bluuebunny/crossref_metadata_2025 huggingface dataset:
The 2010 Dietary Guidelines for Americans food groups associated with decreased inflammation (370.3)
Authors: El‐Abbadi Naglaa , Jacques Paul , Lin Honghuang , Rogers Gail , Troy Lisa
Published: June 2021

Chronic systemic inflammation has been associated with cardiometabolic diseases such as heart disease, stroke and diabetes. We previously demonstrated that diet quality, as assessed by the Dietary Guidelines for Americans Adherence Index (DGAI_2010), is associated with overall inflammation status. To further explore this relationship, associations between DGA food groups and inflammation were examined among 715 male and 833 female participants of the Framingham Offspring Cohort. DGAI_2010 component scores for adherence to the recommended energy‐specific consumption of 12 food groups were determined, and an inflammation total score (IS‐pro15) was calculated from the ranked values of 15 pro‐inflammatory biomarkers (CRP, fibrinogen, IL‐6, TNFα, TNFR2, OPG, MCP‐1, P‐selectin, CD40L, ICAM, MPO, LPL A2 mass, LPL A2 activity, isocreatine and resistin). A multivariable linear regression model for IS‐pro15 was established through backward selection, and after adjustment for age, sex, BMI, smoking and use of NSAIDS showed inverse associations between IS‐pro15 and the recommended intakes of dark green vegetables, orange and red vegetables, meat, poultry & eggs, seafood and sugar (p<0.05). These suggest that specific food groups in the DGA are protective against inflammation.

Grant Funding Source: Supported by NIH (N01‐HC 25195, RO1 HL64753; R01 HL076784; R01 AG028321) and USDA (#58‐1950‐0‐014).
A multi‐component nutrition intervention utilizing a learner‐centered, garden‐based nutrition curriculum improves nutrition knowledge among fourth‐grade children (379.4)
Authors: Linnell J , Smith M , Scherr R , Briggs M , Nicolson Y , Spezzano T , Brian K , Hillhouse J , Zidenberg‐Cherr S
Published: June 2021

The nutrition education and promotion component of the Shaping Healthy Choices Program (SHCP) included classroom education, garden‐based learning, and cooking demonstrations in fourth‐grade classrooms. Discovering Healthy Choices (DHC), a learner‐centered, garden‐based nutrition curriculum, was developed for SHCP. The objective of DHC is to facilitate understanding of nutrition concepts and development of critical thinking skills so youth can make evidence‐based decisions about the foods they eat. DHC supports Common Core State Standards. Cooking demonstrations were linked to DHC concepts. It was hypothesized that students participating in SHCP would have improved nutrition knowledge and critical thinking skills compared to controls. Schools meeting inclusion criteria in Northern (NC) and Central Valley California (CV) were randomly assigned to control (C) or intervention (I) groups. Pre‐ and post‐assessments included a Nutrition Knowledge Questionnaire and the Test of Basic Science Process Skills (BAPS). Districts were analyzed separately. At baseline, a correlation between nutrition knowledge and BAPS was observed (Pearson’s r=0.51, p<0.01). This correlation was maintained at follow‐up. Significant improvements in nutrition knowledge were found between NCI and NCC (P<0.01) and CVI and CVC (P<0.05). There were no significant differences in BAPS between C and I groups in either district.

Grant Funding Source: Supported by UCANR #11‐1018 and USDA 2011‐38420‐20082
Cellular geometry regulates traction stresses (479.4)
Authors: Oakes Patrick , Banerjee Shiladitya , Marchetti M. Cristina , Gardel Margaret
Published: June 2021

Cells generate mechanical stresses via the action of myosin motors on the actin cytoskeleton. While the molecular origin of force generation is well understood, we currently lack an understanding of the regulation of force transmission at cellular length scales. Here we experimentally decouple the effects of substrate stiffness, focal adhesion density and cell morphology to show that the total amount of work a cell does against the substrate to which it is adhered is regulated by the cell spread area alone. Surprisingly, the number of focal adhesions and the substrate stiffness have little effect on regulating the work done on the substrate by the cell. For a given spread area, the local curvature along the cell edge regulates the distribution and magnitude of traction stresses to maintain a constant strain energy. A physical model of the adherent cell as a contractile gel under a uniform boundary tension and mechanically coupled to an elastic substrate quantitatively captures the spatial distribution and magnitude of traction stresses. With a single choice of parameters, this model accurately predicts the cell's mechanical output as a function of cell geometry.

Grant Funding Source: Supported by the NSF, NIH and the Burroughs Wellcome Fund.
Ethanol exposure alters zebrafish morphology and development, evidence of impaired adipogenesis and adiponectin production (540.7)
Authors: Shafiei Mahnoush
Published: June 2021

Adipose tissue secretes numerous bioactive peptides (“adipocytokines”). Visceral fat mediates inflammatory cytokines. In contrast, subcutaneous fat is the principal source of adiponectin production. Adiponectin, consisting of 247 amino acids and single nucleotide polymorphisms (mutations), causes hypoadiponectinemia. Individuals with this mutation are highly susceptible for development of insulin resistance, cardiovascular disease, hypertention, and left ventricular hypertrophy. We hypothesize that alcohol consumption during pregnancy may alter endocrine signaling, by increasing pro‐inflammatory cytokines thus inducing fetal Alcohol Syndrome (FAS). Even low‐dose/or short duration ethanol exposure early in development can produce more subtle effects by alteration of subcutaneous fat and subsequently, decrease in adiponectin production. Zebrafish eggs (AB strain) were raised in water (control) or in 0.5%, 1%, 1.5%, 2%, and 2.5% ethanol solution for one of 3dpf, 5 dpf, 10 dpf, 15dpf, or 20dpf. Anatomical measures including heart diameter and total body length were compared across groups. At the concentration of 2.5%, there were more abnormal physical distortions and significantly higher mortality rates than any other group.

Nile Red (NR) and Oil Red O (ORO) staining was used to detect adipogenesis (visceral vs subcutaneous) fat depots. Adipogenesis during zebrafish development was altered and subcutaneous fat was depleted at 15pft after 2% alcohol exposure compared with control. The expression of an “adipogenic” transcriptional program including PPARγ, SREBP1c, CEBPα □ and pro‐inflammatory cytokines IL‐6, IL‐8, HIF‐1α ROS, TNF‐α, and NF‐κB was altered, as evident from quantitative RT‐PCR analysis. We conclude that exposure of zebrafish larvae to ethanol during development results in modified external and internal morphological development and that the severity of these effects is due to remarkable changes in adiposity and hypoadiponectinemia.

Grant Funding Source: NSF
Splicing regulation of alternative exon 23a of the neurofibromatosis type 1 pre‐mRNA by DBIRD complex (560.5)
Authors: Nguyen Hieu , Lou Hua
Published: June 2021

Hu proteins are neuron‐enriched RNA‐binding proteins that regulate alternative splicing by binding to AU‐rich sequences of pre‐mRNAs. Recently, we showed that Hu proteins participate in epigenetic regulation of the Neurofibromatosis type 1 (NF1) exon 23a splicing by directly interacting with HDAC2 and inhibiting its deacetylation activity to induce local histone hyperacetylation surrounding exon 23a, leading to increased local transcriptional elongation rate and decreased exon inclusion (Zhou et al., PNAS, 2011).

DBIRD complex, formed by DBC1 and ZIRD, increases transcriptional elongation rate across A/T‐rich exons, promoting exon skipping. NF1 exon 23a was identified as a potential DBC1‐dependent alternative exon (Close et al., Nature, 2012). Other studies showed that DBC1 is an inhibitor of two deacetylases. These studies prompted us to propose an intriguing hypothesis that, at the NF1 transcription site, DBIRD, interacting with DNA, and Hu proteins, interacting with RNA, act in a cooperative manner to inhibit HDAC activities, leading to increased local transcriptional elongation rate to skip the NF1 exon 23a.

We are using biochemical and genetic approaches to test this hypothesis. Our preliminary data showed that Hu proteins interact with DBIRD complex and that DBC1 alone and DBIRD complex significantly change the NF1 exon 23a splicing. We are currently investigating DBIRD‐ and Hu proteins‐mediated epigenetic control of splicing regulation in CNS neurons isolated from mouse brains.

Grant Funding Source: NIH and DOD grants to Hua Lou and Vietnam Education Foundation (VEF) support of Hieu Nguyen
Structural comparison of mutant and wild‐type acidic leucine aminopeptidase suggests the basis for secondary chaperone activity (567.11)
Authors: DuPrez Kevin , Scranton Melissa , Walling Linda , Fan Li
Published: June 2021

The acidic leucine aminopeptidase (LAP‐A) of tomato is involved in wound signaling and insect defense. Observation of chaperone activity by LAP‐A suggests new mechanisms of defense against environmental stresses. To evaluate how this chaperone activity is facilitated, the crystal structures of wild‐type and a loss of function mutant of LAP‐A were determined to 2.20 and 2.30 Å, respectively. LAP‐A is a dimer of trimers composed of 6 bilobal monomers. Six substrate entry channels formed as a result of the hexameric structure connect in a central cavity in which all six active sites are localized. Coordination of two active‐site magnesium ions required for catalysis is facilitated by a highly conserved network of amino acid residues. Mutation of select residues within this network can result in loss of activity and/or breakdown of hexameric structure. The structure of the K354E mutant shows disruption of a loop from residue 350 through 361, resulting in the loss of intra‐subunit contacts that stabilize binding of the catalytic metal ions, thus resulting in loss of peptidase activity. These residues also provide inter‐subunit contacts at the trimerization interface, presumably stabilizing the hexameric form of the enzyme. However, breakdown of the hexamer into a tetrameric form induced by the K354E mutation results in increased chaperone activity, likely through heightened exposure of hydrophobic surfaces. These data reveal insights into the structural basis for LAP‐A's secondary function as a molecular chaperone.

Grant Funding Source: Partially supported by a National Science Foundation grant (IOS 0725093) to LLW.
Probing the active site of the Rv0045c esterase from Mycobaterium tuberculosis (580.4)
Authors: Hoops Geoffrey , Johnson R
Published: June 2021

The active site of the Rv0045c esterase from Mycobaterium tuberculosis was probed with a combination of site‐directed mutagenesis and a series of substrate analogues. The thermal stability of enzyme variants was measured using differential scanning fluorimetry with Sypro Orange dye. The catalytic efficiency of the enzyme variants was measured using fluorogenic ester substrates. Variants of active site residues confirmed a classic serine catalytic triad, as well as revealing the relative importance of several other active site residues. Two active site variants (G90A and H187A) displayed increased catalytic efficiency relative to the wild type, but the corresponding double variant lacked a synergistic activation effect. A series of other H187 variants revealed that further fine‐tuning of enzymatic activity was possible. Additionally, the relative catalytic efficiencies of the hydrolysis of a series of flourogenic ester substrates revealed structure activity relationships, in which short chain, unbranched, hydrogen bond accepting esters displayed maximal activity.

Grant Funding Source: Supported by the National Science Foundation
The stops and starts of protein splicing: regulation and mechanism of non‐canonical inteins (584.1)
Authors: Mills Kenneth , Reitter Julie , Nicastri Michael , Williams Jennie , Colelli Kathryn
Published: June 2021

Protein splicing is the post‐translational excision of an intervening polypeptide (intein) from its flanking domains (the exteins), concomitant with extein ligation. Both the Methanoculleus marisnigri (Mma) and the Pyrococcus abyssi (Pab) PolII inteins promote efficient splicing with C‐terminal Gln in place of the highly conserved Asn, which plays a central role in the third step of the mechanism. The Mma intein is regulated by disulfide bond formation, and the third step of splicing is very slow with C‐terminal Gln. The Pab intein is regulated by both temperature and disulfide bond formation. The unusual stability of the unspliced precursor of the Pab intein has allowed for structural study and analysis of the kinetics of each step of splicing in vitro. An NMR solution structure of the Pab PolII intein reveals a particularly rigid structure, a disordered loop absent in a highly similar P. horikoshii intein, and a beta‐hairpin specific to inteins from archaebacteria.

Grant Funding Source: This material is based upon work supported by the National Science Foundation under grant MCB‐124408
A roadmap for the design and implementation of preparatory or “Gateway” courses for biochemistry and molecular biology (618.21)
Authors: Canfield Jennifer , Osborn JoDi Lynn , Felder‐Sumter Takita , Bell Ellis
Published: June 2021

The “Vision and Change in Undergraduate Biology Education: A Call to Action” report urges life science educators to prepare students for science careers by better aligning scientific instruction with practice. To this end, introductory or “gateway” courses should demonstrate the interdisciplinary nature of the sciences to best prepare students for early engagement in research. We recently published a detailed description of the essential concepts that students must have from introductory chemistry, biology, math, and physics courses based on ASBMB’s national “Vision and Change” dialogues. We now expand the study to include suggestions for implementing and assessing outcomes of these foundation experiences. Effective practices include boot‐camp style remedial sessions; problem based learning approaches; facilitated transition from community colleges; and involves crafting well‐defined outcomes as means of developing critical thinking skills. Moreover, best practices in course delivery and approaches to shift the emphasis from content to process driven courses are presented. Finally, we present assessment models that may be coupled to these approaches. Collectively, the findings underscore the importance of introductory foundational concepts, skills, and allied field topics in bridging the gaps between “gateway” and advanced biochemistry and molecular biology courses. This work is supported by NSF Grant 0957205 to EB.

Grant Funding Source: Supported by NSF Grant 0957205 to EB
Best practices in direct nutrition education for low‐income audiences: program design and delivery (625.2)
Authors: MacKinnon Chloe , Auld Garry , Baker Susan
Published: June 2021

The project objective was to identify best practices in nutrition education for low‐income audiences. Best practices are evidence‐based elements or strategies that have shown effectiveness in achieving outcomes. Utilizing best practices increases stakeholder confidence that programs will be effective in helping low‐income participants make positive nutrition‐related behavior changes. A seven‐member expert panel identified best practices criteria via telephone and electronic correspondence; the criteria were confirmed via literature review. Eight criteria were identified as characteristics of effective nutrition education programs within the following areas: a strong theoretical basis, research‐based content, emphasis on goal setting, and audience‐appropriate languages, activities, recipes, and reading level. Five criteria were identified for program delivery: the use of experiential learning and enhancement items, accommodation of learning styles, inclusion of contacts of appropriate frequency and duration, and maintenance of program fidelity. Best practices criteria for program design and delivery may promote consistency and efficacy within and across low‐income nutrition education programs.

Grant Funding Source: Supported by the National Institute of Food and Agriculture no. 2012‐48757‐20337
Telephone interviews with parents of 4th graders support nutrition education using innovative strategies for low‐income families and after‐school programs (625.5)
Authors: Cheng Feon , Smith Stephanie , Cunningham‐Sabo Leslie , Lohse Barbara
Published: June 2021

Fuel for Fun: Cooking with Kids is a school‐based cooking program for 4th graders (n=415, 8 schools). To design a family component, professional phone interviews were held with their parents (n=24, 41.7 ± 5.7 y ) who represented all surveyed parents (n=85)‐white, female, high school graduates, 63% eating competent (EC), 54% normal weight. 38% used at least 1 assistance program (WIC 25%; food banks 21%; SNAP 17%). Assistance program usage was associated with greater interest in label reading (P=.022), meal planning (P=.019), and affordable meals (P=.013). Parents preferred programs on Friday (63%) or Thursday (50%) from 6‐8 PM (63%) on cooking skills, active indoor and nutrition games. Preferred participation incentives were having a child involved in activities (96%), give‐aways (83%) and being served dinner (79%). 3 topics with definite interest were easy‐to‐fix home meals, food prep with children, and affordable home meals. Families being more physically active was of greater interest than food prep. EC parents were more likely to be physically active (P=.005) and cook (P=.047). Interest in learning food prep was not associated with EC status. 21% read blogs at least weekly but most rarely commented; 13% wanted to learn more about blogs. Children did homework at home (92%), 83% used a computer; 67% of parents spent 30 min helping. Findings supported novel strategies and nutrition education in after‐school programs.

Grant Funding Source: Agriculture and Food Research Initiative Grant no. 2012‐68001‐19603 USDA NIFA
Curcumin and 3’ 4’didemethylnobiletin in combination synergistically inhibit cell proliferation and potentiate apoptosis in HCT116 colon cancer cells (647.37)
Authors: DiMarco‐Crook Christina , Zheng Jinkai , Song Minyue , Xiao Hang
Published: June 2021

Curcumin and 3’, 4’ didemethylnobiletin (DDMN) a major metabolite of nobiletin, have each been shown to exhibit anti‐carcinogenic effects in various colon cancer models. However, the efficacy and molecular mechanisms of these dietary compounds in combination has yet to be explored. In this study the interaction of curcumin and DDMN was investigated in the HCT116 human colon cancer cells. Cell viability assays showed a greater inhibition of growth in cancer cells treated with the combination of curcumin and DDMN than with either treatment alone. Isobologram analysis confirmed the synergy between curcumin and DDMN in inhibiting colon cancer cell growth. Flow cytometry results revealed significant G2/M phase cell cycle arrest and extensive apoptosis induced by the combination, which were much stronger than the effects induced by the treatments with curcumin or DDMN alone. Pursuant to these findings, proteins associated with G2/M arrest and apoptosis were examined by Western Blot analysis to confirm increased expression levels of p21, p53, and C‐PARP, and decreased expression level of cdc2. Indeed, the change in expression of these proteins was much greater in response to the combination treatment of curcumin and DDMN than each compound alone. The synergy between curcumin and DDMN warrants further investigation on their combination to determine its chemopreventive potential for colorectal cancer in vivo.

Grant Funding Source: Supported by NIH, AICR and USDA
The presence of histamine in ganglia and tissues of the bivalve mollusc, Crassostrea virginica (656.7)
Authors: LaFleur Kisha , Harrison Jarreau , Mantone Daniel , Carroll Margaret , Catapane Edward
Published: June 2021

Histamine (HIS), a biogenic amine, is a neurotransmitter in neurons and sensory receptors in invertebrates. HIS has rarely been reported in bivalves. Our physiology work show it involved in sensory reception in sensory‐motor integration of gill lateral cell cilia in the bivalve Mollusc, Crassostrea virginica. We hypothesize HIS is present in ganglia and tissues of C. virginica. We used HPLC to measure HIS in cerebral and visceral ganglia, and peripheral tissues using a Phenomenex Gemini 5µ C18 column, Beckman HPLC system and Jasco FP 2020 Spectrofluorometer. The mobile phase was 40/60 v/v acetonitrile/phosphate buffer (50 mM, pH 6.8, 2 ml/min flow rate). Tissues were dissected, blotted dry, weighed and homogenized on ice with a Brinkman Polytron in 0.4 M HCl. Homogenates were centrifuged (4̊C, 600g, 15 min), then recentrifugation (12,000g, 15 min). Supernatants were filtered through 0.24µ filters, adjusted to pH 9.5 with NaOH, derivatized for 15 min with borate buffer (0.3 ml, pH 9.5) KCN (0.3 ml, 20 mM) and NDA (2,3‐naphthalene‐dicarboxaldehyde, 0.3 mM in methanol), injected into the HPLC and fluorescence measured (450 nM ex, 484 nM em). Results show HIS present in ng amounts in cerebral and visceral ganglia, and peripheral tissues. The study confirms HIS in the nervous system and innervated organs, including sensory tissue, and coupled with our other work shows HIS to be an endogenous biogenic amine in C. virginica.

Grant Funding Source: NIH‐ 2R25GM0600309, NSF‐ 0622197, NYSESD‐ 0516041071
Zn disrupts Cd‐induced stimulation of apical choline uptake in cultured choroid plexus independent of GSH availability (685.4)
Authors: Villalobos A. , Young R , Francis Stuart S
Published: June 2021

In cultured neonatal rat choroid plexus we showed cadmium (Cd) stimulated apical choline uptake and induced oxidative stress and glutathione (GSH) synthesis. Zinc (Zn) supplementation disrupted Cd‐induced oxidative stress and stimulation of choline uptake. Our objectives are to further elucidate GSH regulation by Cd and Zn and determine whether GSH is critical to disruption of Cd‐induced stimulation of choline uptake by Zn. Cultured choroid plexus cells were supplemented (48 h) with 25 μM ZnCl2 in complete medium before 12‐h treatment with 0 or 500 nM CdCl2±10 μM ZnCl2 without serum. By immunoblot analysis and qRT‐PCR, we assessed induction of HO‐1, HSP70, Cu/Zn‐SOD (SOD1), metallothionein‐1 (MT‐1) and both modifier (GCLM) and catalytic subunits (GCLC) of glutamate‐cysteine ligase, the rate‐limiting enzyme in GSH synthesis. Cd induced GCLM, GCLC, MT‐1, HO‐1, and SOD1 mRNA levels by 6‐, 3‐, 16‐, 12‐ and 3‐fold; Zn supplementation attenuated induction. Cd alone increased GSH and GSSG by 2‐ and 30‐fold; Zn supplementation abated increases in each. To determine whether the effects of Zn were dependent on GSH availability, we treated cells with 100 μM buthionine sulfoximine (BSO) to inhibit GCL during Zn supplementation and subsequent 12‐h treatment with 250 nM CdCl2±10 μM ZnCl2. BSO alone or with Zn±Cd reduced GSH by 90% and increased GSSG by 15‐fold above control. In Cd‐treated cells, BSO elicited a compensatory induction of HO‐1, HSP70, SOD1, GCLM, GCLC and MT‐1 and further stimulated apical choline uptake. Zn supplementation attenuated induction of cytoprotective genes and proteins and stimulation of apical choline uptake in cells treated with Cd±BSO. Thus, Zn abates Cd‐induced oxidative stress and GSH regulation, but disrupts stimulation of choline uptake independent of GSH availability.

Grant Funding Source: NSF #IOS‐1052654
The Kölliker‐Fuse nucleus drives respiratory modulation of cardiac vagal tone (686.15)
Authors: Farmer David , Dutschmann Mathias , Paton Julian , Pickering Anthony , McAllen Robin
Published: June 2021

Cardiac vagal tone is a major determinant of health, but its origin is unknown. It is present and well developed in the decerebrate perfused working heart‐brainstem preparation (WHBP) of rat, indicating that it arises from within the brain stem. Direct recordings from the cardiac vagal branch in the WHBP (juvenile rats day 17‐25, n=10) showed spike activity that peaked in late inspiration/postinspiration, correlating with the bradycardia of respiratory sinus arrhythmia (RSA). Bilateral inhibition of neurons in the Kölliker‐Fuse nucleus (KF) of the pons by microinjections of the GABAA receptor agonist, isoguvacine (10 mM, 40‐60nl), abolished the post‐inspiratory peak but had little effect on mean cardiac vagal nerve activity. That activity assumed a tonic pattern, interrupted by brief pauses around the onset of inspiration. RSA was abolished, and heart rate rose modestly (~ 20 beats/min). Transection of the caudal pons at the level of the facial nuclei substantially reduced mean cardiac vagal activity (50‐80%) and raised heart rate substantially (~ 50 beats/min). These results indicate that cardiac vagal tone is distinct from RSA, though both depend on supramedullary connections. Drive from the KF is responsible for RSA, while the origins of most cardiac vagal tone remain to be determined.

Grant Funding Source: ARC
Disinhibition of the pontine and dorsal respiratory groups reveals postinspiratory pacemaker activity (714.2)
Authors: Dutschmann Mathias , Bautista Tara
Published: June 2021

The ‘pontine respiratory group’ in dorsolateral pons overlaps with the Kolliker‐Fuse nucleus (KF) while the ‘dorsal respiratory group’ is associated with ventrolateral areas of the nucleus of the solitary tract (NTS). The contribution of both to the generation of the eupneic respiratory motor pattern remains controversial. This study describes the severe disruption to normal breathing following focal microinjection of bicuculline (GABAA receptor antagonist) into the KF and NTS in the juvenile in in situ preparation of rat. Disinhibition of the KF produced an increased variability in the sequential patterning of the eupneic 3‐phase respiratory motor pattern. The chaotic respiratory pattern after KF disinhibition is largely associated with instability of postinspiratory gating mechanisms. In contrast, disinhibition of the NTS caused repetitive barrages of rhythmic, non‐swallow related discharges in vagus nerve that suppressed inspiratory activity. These rhythmic discharges were likely pacemaker‐driven since they were abolished by the persistent sodium channel blocker riluzole. Moreover, riluzole microinjection into KF or NTS alone triggered delayed inspiratory off‐switch, suggesting that pacemaker currents indeed contribute to the timing of postinspiratory mediated inspiratory off‐switch and respiratory motor patterning.

Grant Funding Source: ARC
Characterization of tauopathy in the ponto‐medullary brainstem nuclei of aged Tau‐P301L mice (728.7)
Authors: Stanic Davor , Bautista Tara , Dutschmann Mathias
Published: June 2021

Neurodegenerative diseases, such as Alzheimer’s disease (AD) and dementia, are often linked to swallowing disorders such that aspiration pneumonia resulting from accidental inhalation of ingested material is a common phenomenon. Growing evidence suggests that malfunction in the coordination of swallowing and breathing leads to increased risk of aspiration. We investigated the underlying pathology linking dementia and swallowing dysfunction in Tau‐P301L transgenic mice, an established mouse model of neurodegeneration. Tauopathy and neurofibrillary tangle‐related neuropathological morphology in the brainstem of 10‐month old Tau‐P301L mice was identified by immunohistochemistry using antisera against: 1) pre‐neurofibrillary tangles (Tau phosphoThreonin 231); and 2) extra‐neuronal neurofibrillary tangles (Tau phosphoSerine 199/202). Cell bodies containing diffuse phospho‐tau‐immunoreactivity within the cytoplasm, together with preserved, collapsed or absent dendrites were identified in the Kölliker‐Fuse (KF) nucleus, ventral swallowing group (VSG) dorsomedial to the nucleus ambiguus, and nucleus of the solitary tract (NTS), regions known to co‐ordinate breathing and swallowing. Our results suggest that neurons in the KF, VSG and NTS are susceptible to neurodegeneration, and provide a platform for examining the effect of tauopathy in functionally defined and experimentally controllable brainstem circuits (for swallowing and breathing). This may facilitate a framework for the development of potential drug therapies for neuroprotection and prevention of aspiration and dementia.

Grant Funding Source: ARC
Characterization of RPA‐protein interactions and their dependence on the phospho‐mimetic state of the N‐terminus of Saccharomyces cerevisiae Rfa2 (735.13)
Authors: Mueller Erica , Piya Gunjan , Severson Amber , Haring Stuart
Published: June 2021

Replication protein A (RPA) is a heterotrimeric complex in all eukaryotic cells that is essential for DNA replication, repair/recombination, and cell cycle regulation. A common intermediate of each of these DNA processes is the formation of single‐stranded DNA (ssDNA) and the major biochemical function of RPA is to bind to ssDNA. Therefore, RPA not only acts as a sensor of incomplete duplex DNA, but also as a recruiter of factors necessary to process this intermediate. It is clear that RPA interacts with a number of proteins; however, many of these interactions have not been well characterized in yeast, making unclear the physiological role of each RPA‐protein interaction in cellular DNA metabolism.It is well known that in response to DNA damage, the N‐terminus of the human 32‐kDa subunit (Rpa2) becomes phosphorylated, and this has been demonstrated to modulate some RPA‐protein interactions. Although phosphorylation of the Rfa2 N‐terminus has not been definitively demonstrated during the DNA damage response, we have demonstrated in our lab that phospho‐mutant forms of the budding yeast Rfa2 N‐terminus show DNA damage‐dependent phenotypes. Therefore, we hypothesize that the phosphorylation state of Rpa2 (or Rfa2) coordinates the response to DNA damage through regulation of RPA‐protein interactions. The goals of this research were: 1) to identify yeast RPA‐protein interactions, 2) to map regions important for protein interaction, and 3) to characterize whether these protein interactions are dependent on the phosphorylation state of the Rfa2 N‐terminus We have identified a number of RPA‐protein interactions that are dependent on the state of the Rfa2 N‐terminus, and we have mapped the region(s) of RPA that are important for some of these interactions. This combined data suggests potential models for how the Rfa2 N‐terminal phosphorylation state may be regulating protein interactions to coordinate RPA cellular function.

Grant Funding Source: Supported by NIH, NIJ, NSF
Probing the role of the E304 and P288 residues in the allosteric site of Agrobacterium tumefaciens ADPglucose pyrophosphorylase (769.19)
Authors: Karzai Hoomai , Bor Simona , Benjamin Vonice , Silva Emmanual , Meyer Christopher
Published: June 2021

ADP‐glucose pyrophosphorylase (ADPG PPase) is the rate limiting enzyme in the glucan biosynthesis pathway, a vital source for renewable and biodegradable carbon. Thus ADPG PPase is an attractive target for protein engineering to produce a more active enzyme to increase the biomass. Agrobacterium tumefaciens (Ag. t) ADPG PPase is activated by F6P and pyruvate and inhibited by phosphate and sulfate via common allosteric site(s). The Ag.t ADPG PPase structure (pdb: 3BRK) indicates that the E304 residue is part of the allosteric site. The P288 residue, which is part of the loop region connecting the N‐terminus domain to the C‐terminus domain, is also involved in allosteric regulation. To probe the roles of these two residues the E304A, E304D, and P288E enzymes were generated, purified and kinetically characterized. The purification scheme included Phenyl Sepharose, anion exchange (DEAE or UNO Q), and Blue A chromatography. Substitution of the P288 residue resulted in changes in allosteric behavior with the largest effects being desensitization to activators as well as higher activity in the absence of activators. A decrease in Vmax as well as stability was observed in the P288E mutant in both the absence and presence of activators. Substitution of the E304 residue also resulted in changes in allosteric behavior with the largest effect being desensitization to both activators and inhibitors. The E304D mutant displayed a higher Vmax (~2.5‐fold) in the absence of activators and it was activated by FBP, a gain of function compared to wild type (WT). Future studies include the characterization of E304N and the generation and characterization of K310A (in the presence and absence of P288D) as this residue may be forming a salt bridge with the introduced aspartate residue. Efforts to crystallize the P288D enzyme are underway to compare to the WT structure.

Grant Funding Source: Supported by NSF award 0448676
Everyone Needs Folic Acid is a digital program with face and content validity for low‐income diverse audiences (807.4)
Authors: Cheng Feon , Wamboldt Patricia , Lohse Barbara
Published: June 2021

Everyone Needs Folic Acid, a nutrition program delivered with a digital photo frame platform, was developed to help low‐income persons appreciate the role of folic acid (FA) for all age groups. Program assessment was conducted in 2 stages: Stage 1 focused on the original program and Stage 2 evaluated the subsequent program revisions. Participants (n=73) were white (97%), overweight or obese (43%), not eating competent (73%), and had at least 2 children (53%). The sample was low‐income; 42% used at least 1 assistance program in the past year and 27% were SNAP participants. The original program was easy to read (90%), understandable (87%), and helpful (66%). Suggested improvements were to provide dosage information, increase pictures of folic acid rich foods, and reduce message redundancy. Stage 2 participants (n=50) were recruited from the Stage 1 sample. When asked if they recalled any information about the original program 27% of Stage 2 respondents remembered a lot, 45% remembered some, and 28% reported little or no information recall. 72% had applied information to themselves or their family and 96% indicated that they would recommend this program to others. Participants compared 6 original with revised slides and preferred the original in 2 pairs, the revised slide in the other 4. Everyone Needs Folic Acid demonstrated face and content validity for SNAP‐eligible persons. Funded by USDA, SNAP Ed.

Grant Funding Source: USDA, SNAP Ed
Association between serum and brain fatty acid profiles in centenarians (821.2)
Authors: Mohn Emily , Vishwanathan Rohini , Lichtenstein Alice , Matthan Nirupa , Poon Leonard , Johnson Mary Ann , Hausman Dorothy , Davey Adam , Green Robert , Gearing Marla , Woodard John , Nelson Peter , Miller L , Johnson Elizabeth
Published: June 2021

Profiles of brain fatty acids (FA) may be important indicators of age‐related cognitive decline. There is limited data on brain FA profiles in centenarians. Serum FA may be related to brain FA profiles. The goal of this study was to test this hypothesis in centenarians. Tissue from the frontal, occipital, temporal cortices, and cerebellum were collected from decedents (n=40) in the Georgia Centenarian Study with available banked serum. Brain and serum FA were measured using gas chromatography. FA profiles were different between serum and brain (P<0.0001). On average, saturated FA (SFA) were predominant in the brain (47 mol%), followed by monounsaturated FA (MUFA) (23 mol%), n‐3 polyunsaturated (n‐3) FA (14 mol%, with 91% being docosahexaenoic acid), n‐6 polyunsaturated (n‐6) FA (12 mol%), and trans FA (0.7 mol%). N‐6 FA were highest in serum (38 mol%), followed by SFA (30 mol%), MUFA (25 mol%), n‐3 FA (3 mol %), and trans FA (3 mol%). Serum n‐3 FA were related to brain levels in the frontal (r=0.43, P<0.01) and temporal (r=0.35, P<0.05) cortices and cerebellum (r=0.41, P<0.05). Serum n‐3/n‐6 was related to brain n‐3/n‐6 in all regions (frontal: r=0.41, occipital: r=0.38, P<0.05; cerebellum: r=0.47, temporal: r=0.55, P<0.01). No other FA were related between serum and brain. The data suggest that both n‐3 FA and n‐3/n‐6 FA can be used as markers of brain fatty acid profiles.

Grant Funding Source: Supported by: USDA (58‐1950‐0‐014), NIH (1P01‐AG17553), and Abbott Laboratories
Role of metabolism and intracellular trafficking in fructose‐induced GLUT5 regulation (822.4)
Authors: Patel Chirag , Douard Veronique , Tharabenjasin P , Topaktas E , Ferraris Ronaldo
Published: June 2021

Marked increases in fructose consumption have been linked to various diseases. Fructose (F) upregulates its own absorption by increasing the expression & activity of the intestinal F transporter GLUT5, but the underlying mechanism is not well understood. Defects or limits in regulation can lead to adult‐onset F intolerance or to F malabsorption in infants. I tested the hypothesis that F transport via GLUT5, metabolism via ketohexokinase (KHK), and GLUT5 intracellular trafficking to the apical membrane via the GTPase Rab11a are required for GLUT5 upregulation. WAdult wildtype (wt), GLUT5‐/‐, and KHK‐/‐, and Rab11a‐/‐ mice were receivedreceiving 30% glucose (G), fructose (F) or lysine (L) by gavage twice a d for 3 d at X.X ml/kgays. 17 d old wt and Rab11a‐/‐ mice were gavaged with either 30% G or F. Afterwards, GLUT5 activity as well as mRNA and protein expression determined in the small intestine. Activity and expression of GLUT5 and of other enzymes involved in F metabolism increased in F‐gavaged mice compared to those gavaged with G or L. This effect was specific because expression of unrelated transporters like SGLT1 did not change. Strikingly, F failed to induce the expression & activity of GLUT5 and of other enzymes involved in F metabolism, in KHK‐/‐, GLUT5‐/‐ and Rab11a‐/‐ mice. Thus, I have demonstrated for the first time the essential roles of membrane transport, metabolism, and Rab11a‐mediated trafficking in F‐ induced GLUT5 upregulation.

Grant Funding Source: NSF IOS‐1121049.
Radiofrequency renal nerve denervation decreases blood pressure, sympathetic activity and renal norepinephrine content in spontaneously hypertensive rats (875.16)
Authors: Gao Juan , Kerut Edmund , Smart Frank , Kapusta Daniel
Published: June 2021

Radiofrequency renal denervation (RF‐RDN) decreases blood pressure (BP) in patients with drug‐resistant hypertension. This study investigated the effects of RF‐RDN on sympathetic activity and BP (Systolic, SBP; Diastolic, DBP) in SHR. Methods Nineteen‐week old male SHR were instrumented with telemetry probes for chronic measurement of BP and BP variability (BPV). After 1‐week, control BP was measured in rats for 3 days. The next day SHR (n=6/group) were anesthetized and randomly received either bilateral Sham‐RDN or RF‐RDN of the renal arteries (Biosense Webster Stockert 70 generator and RF‐probe). BP was then measured in SHR for 8 weeks. Results Following RF‐RDN, SBP/DBP was significantly decreased the day after (153±8/102±4 mmHg) and throughout the post‐RDN study (4‐week SBP/DBP, 173±7/120±5; 8‐weeks, 171±6/116±4 mmHg) compared to pre‐RDN control levels (SBP/DBP, 187±9/130±5 mmHg). RF‐RDN significantly decreased the low frequency component of BPV in SHR with the peak attenuation observed at 8 weeks (LF/HF 2.2±0.2 vs 0.9±0.2); this correlated with a blunting of the hypotensive response to i.p. chlorisondamine in RF‐RDN rats. Finally, at the end of week 8, kidney norepinephrine (NE) levels in RF‐RDN rats (left, 10±10; right, 0.0±0.0 ng/g kidney) were dramatically decreased compared with levels from sham SHR (left, 130±10; right, 110±10 ng/g kidney). Conclusions These data demonstrate that in hypertensive SHR, bilateral RF‐RDN of the renal arteries decreases BP in part by chronic inhibition of sympathetic neural pathways and a decrease in renal tissue NE levels.

Grant Funding Source: Biosense Webster grant IIS‐175 to DRK and FS; NIH NIGMS P20 GM103514 to DK
Electrocardiographic markers for cardiac hypertrophy in mice bred for high voluntary wheel running (881.11)
Authors: Kay Jarren , Hampton Thomas , Garland Theodore
Published: June 2021

The left ventricle becomes enlarged during cardiac hypertrophy, caused by either physiological or pathological factors. Electrocardiogram (ECG) recordings can provide insights into cardiac hypertrophy. We used 100 mice from an ongoing experiment which breeds for high voluntary wheel running. ECGs were recorded before and after 6 days of wheel access using an ECGenie (Mouse Specifics, Quincy, MA). After wheel access, and adjusting for body mass, mice from the four replicate High Runner lines had significantly lower heart rates than those from the four non‐selected Control lines (650 ± 30 bpm vs. 709 ± 14 bpm, LSM ± SE, p<0.05); females had a shorter PQ interval duration than males (19.2 ± 0.3 ms vs. 20.9 ± 0.8 ms, p<0.05). A subset of the HR mice are homozygous for a recessive mutation that causes the mini‐muscle (MM) phenotype, characterized by a ~50% reduction in hindlimb muscle mass and larger hearts than wild types. MM mice had longer PQ intervals after wheel access, and a longer QRS complex compared to Control both before (11.1 ± 0.3 ms vs. 10.2 ± 0.1 ms, p<0.05) and after wheel access (11.2 ± 0.3 ms vs. 10.5 ± 0.1 ms, p<0.05). The longer QRS duration in the MM mice before wheel access points to left ventricular hypertrophy, but whether this is a physiological adaptation to selective breeding or a pathological state requires further study. MM ventricular hypertrophy may be a useful model of cardiac enlargement in human athletes.

Grant Funding Source: NSF IOS‐1121273
Sex differences in the ability of high fat diet to inhibit glucose‐induced modulation of 5‐HT3 receptor responses in GI‐vagal afferents in rats (905.4)
Authors: Troy Amanda , Browning Kirsteen
Published: June 2021

Previous studies in this lab have shown that extracellular glucose modulates 5‐HT3 receptor (5‐HT3R) mediated currents GI vagal afferent neurons and that this response is lost following exposure to a high fat diet (HFD). The aim of this study was to investigate whether this loss of glucose‐induced modulation showed any sex dependent differences.

Sprague‐Dawley rats were placed on HFD (60% kcal from fat) at 28 days. Whole cell patch clamp recordings were made subsequently from acutely dissociated identified GI‐projecting vagal afferent neurons. The ability of extracellular glucose (2.5‐10mM) to modulate the magnitude of the 5‐HT‐induced inward current was assessed.

After 4 weeks HFD exposure, glucose modulated the 5‐HT‐induced response in 7/8 neurons (87%) from female rats. In contrast, glucose modulated the 5‐HT‐induced inward current in only 4/7 neurons (57%; 4 weeks) from male rats (P<0.2). At this time point, there was no difference in body weights (male: 189±12g vs female: 168±5g; P>0.05) although male rats had significantly higher blood glucose levels (363±13mg/dl vs 251±7mg/dl; P<0.05). These data suggest that, following HFD (i) male rats may be more susceptible to loss of glucose‐induced modulation of 5‐HT‐induced currents in GI vagal afferent neurons and (ii) this loss of glucose‐induced modulation is determined primarily by dysregulated glycemic control rather than diet or obesity.

Grant Funding Source: NIH DK078364, NSF IOS1148978
Substrate recognition and processing by the eukaryotic 26S proteasome (936.3)
Authors: Bashore Charlene , Martin Andreas
Published: June 2021

The 26S proteasome degrades a multitude of substrates and is essential for eukaryotic cell survival. The 2.5 mega‐dalton protease processes substrates through its 19S regulatory particle: this intricate complex binds, unfolds, and feeds substrates into the proteasomal peptidase, where they are irreversibly degraded. Although the mechanism of proteasome proteolysis has been well characterized, little is known about how the processing steps prior to degradation culminate to give an overall degradation rate. The complexity of substrate processing and the requisite of proteasome function for cell viability pose a significant challenge to quantitative analysis. I have developed an in vitro system to measure degradation and deubiquitination rates as a function of substrate thermodynamic stability and lysine placement. These tools lend insight on the rate limiting step in substrate degradation and how polyubiquitin tags are recognized and processed by this intricate proteolytic machine.

Grant Funding Source: Supported by the National Science Foundation
Messenger RNAs and cognate microRNAs in the nucleolus (940.3)
Authors: Reyes‐Gutierrez Pablo , Politz Joan , Pederson Thoru
Published: June 2021

During studies of muscle cell differentiation in a rat myogenic cell line we discovered, to our surprise, that several microRNAs are concentrated in the nucleolus (Politz et al., PNAS 103: 18957‐18962, 2006; RNA 15:1705‐1715, 2009). Since these nucleolus‐localized microRNAs were also present in the cytoplasm, it seemed plausible that their association with nucleoli represented a station on their nuclear export pathway. These findings prompted us to ask whether certain messenger RNAs might also localized in the nucleoli of these cells. Microarray analysis of RNA from highly purified nucleoli revealed a set of mRNAs present at 30 to 650‐fold higher concentrations than in the nucleoplasm and this was confirmed by in situ hybridization for one of these mRNAs, that encoding insulin‐like growth factor 2 (IGF2), an well‐known autocrine regulator of myoblast proliferation and differentiation. RT‐PCR experiments demonstrated that these nucleolus‐localized IGF2 transcripts are spliced mRNAs, suggesting that they arrive from distal sites in the nucleoplasm. Bioinformatics analysis predicted stable interactions between all of the nucleolus‐localized microRNAs and the 5′ or 3′ untranslated regions of IGF2 mRNA. Additional considerations suggest that at least one of the nucleolus‐localized microRNAs, miR‐206, may be involved in the positive regulation of IGF2 mRNA translation and we speculate that transit through the nucleolus commits IGF2 mRNA to this mode of regulation.

Grant Funding Source: NSF MCB‐1051398
A comparative study of xanthine dehydrogenase regulator proteins in Agrobacterium tumefaciens and Streptomyces coelicolor (946.3)
Authors: Sivapragasam Smitha , Grove Anne
Published: June 2021

A. tumefaciens and S. coelicolor are among the most studied bacteria with the former being the 3rd most important plant pathogen. The transcription factor XdhR of S. coelicolor (ScXdhR) regulates its own gene expression along with the divergently oriented xanthine dehydrogenase ABC (xdh) gene cluster. A similar genomic locus was observed in A. tumefaciens. Xanthine dehydrogenase converts hypoxanthine and xanthine to urate. Based on elecrophoretic mobility shift assays we show that ScXdhR and XdhR of A. tumefaciens (AtXdhR) bound to their respective intergenic regions specifically but with different binding affinities. The ligands GMP, rGTP and xanthine compromised the DNA‐XdhR binding in both cases. The ScXdhR‐DNA complex also dissociated upon addition of H2O2, cumene hydroperoxide and tert‐butyl hydroperoxide, whereas DNA binding by AtXdhR was unaffected. SDS‐PAGE analysis revealed that ScXdhR dimerized upon addition of these oxidants, whereas AtXdhR did not. AtXdhR dimerized in the presence of Co2+ whereas ScXdhR did so in the presence of Cu2+ as well as Co2+. This study suggests that both XdhR proteins may regulate expression of the xdh gene cluster in response to intermediates in the purine degradation pathway. Notably, ScXdhR also responds to oxidation, suggesting upregulation of xdh during oxidative stress.

Grant Funding Source: NSF