Kerwin Sean M. , David Wendi
Published: June 2021

G‐quadruplexes and H‐DNA are non‐canonical or non‐B form DNA structures. G‐quadruplexes are three dimensional structures formed by planar stacking of guanine quartets in the DNA. H‐DNA structures form at inverted repeats consisting of poly(purine)•poly(pyrimidine) sequences, in which the poly(purine) portion of one half of the repeat forms a Pu•Py‐Pu triplex by binding to the duplex form of the other half. Genetic “hotspots” are regions of DNA that are especially susceptible to double strand breaks. It has been shown that the structural conformation of DNA could be the cause for some of these breaks in “hotspot” regions. In particular, the well‐known transpositions involving the c‐Myc oncogene in Burkitt Lymphoma involves a “hotspot” region of the gene that is proposed to form two non‐canonical structures, a G‐quadruplex and H‐DNA structure, which are in competition which each other.

Surface plasmon resonance (SPR) was used to detect H‐DNA or G‐quadruplex DNA formation in c‐MYC. An immobilized single‐stranded DNA construct encompassing the c‐MYC hotspot region was probed with short oligonucleotides complementary to different regions of the putative H‐DNA and G‐quadruplex DNA structures. These studies demonstrated that both the duplex portion of the putative H‐DNA as well as the poly(purine) track displayed limited hydridization to complement probes, commensurate with formation of an H‐DNA structure.

Spectroscopic methods such as UV/vis melting, Förster resonance energy transfer (FRET), and circular dichroism (CD) have been used in previous studies to detect and characterize G‐quaduplexes. Using these spectroscopic methods, we were able to characterize the formation of non‐canonical DNA structure from an oligonucleotide comprised of the entire poly(purine) tract from the c‐MYC hotspot. These studies demonstrate that this poly(purine) sequence adopts a stable intermolecular G‐quadruplex in preference to the intramolecular G‐qaudruplex predicted earlier. This means that the H‐DNA in c‐MYC is likely not in competition for formation with the intramolecular G‐quadruplex suspected to form from this same region.

Support or Funding Information

Cancer Prevention and Research Institute of Texas

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Long and Short of Synaptonemal Complex Assembly: Investigating the genesis and functional relevance of a smaller Zip1 isoform
Authors: Shames Alexander , MacQueen Amy J.
Published: June 2021

The proper segregation of homologous chromosomes (homologs) during meiosis is a paramount task for living cells. Homologs successfully disjoin from one another on the meiosis I spindle so long as they have undergone prior alignment and have formed at least one crossover attachment. The Zip1 protein promotes an intimate alignment between homologs during budding yeast meiosis through its role as a building block of the synaptonemal complex (SC), a proteinaceous structure that assembles along the full length of aligned partner chromosomes. Independent of its role in building SC, Zip1 also plays a critical role in promoting interhomolog crossover recombination, although the molecular underpinnings of this activity are unclear. Through a structure‐function study to identify residues that are essential for Zip1's crossover recombination or SC assembly functions, we identified two zip1 mutants that produce an unexpectedly smaller form of Zip1. In meiotic cells that express either a zip1 allele in which six contiguous residues are changed to alanine, or an allele in which only two of the aforementioned six residues are changed to alanine, a large fraction of Zip1 migrates on a protein gel as if it has lost ten kilodaltons of its wild‐type molecular weight. The change in apparent molecular weight of Zip1 observed in our mutants raises the unanticipated possibility that most Zip1 proteins within the wild‐type meiotic cell are constitutively modified, perhaps by a ten kilodalton protein, and that this post‐translational modification is dependent on the residues that are altered in our two zip1 mutant alleles. Here we describe the results of two experiments designed to ask whether Zip1 in wild‐type meiotic nuclei is constitutively modified by the SUMO protein. First, we will examine the apparent size of Zip1 protein when two SUMO ligase proteins, Siz1 and Siz2, are simultaneously removed. Second, we will determine whether Zip1's molecular weight appears increased in a strain that expresses a larger molecular weight version of SUMO, 3xHA‐SUMO. Finally, we will examine SC assembly and crossover recombination in the two zip1 mutant strains that produce the putative unmodified form of Zip1, in order to determine a function for this potential post‐translational modification.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Molecular insights into replication‐associated genome instabilities caused by CST deficiency
Authors: Chai Weihang
Published: June 2021

The RPA‐like ssDNA binding protein complex CTC1/STN1/TEN1 (CST) has recently been implicated in promoting genome stability under replications stress. Deficiency in CST genes cause the Coats plus syndrome, a complex genetic disorder affecting multiple systems including brain, retina, bone, gastrointestine, and other parts of the body. Yet the precise role of CST in countering replications stress is unclear. Recently we have reported that CST is enriched at GC‐rich and repetitive fragile sequences genome‐wide in response to hydroxyurea (HU)‐induced replication stress (Chastain et al, 2016, Cell Rep 16, 1300–1314). CST deficiency leads to spontaneous chromosome breakage and high‐level chromosome fragmentation that can be further exacerbated by replication stress. Mechanistically, we have found that upon fork stalling, CST proteins form distinct nuclear foci that colocalize with RAD51. Furthermore, replication stress induces physical association between CST with RAD51 in an ATR‐dependent and DNA‐independent manner. Strikingly, CST deficiency diminishes HU‐induced RAD51 foci formation and reduces RAD51 recruitment to GC‐rich fragile sequences. Collectively, our findings establish that CST promotes RAD51 recruitment to GC‐rich repetitive sequences in response to replication stress to facilitate replication restart, thereby providing insights into the mechanism underlying genome stability maintenance. In the present research, we characterize eleven reported disease‐causing CTC1 missense mutations in maintaining genome stability. Our results show that these mutations induce spontaneous chromosome breakage that leads to global genome instabilities. These mutations abolish or reduce CST interaction with RAD51, disrupt RAD51 foci formation, and/or diminish binding to GC‐rich genomic fragile sites under replication stress. Furthermore, CTC1 mutations limit cell proliferation under unchallenged condition and significantly reduce clonal viability under replication stress. Our findings thus provide molecular evidence linking replication‐associated genomic defects with Coats plus disease pathology.

Support or Funding Information

Research is supported by NIH R01GM112864.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Rad53p Activation Alters Chromatin Structure, Induces Respiration and Elevates Cellular ATP Level
Authors: Bu Pengli , Shah Ankita , Zeng Joey , Bhagwat Madhura S. , Nagar Shreya , Vancura Ales
Published: June 2021

Rad53p is an essential kinase in the DNA damage response in Saccharomyces cerevisiae. Here we show that activation of Rad53p, either via genetic manipulation or by treatment with genotoxic chemicals, induces mitochondrial respiration. Activation of Rad53p results in reduced transcription of histone genes and globally decreased DNA nucleosome occupancy. This globally altered chromatin structure leads to increased expression of genes encoding enzymes of tricarboxylic acid cycle (TCA), electron transport chain (ETC), and oxidative phosphorylation (OXPHOS) and increased mitochondrial DNA (mtDNA) copy number, oxygen consumption, and ATP synthesis. These findings are surprising and counterintuitive, since it is widely believed that DNA damage results in downregulation of oxidative metabolism to protect DNA from the effects of reactive oxygen species, produced during ETC and OXPHOS. However, our results indicate that the elevated ATP synthesis improves cell survival under conditions of DNA damage and Rad53p activation. In summary, our study reveals a novel role for Rad53p activation in the restructuring of global nucleosomal chromatin architecture and concomitant metabolic transition from fermentation to respiration.

Support or Funding Information

Supported by NIH GM120710

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Genome‐Wide Analysis of Bromodomain Histone Di‐Acetyllysine Ligands
Authors: Signorelli Sylvia , Gerrard Diana , Glass Karen , Frietze Seth
Published: June 2021

Combinatorial patterns of epigenetic modifications on core histone proteins serve in the marking of genomic elements for the recruitment of regulatory proteins to effect critical biological functions. The recognition of acetylated histone lysine residues by bromodomain chromatin readers appears to be an essential cellular mechanism that regulates protein‐protein interactions, chromatin remodeling, and gene expression. However, the functional relevance of the association of bromodomain chromatin readers and their genome‐wide association with various combinations of histone acetylations remains poorly understood. ATAD2 is a bromodomain‐containing protein highly overexpressed in solid tumors, and has been correlated to an increased risk of cancer metastasis and recurrence. ATAD2B (KIAA1240) is a poorly studied paralog of the ATAD2 gene, and although ATAD2 and ATAD2B are highly conserved, there is little known about the function of ATAD2B. In vitro binding assays demonstrate ATAD2 and ATAD2B bromodomains selectively recognize H4K5/H4K12 and H4K5/H4K8 di‐acetyllysine marks, respectively. To study the genome‐wide patterns of ATAD2/2B histone ligands, we used a Chromatin Immunoprecipitation followed by high‐throughput sequencing (ChIP‐seq) assay to characterize the genomic distribution of H4 tail lysine acetylation patterns in MCF7 breast cancer cells. We performed replicate H4K5, H4K8, H4K12 ChIP‐seq assays as well as H4K5/K8 or H4K5/K12 sequential ChIP‐seq assays. We provide a high‐resolution map of combinations of histone tail acetylation in MCF7 cells. Our results reveal unique patterns of chromatin states, genes and putative transcription factor binding sites associated with different combinations of diacetyllysine residues. Ongoing work seeks to define the recruitment of ATAD2/B, as well as other chromatin effector proteins, and to understand the functional relevance of histone ligand recognition on breast cancer cell gene expression programs.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Salt Bridge Formation within the RKS Motif of Histone H3 Detected by NMR Spectroscopy
Authors: Estrada‐Tobar Zoila M. , Fuentes Daniel , Zurita‐Lopez Cecilia I.
Published: June 2021

Histone N‐terminal tails are subject to covalent post‐translational modifications (PTMs) including methylation and phosphorylation which can help signal gene activation. More recently, PTMs are studied for their impact on each other. In many cases, one modification may prevent that of another. We set out to physically characterize the formation of a salt bridge between arginine‐8 and phosphoserine‐10 in histone H3 by nuclear magnetic resonance (NMR) spectroscopy. We hypothesize that both methylated arginine and phosphorylated serine do not co‐exist when in close proximity; phosphorylation of serine‐10 forms a salt bridge with neighboring unmodified arginine‐8, preventing arginine‐8 methylation. To investigate the contributions of these two modifications, 31P NMR was used for salt bridge detection because the natural isotopic abundance of phosphorus (100 %) is high enough to observe without the need for isotopic labeling. To calculate the relative free energy of the salt bridge, we carried out a 31P NMR titration using four modified peptides that correspond to histone H3. Our NMR data confirm the formation of a salt bridge. The detection of a salt bridge provides evidence that this interaction changes the conformation of the histone H3 tail and also prevents subsequent methylation of arginine‐8. This work can lead to mechanistic insights into the modification of histone H3 tails in both healthy and diseased states.

Support or Funding Information

National Science Foundation Louis Stokes Alliance for Minority Participation HDR‐1602210

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Stoichiometry of Multi‐protein Complexes containing RTT109, Vps75, and Histone H3–H4
Authors: D'Arcy Sheena , Akhavantabib Noushin , Krzizike Daniel
Published: June 2021

RTT109 is a fungal‐specific acetyltransferase that acetylates lysine residues in histone H3–H4. These acetylations play a critical role in nucleosome dynamics as required for DNA transcription, replication and repair. RTT109 acetylation of specific histone H3 lysine residues requires Vps75. Vps75 is a histone chaperone belonging to the Nap‐family; it directly binds the RTT109 enzyme and the histone H3–H4 substrate. The goal of this study was to determine the absolute stoichiometry of the active complex containing RTT109, Vps75, and histone H3–H4. The stoichiometry was unclear as a Vps75 homodimer can crystalize with either one or two copies of RTT109. A high‐resolution structure of the complex between Vps75 and histone H3–H4 is not available, and both Vps75 and histone H3–H4 are known to form oligomers in solution. To measure absolute stoichiometry, we have used an elegant series of analytical ultracentrifugation experiments. We characterize all possible combinations of RTT109, Vps75, and histone H3–H4. These centrifugation experiments are complemented by other biophysical approaches. Our final stoichiometry, combined with targeted mutagenesis, reveals the organization of the active complex containing RTT109, Vps75, and histone H3–H4. Our results pave the way for high‐resolution structural studies and have repercussions for the mechanism and preferred substrate of RTT109 bound by Vps75.

Support or Funding Information

Startup funds to S. D'Arcy from The University of Texas at Dallas.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.