bluuebunny/crossref_metadata_2025 huggingface dataset:
Prolonged enteral leucine supplementation improves lean growth in neonatal pigs fed a restricted protein diet (258.6)
Authors: Columbus Daniel , Steinhoff‐Wagner Julia , Suryawan Agus , Kao Michelle , Hernandez‐Garcia Adriana , Boutry Claire , Nguyen Hanh , Almonaci Rosemarie , Scull Brooks , Fiorotto Marta , Davis Teresa
Published: June 2021

Many low birth weight infants experience growth failure due to feeding intolerance resulting in reduced nutrient intake. The branched‐chain amino acid, leucine, has been shown to have anabolic effects on skeletal muscle. The objective of the study was to determine if prolonged enteral leucine supplementation improves lean growth in neonatal pigs fed a restricted protein diet. Five‐day‐old pigs (N = 22; 1.9 ± 0.3 kg initial BW) were fed by gastric catheter either a control protein (CP; 22.5% protein, 0.98% leucine) or restricted protein (RP; 11.2% protein, 0.49% leucine) milk replacement diet or RP supplemented with leucine to the same level as in the CP diet (RP+L). Pigs were fed 40 ml/kg BW per meal every 4 h for 21 d. Body composition was determined by dual‐energy x‐ray absorptiometry on d 0 and 20. Feeding the CP diet resulted in greater total body weight and lean body mass after 21 d compared to RP fed pigs (P < 0.05). Mass of the longissimus dorsi muscle, heart, and kidney were also greater in the CP than RP fed pigs (P < 0.05). Body weight, lean body mass, and mass of the longissimus dorsi, heart, and kidney in pigs fed the RP+L diet were intermediate to the RP and CP fed pigs. Fat content and gain were similar for all groups. Prolonged leucine supplementation of a marginal protein diet has the potential to improve overall growth in neonatal pigs and infants with restricted protein intake through an increase in lean tissue growth.

Grant Funding Source: Supported by NIH HD072891 and USDA/ARS 6250‐51000‐055
The more sugar and calories, the better? Children’s perspective on sugar‐sweetened beverages (262.7)
Authors: Kharmats Anna , Hopkins Laura , Anderson Steeves Elizabeth , Mui Yeeli , Gittelsohn Joel
Published: June 2021

Caloric menu labeling is currently used as a strategy to promote healthier dietary choices. However, because caloric menu labels may be challenging to interpret, providing information about the amount of time it would take to burn off the calories consumed has been suggested. Unfortunately, children may interpret this information in an unintended way. Three focus groups were conducted with youth participants (n=17) ages 10 to 14 to investigate their understanding of promotional materials that depicted the high sugar and caloric content of sugar‐sweetened beverages (SSBs) and the amount of jogging time it would take to burn off the calories. Before viewing the promotional materials, children expressed positive opinions of water, and negative opinions of soda. Younger boys commented that soda makes them hyper. Surprisingly, when reviewing promotional materials, younger boys expressed positive views of sports drinks, because the sugar and calories in these beverages would give them more energy to exercise, even when the messaging portrayed added sugar negatively. Girls and older participants interpreted the promotional materials as intended by the researchers. These results indicate the need for additional research to find effective communication strategies to promote consumption of healthier beverages among younger children, and to counteract marketing campaigns that depict SSBs as healthy options.

Grant Funding Source: NIH
Meetings Outside the Box: a program to build skills in science communication through networking of graduate level trainees and high school teens (29.1)
Authors: Evans Teresa , Blake Nicquet
Published: June 2021

The vision of this project is to leverage an existing program for graduate level trainees (Trainee Meetings Outside the Box, TMOB) in the launch of a newly developed program for high school teens (Teen Meetings Outside the Box, TeenMOB). TMOB has been extremely well received amongst graduate students and postdoctoral fellows at the University of Texas Health and Science Center at San Antonio (UTHSCSA). The mission of TMOB is to provide a structured setting for the discussion and development of scientific communication skills for trainees, which will serve as the foundation for the continued effective communication of science and career development. Following this model, we will create an environment that allows TMOB trainees to work closely with teens interested in science to organize a sustainable teen science café (TeenMOB). The goals of TeenMOB are to: 1. Provide dynamic, engaging, and facilitated group discussions designed to promote scientific career building skills and mentoring relationships among UTHSCSA research trainees and San Antonio teens with a focus on underrepresented minority (URM); 2. Increase STEM education experiences for high school students who are underrepresented in STEM fields; 3. Provide experiences for graduate trainees to increase science communication and mentoring skills; and 4. Establish the first sustainable teen science café network within San Antonio.

Grant Funding Source: Supported by ASBMB Science Outreach Seed Grant
Outreach and the Otterbein University UAN: Campus and Community (29.4)
Authors: Bowman Jacob , Saari Sofia , Bickers Bridget , Geiger Robert , Bennett Jennifer , Tansey John
Published: June 2021

The Otterbein University UAN chapter is an active chapter that recognizes the value of scientific outreach. Our outreach efforts can be categorized as those that happen for the campus community and those that are for the broader community. Each week our UAN meets for a coffee and tea hour (currently called the BMB tea or BMBT). These informal meetings are open to the campus and frequently draw students and faculty from other branches of science. Currently we are focused on three other ongoing or upcoming endeavors. We have started a standardized test study group to assist students in preparing for the GRE or MCAT. We have started conversations with various constituencies on campus to start a Women in Science group and finally we are running an outreach session to a career fair this coming April.
Sharing science around the bay: Public outreach efforts of the San Francisco State University UAN Chapter (29.5)
Authors: Baird, Jr Teaster
Published: June 2021

The San Francisco State University Chapter of the ASBMB Undergraduate Affiliate Network (UAN) has made public outreach one of its missions. One of our goals in carrying out public outreach activities is to make science more accessible, interesting and relevant to the general public by presenting science in entertaining and interactive ways. In our efforts to achieve this goal, our UAN chapter has participated in local and regional activities, described in this poster, in the San Francisco Bay Area. We have hosted booths and carried out the “Taste of Genetics” demonstrations at AT&T Park during Discovery Days as part of the Bay Area Science Festival, where an estimated 30,000 people attended, and at local elementary schools for classes of 3rd ‐ 6th graders. One of our more unique and well‐received events was a science‐themed costume party that was held in October 2013. The “Enlightenment Party” was open to the public and arose through a collaborative effort between our SFSU UAN chapter, the ASBMB Public Outreach Committee (POC), and Guerilla Science (http://guerillascience.co.uk). Presenting science in unconventional ways has proven to be an effective means for the SFSU UAN chapter to engage the public in science.
The social network: how are students communicating outside of the classroom? (336.2)
Authors: Doubleday Alison
Published: June 2021

As the number of available tools for collaboration and social communication increase, some students are moving informal class discussions to these venues. How are students using these tools to communicate with each other about course material and is it possible to determine whether this type of informal learning impacts what happens in the formal classroom? Qualitative analysis of focus groups and interviews with dental students provides insight into these questions. Results indicate that participating students are deliberate and specific in their use of social networking tools outside of class and that students have expectations for how, when and why these tools are used. Knowing which factors play a role for students as they develop collaborative learning strategies and as they select communication tools will allow faculty to incorporate intuitive and accessible tools and technologies into the classroom and as part of the overall curriculum.
The role of aggrecan in embryonic growth plate cytoarchitecture and differentiation: a rescue model (344.6)
Authors: Lauing Kristen , Cortes Mauricio , Domowicz Miriam , Henry Judith , Schwartz Nancy
Published: June 2021

The homozygous cmd mouse harbors a mutation resulting in the complete absence of the proteoglycan aggrecan in the extracellular matrix of growth plate cartilage. In this study, we examine specifically how this proteoglycan influences the cytoarchitecture and differentiation of chondrocytes within growth plate zones. We utilize the cmd mouse (cmd/cmd) and a novel transgenic mouse expressing chick aggrecan driven by the Col2a1 promoter (tg/+) to elucidate the role of this proteoglycan on chondrocyte survival and proliferation, as well as growth factor production and chondrocyte transcription factor expression. E18.5 cmd embryos display a 55% decrease in limb length, aberrant spatial and temporal mRNA expression patterns of chondrocyte growth factors, and a significant increase in cell death levels in chondrocytes adjacent to the perichondrium compared to wild type controls. Expression of the novel transgene in “rescue” embryos (cmd/cmd::tg/+) results in a 20% increase in limb element length compared to cmd mutant embryos. In situ hybridization demonstrates a partial restoration of normal growth factor and transcription factor gene expression patterns for Indian hedgehog, Patched, Collagen X, FGFR3, and Sox9 compared to cmd growth plates. In addition, the growth plates of rescue embryos display improved chondrocyte morphology and matrix production, particularly within the proliferative and hypertrophic zones. Notably, rescue growth plates display significantly decreased cell death levels compared to cmd embryos in the perichondrial region while concomitantly increasing proliferation levels in the growth plate. The results of this study suggest that aggrecan influences the temporal and spatial patterns of growth factor production by chondrocytes in the embryonic growth plate, and is essential to chondrocyte organization, morphology, and survival during embryonic limb growth.

Grant Funding Source: Supported by the National Institutes of Child Health and Human Development (R01 HD017332)
Translating DBERese (348.2)
Authors: Osgood Marcy
Published: June 2021

Every discipline has its own language. Becoming comfortable with the vocabulary and conceptual rules of a language allow a new speaker to communicate with other natives. Discipline‐Based Education Research (DBER) has its own jargon, which incorporates terms and ideas from both the specific STEM discipline, and from the learning sciences. This interactive presentation will define key phrases, suggest the best resources for improving eloquence, and encourage practice in translating and speaking DBERese. Several different routes to becoming fluent will also be outlined.
Mitochondrial SUMOylation as a critical regulator of proliferation and death (349.3)
Authors: McBride Heidi , Zunino Rodolfo , Goyon Vanessa
Published: June 2021

In our search for the mechanisms that regulate mitochondrial division, we uncovered a role for SUMOylation as a post‐translational modification that regulates the activity of the fission GTPase Drp1. Although we first documented the SUMOylation of Drp1 in the activation of mitochondrial fission, there are a large number of uncharacterized SUMOylated substrates on the mitochondria. To better understand the global impact of mitochondrial SUMOylation, we identified a mitochondrial anchored SUMO E3 ligase called MAPL (mitochondrial anchored protein ligase, also called MULAN/MUL1), and SenP5, a protease that acts upon mitochondrial substrates. We previously showed that SUMOylation on the mitochondria is stabilized during cell death, and is lost during mitosis. The mitotic loss is due, at least in part, to the recruitment of SenP5 to the mitochondrial during the G2/M transition. To examine the role of mitochondrial SUMOylation in vivo, we generated a MAPL knock‐out mouse strain, which provides further evidence for a global role of mitochondrial SUMOylation in the regulation of cell survival and proliferation. These mice present with increased proliferation within the gut, resulting in a lean phenotype due to nutrient maladsorption. After 12 months, nearly 90% of the MAPL KO animals develop hepatocellular carcinoma. Combined with an analysis of the human tumor samples, our data strongly implicate MAPL as a novel mitochondrial tumor suppressor. We will discuss these data, highlighting the importance of mitochondrial SUMOylation as a critical regulator of signaling from the mitochondria.
Superhero superoxide dismutase (360.2)
Authors: Culotta Val
Published: June 2021

From archaic bacteria to humans, the superoxide dismutase (SOD) metallo enzymes play important roles in controlling reactive oxygen species. SOD enzymes fall into three classes based on metal co‐factor used: (i) copper, (II) nickel or (iii) iron/manganese. These three classes are completely unrelated in sequence but have converged in evolution to catalyze the disproportionation of superoxide at rates limited only by diffusion. Like all metalloproteins, SOD enzymes face the challenges of acquiring the proper metal co‐factor under conditions of limited metal availability and in a sea of competing non‐cofactor metals. This issue is particularly problematic at the host‐pathogen interface, where the host attempts to starve the pathogen of certain essential metals and bombard with toxic levels of other metals. In spite of these harsh metal conditions, pathogenic bacteria and fungi manage to properly activate their SODs with the correct metal co‐factor and combat the oxidative burst of host immunity. Two such examples include the Lyme disease pathogen Borrelia burgorferi and the infectious yeast Candida albicans. Both organisms have evolved with a novel family of SOD enzymes and metal homeostasis mechanisms that ensure SOD enzyme activation at the host‐pathogen interface. These adaptations in metals and SOD enzymes will be the focus of discussion.
Molecular interplay at the ribosome exit site (362.1)
Authors: Shan Shu‐ou , Ariosa Aileen
Published: June 2021

The ribosome exit site is a crowded environment where a variety of protein biogenesis factors interact with a nascent protein. Accurate decision‐making must be made at the ribosome exit site to ensure that the nascent protein engage the correct factors, and thus enter the proper biogenesis pathway. The molecular mechanisms by which these decisions are made have remained elusive. To address this question, we investigated the molecular interplay between the signal recognition particle (SRP), a conserved protein targeting machine that mediates the localization of proteins to the cellular membrane, and trigger factor (TF), a major co‐translational chaperone in bacteria. The results revealed multiple mechanisms by which TF and SRP influence one another, and provide a conceptual framework to understand molecular interplay in the crowded environment at ribosome exit site.
Iodized salt benefits infant mental development: preliminary results from a cluster randomized trial in Ethiopia (389.4)
Authors: Bougma Karim , Marquis Grace , Aboud Frances , Frongillo Edward , Melka Tizita , Samuel Aregash
Published: June 2021

Iodine deficiency is thought to contribute to delayed mental development. The effectiveness of iodized salt programs to eliminate this delay has not been studied in children under 36 months. Using a cluster randomized design, 1880 infants 6 to 10 mo old were administered Bayley Scales of Infant and Toddler Development (BSID‐III) in 60 villages in Amhara, Ethiopia. Following salt iodization legislation, iodized salt was forced early into the markets of 30 villages (intervention group) before it became available in the 30 control villages. The two groups had similar socioeconomic, nutrition, and mental development scores at baseline. The same children (86% of the baseline sample) were re‐assessed with the BSID‐III when 20 to 29 mo of age. Analysis was done with a linear mixed model. The intervention group had significantly higher scores at reassessment on three of the four subscales: cognitive (std mean 6.4 vs 6.1, p<0.05), receptive language (std mean 7.2 vs 6.9, p<0.05), expressive language (std mean 6.4 vs 6.2, ns), and fine motor (std mean 7.94 vs 7.63, p<0.05). Effect sizes were small (d=0.11), possibly due to the low socioeconomic status, diet diversity, and stimulation as well as high prevalence of stunting and access to iodized salt in some control areas that may have hampered greater benefits that iodine alone could provide.

Grant Funding Source: Supported by Micronutrient Initiative, Canada
Nanoparticles in lungs: a model of effects after infection (406.3)
Authors: Fotouhi Ariana , Fotouhi Anna , Boyer Daniella , Herndon Betty , Quinn Tim , Molteni Agostino
Published: June 2021

Single wall nanoparticles (CN) are considered toxic with pulmonary effects that have been compared to asbestos. Our previous studies suggested only acute pulmonary toxicity by CN, but to evaluate synergistic effects this study compared acute (3 hr) intratracheal CN effects on rat lungs in animals with or without an ongoing “second hit” of an intraabdominal Staph. Aureus infection (1 week). Comparison groups: (1) CN only, (2) CN + infection, (3) infection only. Histopathology on 8 features of pulmonary tissue was scored. Lumen patency of pulmonary vessels was averaged. Macrophage (mφ) counts on CD68 stained sections and PMN counts on the three groups were made.

Histopathology showed significant acute toxicity in lung septa (p=0.04), small bronchi (p=0.045) and pulmonary vessels (0.001). Lumen of lung vessels showed greater patency with nanoparticles (3 hr) than either group with infection, although the values had a p >0.05.

Both macrophages (Mφ) and neutrophils (PMN) were elevated in lungs. The infection only group was highest, nanoparticles alone second highest, and nanoparticles plus infection had the lowest total cell counts, but group comparisons were below significance. We conclude that the initial steps to pulmonary toxicity by nanoparticle exposure are not exacerbated by the “second hit” of an ongoing distant infection in this model although acute toxicity was found histologically.

Grant Funding Source: St Luke's Hospital foundation grant
The role of nuclear pore proteins in epigenetic regulation and spatial organization of the genome (471.2)
Authors: Brickner Jason
Published: June 2021

In yeast and metazoa, active genes interact with nuclear pore proteins, which promotes stronger transcription. Targeting to the NPC in yeast is mediated by cis‐acting “DNA zip codes” that are necessary and sufficient to confer targeting to the nuclear periphery. Loci with the same zip codes cluster together. Some inducible genes remain associated with the NPC for several generations after being repressed, a phenomenon called epigenetic transcriptional memory. This interaction poises genes for future reactivation and requires the nuclear pore protein Nup100, a homologue of human Nup98. A similar phenomenon occurs in human cells; for at least four generations after treatment with interferon gamma (IFNg), many IFNg‐inducible genes are induced more rapidly and more strongly than in cells that have not previously been exposed to IFNg. In both yeast and human cells, the recently expressed promoters of genes with memory exhibit persistent dimethylation of histone H3 lysine 4 (H3K4me2) and physically interact with Nups and a poised form of RNA polymerase II. Transcriptional memory in yeast requires several chromatin modifications: H2B ubiquitination, H3K4 methylation and the SET3C histone deacetylase. In human cells transiently depleted of Nup98 or yeast cells lacking Nup100, transcriptional memory is lost; RNA polymerase II does not remain associated with promoters, H3K4me2 is lost, and the rate of transcriptional reactivation is reduced. Thus, Nup binding to active genes plays a conserved role in promoting stronger transcription and Nup binding to recently expressed promoters plays a conserved role in promoting epigenetic transcriptional memory.
Health and Anatomy in the Wiregrass: a series of educational workshops (534.5)
Authors: Collier Larissa , Foster James , Senn Sarah
Published: June 2021

Resources for the study of biological and health sciences are often limited for high schools, especially in rural areas. Anatomy is frequently offered as an elective rather than a required course, and students, particularly those interested in medicine, do not receive hands‐on exposure to human anatomy until college. The anatomy faculty at the Alabama College of Osteopathic Medicine (ACOM) offered two workshops to high school students and educators to increase awareness of the importance and practical application of anatomical sciences in clinical and medical education in the Wiregrass region (13 counties in AL, GA, and FL). ACOM provided the facilities and expertise for the workshops for high school students and educators in 25 high schools. The workshop for students was designed to cultivate interest in the fields of anatomy and medicine, perhaps engaging them to pursue education tracts in health sciences. For teachers, it was an opportunity to connect with other health science educators and build classroom experiences in anatomy. Pre‐ and post‐workshop surveys were administered to participants to determine if interest in anatomy and medicine changed as a result of attending the workshops.
Effects of psychological stress in histone acetylation (537.2)
Authors: Gómez Padilla Paola , Colón Caraballo Mariano , Rivera Ivelisse , Appleyard Caroline , Flores Idhaliz
Published: June 2021

Recently, epigenetic factors, including histone modifications, and environmental factors have shown to play an active role in endometriosis. We have recently shown that the levels of histone de‐acetylates (HDACs) are higher and that histone acetylation marks are decreased in endometriosis lesions. Also, stress contribute to the development of higher number of lesions and inflammatory parameters are exacerbated. Psychological stress has shown to cause changes in the epigenome. We sought to test the hypothesis that in endometriotic tissue from rats exposed to stress, histone acetylation is modulated, which would impact gene expression and lead to higher severity of symptoms. This randomized controlled study consists of two groups of rats, both with surgically induced endometriosis: the experimental group, which was exposed to a 10‐day swim stress protocol and a control group, which was not exposed to stress. On day 60, all rats were sacrificed and examined for the presence of endometriotic vesicles, which were processed for protein extraction and western blot analysis for HDACs, and for acetylated H3K9 and H4K8, as well as the housekeeping gene GAPDH. Densitometry and statistical analyses were performed to the bands. We have standardized the conditions for Western blot analysis of HDAC2 (57kD), H3K9ac and H3K8ac (~20Kd). Our preliminary results identified a specific HDAC2 band, which is expressed at the highest level by a rat exposed to stress, characterized by higher number of lesions and worse inflammatory parameters. Next, we will conduct Western Blot for HDACs and histone marks in both groups and will evaluate possible correlations between protein levels and inflammatory parameters in this rat model. These studies may produce evidence in support of a possible role of stress in modulation of epigenetic markers in endometriosis. Due to the fact that HDACs can be targeted using inhibitors, which can become potential new therapies, these results have a high translational potential.

Grant Funding Source: Supported by: NIH Grant #R25GM096955 and (NCCAM) #3 NC1R15AT006373.
Kruppel‐like factor‐3: role in lipid secretion, transport and beta oxidation (537.3)
Authors: Alnassar Nafla , Parhar Ranjit , Al‐Mohanna Futwan , Hashmi Sarwar
Published: June 2021

Lipid metabolic disorder is a critical risk factor for metabolic syndrome,triggering diseases like obesity and diabetes.Obesity raises the risk for many chronic illnesses including diabetes,and heart disease.A unifying view is that insulin resistance,which arises from an excess of body fat,links T2D to obesity.Consistent with this view it has been observed that fat deposition in insulin effector cells decreases their sensitivity to insulin causing insulin resistance.While genetics and environmental components play a major role in the onset of disease caused by fat accumulation.There has been progress in our understanding of metabolic diseases given the complexity of vertebrate systems that employ many factors acting in overlapping and partially redundant pathways.Highly complicated metabolic networks and sensing mechanisms coordinate the flow of fats through synthesis,storage and breakdown pathways and therefore a composite view of nutrient uptake,transport,storage and utilization is required for its understanding.Mammalian Krüppel‐like factors KLFs play essential function in over all energy metabolisms.Many important factors controlling energy metabolism are conserved between mammals and C.elegans providing a new and powerful strategy to dissect the role of KLF in molecular pathways involved in fat metabolism.The worm intestine is our model system where genetics,molecular biology,and cell biology are used to identify and understand genes required in fat metabolism.Thus,we have found an important role of worm KLF in FA biosynthesis,mitochondrial proliferation,lipid secretion,and beta‐oxidation.We have began to understand the mechanism by which KLF controls these events in lipid metabolism.We have recently observed that worm KLF3 selectively acts on insulin components to regulate insulin pathway activity.There are many factors that control energy metabolism and defects in this control system are implicated in the pathogenesis of human obesity and diabetes.
WhmD promotes the assembly of Mycobacterium smegmatis FtsZ (538.2)
Authors: Bhattacharya Dipanwita , Kumar Ashutosh , Panda Dulal
Published: June 2021

FtsZ assembles into a ring like structure known as Z‐ring at the mid‐cell during cytokinesis. Several proteins are known to assist in the appropriate formation and functioning of the cytokinetic Z‐ring. WhmD, a Mycobacterium smegmatis protein, is essential for septum formation and bacterial cell division. Deletion of WhmD causes deformed septum formation and filamentation of Mycobacterium smegmatis. In the present study, we sought to determine the effect of WhmD on the assembly of Mycobacterium smegmatis FtsZ. Light scattering experiment showed that WhmD increased the assembly of Mycobacterium smegmatis FtsZ in vitro in a concentration dependent manner. Further, WhmD also increased the sedimentable polymerized mass of Mycobacterium smegmatis FtsZ. For example, the amount of FtsZ polymerized was increased by 54% in the presence of 1:1 WhmD and FtsZ. Under the conditions used, FtsZ formed thin filaments. Electron microscopic analysis suggested that WhmD enhanced the assembly and bundling of Mycobacterium smegmatis FtsZ filaments. In addition, WhmD increased the GTPase activity of Mycobacterium smegmatis FtsZ in vitro. For example, 6 µM WhmD increased the GTPase activity of 6 µM FtsZ by ~120%. The results indicated that WhmD may function as a positive regulator of the assembly of Mycobacterium smegmatis FtsZ.
A proteomic strategy for the identification of SGT‐interacting proteins in prostate cancer cells (539.6)
Authors: Prieto Luis
Published: June 2021

A Proteomic Strategy for the Identification of SGTAα‐ Interacting Proteins in Prostate Cancer Cells Molecular chaperones facilitate proper folding and regulation of steroid hormone receptors (SHRs). Upon proper folding, SHRs bind its ligand with high affinity, translocate to the nucleus and initiate gene expression. A fairly novel cochaperone, human small glutamine rich TPR (tetratricopeptide repeat) containing protein alpha (SGTA), is a down‐regulator of aAndrogen (AR), gGlucocorticoid (GR), and pProgesterone (PR) receptors. It binds to heat shock protein (Hsp) 70 and 90 kDa. Additionally, SGTA plays a role in cellular processes such as cell cycle progression and apoptosis. Therefore, a top‐down proteomic approach was used to determine novel interactors in LNCaP human prostate cancer cells line LNCaP. Recombinant FLAG‐A tagged ‐SGTA version was purified using a FLAG peptide and further analyzed for unknown interacting proteins with liquid chromatography mass spectrometry (LC‐MS/MS). A comparison of normal (150 mM) versus high (300 mM) salt concentration was used to determine strong interactors. Preliminary studies have identified RanBP2 E3 SUMO‐Ligase and microtubule to be present along with SGTA in both concentrations. Thus, future studies aim to validate these interactions with SGTA using nickel purification and assess their functional cellular relevance, which will contribute to the understanding of the role SGTA plays within the chaperoning pathway.
Hypoxia during embryonic development of myocardium (542.1)
Authors: Nanka Ondrej , Krejci Eliska , Barth Jeremy , Sedmera David
Published: June 2021

Our previous results demonstrate that normal embryonic development involves physiological tissue hypoxia. Through analysis of avian embryos we found that hypoxic conditions caused alteration in myocardial architecture. We also found that the extent and distribution of hypoxic regions differed in the embryos in relation to HIF1α levels. Now we have used microarray analysis to examine how gene expression levels are affected under hypoxic conditions at embryonic day 8 in comparison to embryos at normoxic conditions.

Chicken eggs were incubated at 38□C under normoxic (21% O2) conditions for the first 48 hrs and subsequently under hypoxic (16% O2) conditions. Controls were incubated in normoxia only. RNA was extracted from isolated normal and hypoxic chick embryonic ventricles. cRNA was hybridized to Affymetrix Chicken Genome array according to manufacturer protocols. We identified 164 transcripts that were differentially expressed between the normoxic and hypoxic group (fc>1.5 and p<0.05). Analysis of gene ontology information for the subset of transcripts up‐regulated by hypoxia (89) detected functional relationships to cell growth, cell differentiation, muscle contraction and signal transduction. Similar ontology analyses of transcripts down‐regulated by hypoxia (75) detected functional associations with cell communication, ion transport, and signal transduction.

Grant Funding Source: Supported by Charles University PRVOUK 1105‐280002 and GACR P302/11/1308; NIH grants GM103342,103499
The type I TGFß receptor ALK‐1 and it’s ligands promote angiogenesis (544.2)
Authors: Duffy Iain , Zundell Joseph , Robinson Jesse , Hawker James
Published: June 2021

Control of angiogenesis is critical in diseases such as cancer, heart disease, or wound repair. Growth factors such as transforming growth factor beta (TGFß) and fibroblast growth factor 2 (FGF‐2) contribute to angiogenesis. The TGFß family signaling receptor Activin‐Like Kinase 1 Receptor (ALK‐1) is expressed in coronary venular endothelial cells (CVEC) at near confluent cell densities on gelatin, but not collagen but it is induced by TGFß + FGF‐2 during tube formation in collagen. Other genes induced during tube formation were Smad 6 and Fibroblast Growth Factor Receptor 1 (FGFR1). By contrast, ALK‐5, a related TGFß type 1 receptor, and TGFßRII were expressed in these cells in both gelatin and collagen. ALK‐1 is also expressed in other EC types, smooth muscle cells, cardiac fibroblasts, embryonic fibroblasts (3T3) but not epithelial cells. Three ligands: TGFß, activin, and bone morphogenetic protein 9 (BMP‐9), have been shown to bind ALK‐1. We show that endogenous activin binds ALK 1 and can be displaced by exogenous TGFß. It may function to regulate control of ALK 1 activation. The response of endothelial cells to various ALK‐1 ligands may depend on the cell matrix of endothelial cells and which other co‐receptors (type II receptors and endoglin) are expressed. We are: 1) testing expression of various ALK‐1 ligands and co‐receptors in endothelial cells under various conditions; and 2) we are generating mutants of ALK‐1, ALK‐5, and endoglin to further test their roles in angiogenesis.

Grant Funding Source: Research support was from AHA‐TX and Dept Veteran’s Affairs grants (JH)
How the initial compensatory response transition to permanent modifications of cardiac structure and heart failure after myocardial infraction? (547.7)
Authors: Aslam Naveed
Published: June 2021

MI is the first and most significant clinical manifestation of coronary artery disease (CAD). MI initiates a complex sequence of events in left ventricle (LV) leading to structural remodeling. Clinically LV remodeling is manifested through changes in the size, shape and function of heart. Though initial reorientation of ventricular structure is an adaptive response to functional loss of myocardium however, these compensatory changes can transition to permanent modifications leading to heart failure. Partially the transition from compensatory response to functional de‐compensation is linked with impaired calcium homeostasis. Here, I examine the proposal that calcium homeostasis is regulated through a local signaling cascade in cardiomyocytes. The proposal is based on observations that only a very few second messengers can control the complex and diverse functions of cardiac cells. Therefore, it is essential that signaling in cardiomyocytes be compartmentalized to allow diverse stimuli to produce stimuli‐specific responses. Here, through a systems biology approach I show that in cardiomyocytes the local DAG signaling is regulated through a two compartment signaling system. My results imply that after large MI local homeostasis of DAG is disrupted and the loss of this balance leads to prolonged activation of PKC a key molecular target linked with LV remodeling and dysfunctional filling and ejection characteristics in mammalian heart. This study also proposes an explanation for how the DAG homeostasis is regulated during normal systolic and diastolic functions.
Metaanalysis of post‐translational modifications on proteasomal proteins (555.17)
Authors: Hornbeck Peter
Published: June 2021

The 26S proteasome plays a critical in regulating protein degradation in multiple cellular compartments. The activity of the proteasome must respond to short term and long‐term cellular and environmental inputs efficiently. The control of phosphorylation and other post‐translational modifications (PTMs) is a central component of cellular signaling, yet little is known about the location, extent and possible roles of proteasomal PTMs. We analyzed data in PhosphoSitePlus that has been curated from >15,000 peer‐reviewed low‐throughput papers, over 100 publications of high‐throughput PTM analyses, and data from over 7,000 mass‐spec analyses performed at Cell Signaling Technology. Only experimentally verified PTMs are included in the study. They include > 150,000 phosphorylation, 50,000 ubiquitinylation, 24,000 acetylation, and ~ 8,000 methylation sites. We have analyzed the location and distribution of multiple types of PTMs on proteasome subunits and associated proteins, and have identified specific residues that are heavily modified. For example, PSMA3 is heavily phosphorylated on Ser250, but not on Tyr residues, while PSMA2 is heavily phosphorylated on four tyrosine residues including Tyr683 but not on serine residues. In contrast phosphorylation occurs infrequently on PSMA1 and PSMA4. Analysis of the topology of PTMs on available molecular structures reveal that some of the observed tyrosine and serine phosphorylation sites occur on interface residues that could potentially regulate processes such as opening the catalytic chamber and interacting with the regulatory subunits. As a contrast to the PTM landscape of the proteasome, and to place it in perspective with another cellular structure, a similar analysis of the PTM profile of nuclear pore complex will be presented.

Grant Funding Source: NCI, NIGMS, NIAAA
Gene loops enhance mRNA export in yeast (560.8)
Authors: Singh Badri , Hampsey Michael
Published: June 2021

Extraordinary progress has been made in recent years toward understanding the structure of the eukaryotic genome and the mechanisms that underlie gene expression. Missing from the picture, though, is a fundamental understanding of the three‐dimensional organization of the genome and how these structures affect gene expression. We are investigating the higher order structure of genes transcribed by RNA polymerase II (RNAP II) in the yeast Saccharomyces cerevisiae. Using a powerful technique, called “chromosome conformation capture”, designed to probe the three‐dimensional architecture of the genome, we have detected gene loops that specifically juxtapose promoter and terminator regions. Gene loops are formed on actively transcribed genes and are dependent upon components of both the initiation and pre‐mRNA 3’‐end processing machineries. However, neither the structural requirements for formation and maintenance of gene loops, nor the functions of gene loops are well understood. Here, we report that gene loops are RNA‐dependent and facilitate export of mRNA from the nucleus to the cytoplasm. Further, our systematic analysis of genetic and physical requirements for looping (1) Define the transcription and mRNA export factors TREX‐1 and TREX‐2 as required for formation and/or maintenance of gene loops; (2) Determine the function of gene loops by defining how gene loops, the nuclear pore and mRNA export are interconnected. Our working hypothesis is that all protein‐encoding genes form loops. Looping is not essential, but rather facilitates mRNA export by interaction with nuclear pore complex. This is consistent with (i) coupling of export to transcription of nascent mRNA and (ii) interaction of actively transcribed genes to the nuclear pore. Our results define a fundamental process involved in gene expression and illuminate the pathway by which transcription is coupled to mRNA export.

Grant Funding Source: Supported by NIH
Understanding the functional interplay between DNA demethylation and transcription (561.1)
Authors: Wang Dong
Published: June 2021

Dynamic regulation and faithful maintenance of proper DNA methylation patterns are essential for many cellular functions. 5‐Hydroxylmethylcytosine (5hmC), 5‐formylcytosine (5fC) and 5‐carboxylcytosine, newly discovered oxidized forms of 5‐methylcytosine (5mC) are involved in active DNA demethylation process. The roles of these oxidized species of 5mC in epigenetic and transcription regulation have been an area of intensive study recently. Here we report a systematic study of the effects of five different forms of cytosine in DNA on mammalian and yeast RNA polymerase II transcription elongation, providing new insights into potential functional interplay between cytosine methylation status and transcription. In addition, we will discuss our recent effort in developing new chemical tools that target the formylcytosine sites and modulate 5fC‐related biological processes.

Grant Funding Source: NIH GM085136, NIH GM102362
Understanding the role of pea enation mosaic virus mRNA 3’ untranslated region in translation initiation (566.5)
Authors: Liu Qiao , Goss Dixie
Published: June 2021

Most eukaryotic mRNAs use two key elements at opposite ends of the mRNA, the 5’ m7GpppN cap and the 3’ poly adenosine (A) tail, to recruit translation initiation machinery and circularize mRNA. Pea Enation Mosaic Virus (PEMV) mRNA lacks both a 5’ m7GpppN cap and a 3’ poly (A) tail. A novel type of cap independent translation elements (PTE) is found in the 3’UTR and is essential for translation initiation, the detailed mechanisms of initiation are not well known. Direct protein fluorescence quenching and labeled RNA fluorescence anisotropy were used to investigate the binding affinity of PTE fragment with two important eukaryotic initiation factors, eIF4E (Kd=~7.2nM) and eIF4F (Kd=~8.4nM). These binding affinities are as high as those for eIF4E and eIF4F binding with the cap analogue m7GTP and much higher than those for m7GpppG. With the fluorescence anisotropy assays, we also demonstrated that eIF4A, eIF4B and Poly A tail binding protein (PABP), which can form a stable complex with eIF4F in vivo, only slightly increased the binding affinity of eIF4F with the PTE RNA (Kd<5nM). These binding affinities suggest the PTE element can sequester eIFs from the host cell. The role of eIF/PTE binding in ribosome assembly is being investigated.

Grant Funding Source: NSF MCB 1157632
Chaperone‐dependent prion selection: prion‐specific bifunctionality in the J‐protein Sis1 (567.3)
Authors: Hines Justin , Harris Julia , Nguyen Phil , Sporn Zachary , Patel Milan
Published: June 2021

Yeast prions are heritable amyloid aggregates of yeast proteins. Their propagation to subsequent cell generations is dependent upon their fragmentation by molecular chaperones for which mounting evidence indicates that the J‐protein Sis1 acts as a specificity factor. Though Sis1 is essential for the propagation of all yeast prions for which data are available, individual prions exhibit distinctive sensitivities to ectopic Sis1 expression; a phenomenon rationalized by positing that prions have distinct stringencies for their requirement of Sis1 activity. By examining Sis1‐domain requirements of multiple [PSI+] prion variants and confirming our observations in two distinct yeast genetic backgrounds, we find that two yeast prions, [RNQ+] and weak [PSI+], have mutually exclusive requirements for Sis1 function which cannot be rationalized by a singular function for Sis1 in prion propagation. Rather, these observations necessitate that Sis1 must have at least two functional roles which individual prions differentially require for propagation. This phenomenon allows for variant‐specific prion selection in the presence of distinct Sis1 constructs or the human homolog Hdj1. Surprisingly, Hdj1 is capable of promoting [PSI+] prion propagation in place of Sis1 but that this conservation of function is also prion‐variant dependent, indicating that only one of the two Sis1‐prion functions is conserved.
Validation of a respiratory phenotyping assay for yeast indicates that aerobic metabolism is regulated by mitochondrial thioredoxin activity (572.1)
Authors: Weber Emily , Padilla‐Lopez Sergio , Pearce David , Vitiello Seasson , Vitiello Peter
Published: June 2021

Thiol oxidoreductase enzymes, including thioredoxins, glutaredoxins, peroxiredoxins, and glutathione peroxiredoxins, play an important role in regulating cellular redox reactions through reduction of oxidized thiols. These enzymes regulate integral processes such as protein folding, detoxification of reactive oxygen species, and metal and sulfur metabolism; however, many thiol oxidoreductase substrates and consequential functional changes are not known. Since thiol oxidoreductase genes are highly conserved across eukaryotic species, the objective of this study was to screen a Saccharomyces cerevisiae thiol oxidoreductase deletion library for respiratory phenotypes. A novel protocol using the XF Flux Analyzer (Seahorse Biosciences) was established to simultaneously measure real‐time oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) from media which respectively correlates to cellular aerobic and anaerobic respiratory activities in S. cerevisiae. This approach was validated by artificially pressuring yeast to preference metabolic pathways by restricting nutrient availability in the media and genetic deletion of critical respiratory enzymes. Simultaneous screening of 15 thiol oxidoreductase deletion strains for OCR and ECAR resulted in a single putative respiratory‐regulating enzyme. Deficiency in thioredoxin‐3 (TRX3) resulted in impaired oxygen consumption, and no other mitochondrially‐targeted thiol oxidoreductases (Grx2, Grx5) recapitulated this phenotype. This suggests that Trx3 may have substrate specificity and regulate the activity of a rate‐limiting aerobic respiratory protein in the mitochondria. Taken together, these studies demonstrate that the XF Flux Analyzer can be used to screen for respiratory phenotypes in S. cerevisiae and that aerobic respiration is regulated through mitochondrial thioredoxin status.
Crystal structure of the bi‐functional quinone reductase/ O‐methyl transferase GilM from the gilvocarcin biosynthetic pathway (584.6)
Authors: Downey Theresa , Tibrewal Nidhi , Tsodikov Oleg , Rohr Jurgen
Published: June 2021

Gilvocarcin V, produced by Streptomyces griseoflavus and various other Streptomyces strains, shows significant antitumor activity and remarkably low toxicity. The enzyme GilM a pivotal enzyme in the gilvocarcin biosynthetic pathway exhibits dual functionality in that it catalyzes a reduction of a quinone intermediate to a hydroquinone and an O‐methylation. GilM mediates its reductive catalysis through the aid of GilR that provides FADH2 for the GilM reaction, through which FAD is regenerated for the next catalytic cycle. This unusual synergy eventually completes the biosynthesis of the polyketide‐derived defuco‐gilvocarcin chromphore. Here we report the X‐ray crystal structure of GilM along with insights into its dual functionality and its interaction with the partner enzyme GilR.
A computational evaluation of the steric and electronic contributions to the stability of the structures of α− and β−D‐glucopyanose Part 2: natural bond orbital analysis of monosubstituted cyclohexanes and pyrans (593.3)
Authors: Baello Rofe , Abdulsalam Maryam , Balilli Rahih , Brutofsky Christian , Kasner Marc
Published: June 2021

The anomeric effect refers to the tendency of an electronegative substituent at C2 in a pyran to be lower in energy when in the axial orientation than in the equatorial orientation. By examining the geometric and electronic parameters of a sequence of monosubstituted tetrahydro‐2H‐pyrans (‐CH3 ‐OH and ‐CH2OH) and comparing them to a set of similarly substituted cyclohexanes the steric factors can be evaluated and the magnitude of the electronic interaction can be evaluated. The changes in the bond lengths and bond angles that occur when an oxygen is substituted for a CH2 in the pairs of molecules (cyclohexanes versus the pyrans) show that steric interactions cannot explain all the structural stabilities and energies. Natural Bond Orbital analysis shows that donation of electron density from the ring oxygen occurs in all three pyrans) but, as expected, is most significant in 2‐hydroxyl tetrahydro‐2H‐pyran. There appears to be little or no donation of electron density is the substituted cyclohexanes .

Recognizing that the cyclic configuration of glucopyranose is a substituted tetrahydro‐2H‐pyran and that the ‐CH2OH substituent plays a major role in determining the energetic stability of the compound, analysis of this series of compounds can determine the significance of many of the electronic interaction in glucopyranose . The NBO analysis helps to quantify the details of the electronic interactions that contribute to the geometrical structure of α− and β−D‐glucopyranose.
The nuclear accumulation of alpha‐synuclein is mediated by importin alpha and promotes neurotoxicity in nucleus (596.6)
Authors: Ma Kai Li , Chen Nai Hong
Published: June 2021

α‐synuclein (α‐syn), a 14 kDa presynaptic protein, is widely involved in the Parkinson’s disease (PD) pathogenesis. It has been noted that the nuclear import of α‐syn may be mediated by importin α and that both the amino acid residues 1‐60 and 103‐140 of α‐syn were indispensable for its nuclear import. After imported into the nucleus, the accumulated α‐syn played a toxic role in the PC12 cells. Thus, it can be concluded that α‐syn can accumulate in nucleus, which is mediated by importin α, and promote neurotoxicity in nucleus.

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