bluuebunny/crossref_metadata_2025 huggingface dataset:
Older Americans Act Nutrition Program Improves Participants' Food Security in Georgia: Georgia Advanced Performance Outcomes Measures Project (GA POMP)
Authors: Lee Jung Sun , Johnson Mary Ann , Brown Arvine
Published: June 2021

With population aging and the current economic recession, it is critical to use convincing research methodology to demonstrate the benefits of nutrition assistance programs targeted to nutritionally vulnerable older adults. We examined the impact of the Older Americans Act Nutrition Program (OAANP) participation on food security and nutrition risk by using the longitudinal data from the ongoing GA POMP. The sample included 717 older Georgians who began OAANP participation (37.1%) and those added to program waitlists (62.9%) between July and early November, 2008 and completed all study variables in two waves of self‐administered mail surveys conducted 4 months apart (mean age 74.6±9.5, 70.9% female, 33.2% black). At baseline, most of the sample were food insecure (53.8%) and at high nutrition risk (74.9%). Waitlisted people reported higher levels of persistent food insecurity (45.9%) or becoming food insecure (10.0%) than the participants (29.3% and 7.1%, respectively) over the 4 month period. While considering potential confounding factors, the estimated odds of achieving food security were 1.69 times higher in participants than in waitlisted people in the 4 month period. This significant reduction in food insecurity suggests that this easily administered and reliable measure of food security can detect the benefits of OAANP participation in as little as 4 months. This study was funded in part by US AoA and USDA.
Use of blood mRNA levels as molecular biomarkers to determine dietary selenium requirements in rats
Authors: Sunde Roger A. , Evenson Jacqueline K.
Published: June 2021

To study use of blood glutathione peroxidase‐1 (Gpx1) mRNA to assess selenium (Se) status and requirements, male weanling rats were fed graded levels of Se as selenite for 28 d. Liver Se and plasma Gpx3 and liver Gpx1 activities indicated that the minimal Se requirement was 0.08 μg Se/g diet. In whole blood mRNA, Gpx1 mRNA in Se‐deficient rats decreased to 10% of levels in Se‐adequate (0.2 μg Se/g) rats. With Se supplementation, blood Gpx1 mRNA levels increased sigmoidally to a plateau with a minimum Se requirement of 0.08 μg Se/g, whereas Gpx4 mRNA levels were unaffected. The preponderance of whole blood Gpx1 mRNA arose from erythroid cells. When rats were bled (1.5% BW) by cardiac puncture at 25 d and killed at 28 d, blood hemoglobin and hematocrit decreased, reticulocyte count increased, but leukocyte counts, plasma Gpx3 activity, and RBC Gpx1 activity were not altered. Gpx1 and Gpx4 mRNA both increased over 200% with bleeding whereas Gapdh mRNA decreased, thus discounting Gapdh mRNA as a control mRNA. β‐globin (Hbb) mRNA levels increased with bleeding and were not affected by Se status; Gpx1 mRNA levels normalized to Hbb mRNA in bled and sham‐bled rats, however, were similarly regulated by Se status, indicating that use of appropriate control mRNAs has potential to allow accurate assessment Se status independent of modest changes in erythrocyte count. USDA 95‐37200‐1799 and UW and Selenium Nutrition Research Fund.
Selenoprotein W regulates tumor suppressor p53 in prostate epithelial cells
Authors: Hawkes Wayne Chris , Printsev Ignat , Alkan Zeynep
Published: June 2021

The anticancer activity of selenium has been demonstrated in myriad animal and in vitro studies, but the mechanisms remain obscure. Selenoprotein W (SEPW1) is a highly conserved protein of unknown function ubiquitously expressed in animals, bacteria and archea that modulates control of cell cycle entry. The tumor suppressor protein p53 is a redox‐active transcription factor that is activated by and responds to various types of stresses with apoptosis, cell cycle arrest, senescence, DNA repair, cell metabolism changes, or autophagy. TP53 is the most frequently mutated gene in human cancers. We set out to examine the functional and biochemical interactions between SEPW1 and p53. SEPW1 silencing increased p53 protein levels by decreasing p53 turnover, suggesting SEPW1 promotes cell cycle progression by destabilizing p53. Silencing SEPW1 greatly increased expression of p21(Cip1/WAF1), the cyclin‐dependent kinase inhibitor responsible for p53‐mediated cell cycle arrest. Western blots did not show any difference in phosphorylation of p53 on Ser15, suggesting a non‐canonical mechanism of p53 regulation. SEPW1 silencing increased mRNAs of stabilizers of p53 (LATS2, GMCL1, TGFB1, ZNF148, YWHAE) and decreased mRNAs of destabilizers of p53 (PIM3, RCHY1, HSPB1), suggesting several possible mechanisms for regulation of p53 by SEPW1. Supported by USDA CRIS Projects 5306‐51530‐014‐00D and 5306‐51530‐018‐00D.
A High Protein Diet Induces Intestinal Iron Transporter Expression and Improves Iron Absorption in Rats
Authors: Cucchi Carrie , Gaffney‐Stomberg Erin , Sun Ben‐Hua , Kerstetter Jane , Insogna Karl
Published: June 2021

Dietary protein has been shown to increase iron (Fe) absorption by improving bioavailability. Little data are available on the effect of dietary protein on Fe transporter expression.

As part of a study designed to evaluate the impact of dietary protein on mineral homeostasis, a whole genome microarray screen was performed using RNA prepared from duodenal intestinocytes of rats ingesting low (5%) or high (40%) casein diets for one week. Of 91 genes that were significantly up‐regulated in the 40% group, 3 were Fe transporters. DMT1 expression was 2.5‐fold greater in the 40% compared to the 5% group, while Cybrd1 and Tfrc were 1.6‐fold greater. Q‐PCR using the same RNA employed in the microarray screen confirmed these findings.

When compared to the level of expression on a 20% diet (usual rat chow), DMT1 was 4‐fold higher in the 40% group (p<0.001), Cybrd1 was 2.2‐fold higher (p<0.05) while Tfrc was not different.

To determine if the observed changes in expression of DMT1 and Cybrd1 were associated with changes in Fe homeostasis, a balance study was performed in rats ingesting 5, 20 and 40% protein diets. By the Kruskal‐Wallis test there was a significant effect of dietary protein on Fe balance and by post hoc analysis Fe absorption was greater in animals consuming the 40% vs. 5% diet (p<0.01).

These data suggest that increasing dietary protein augments Fe absorption in part by inducing the expression of duodenal Fe transporters.

Grant Funding Source: USDA
Characterization of plasma ceruloplasmin activity and expression following dietary copper deficiency
Authors: Prohaska Joseph , Broderius Margaret , Mostad Elise , Wendroth Krista
Published: June 2021

Ceruloplasmin (CP) is a multicopper oxidase and the most abundant copper binding protein in mammalian plasma. Loss of function mutations in humans or experimental deletion in mice result in iron overload consistent with CP's putative ferroxidase function. Prior work suggested plasma may contain multiple ferroxidases. Studies were conducted in Holtzman rats, albino mice, Cp −/− mice, and adult humans to investigate the copper‐iron interaction and further characterize CP as a biomarker. Dietary copper‐deficient (CuD) rats and mice were produced using a modified AIN‐76A diet. Results confirmed that o‐dianisidine is a better substrate than paraphenylene diamine (PPD) for assessing diamine oxidase activity of Cp. Plasma from CuD rat dams and pups, and CuD and Cp −/− mice contained no detectable Cp diamine oxidase activity. Importantly, following western blotting no detectable ferroxidase activity was observed for CuD rats, mice, or Cp −/− mice compared to robust activity for copper‐adequate (CuA) rodent controls. Immunoblot protocols detected major reductions (60–90%) in Cp protein in plasma of CuD rodents but no alteration in liver mRNA levels by qRT‐PCR. Data support that apo‐Cp is less stable than holo‐Cp. Further research is needed to explain normal plasma iron in CuD mice. Reduction in CP activity is a sensitive biomarker for copper deficiency.

Supported by NIH HD‐39708 and USDA NIFA 2006‐35200‐17378.
How the family started: Origins of hominids
Authors: Ward Carol
Published: June 2021

The transition to committed bipedality is a key characteristic of hominins. Fossil evidence documenting hominin origins has grown substantially, but much of our understanding of the earliest members of the Australopithecus‐human clade still comes from Au. afarensis. Understanding selective pressures shaping Australopithecus requires knowledge of its ancestral morphology. Here I compare evidence from Miocene and Pliocene hominoids and consider the implications of their morphology for reconstructing the ancestral hominin morphology. Data indicate high levels homoplasy among hominoids, and that ancestral hominins likely differed from extant great apes in important ways, such as pelvis, lumbar and hand proportions. However, some characteristic of extant apes are likely primitive. Homoplasy is common and involved different experiments in exploiting arboreal habitats. So rather than thinking about progressive stages leading towards Australopithecus from an ancestral taxon like Proconsul through one like a modern great ape or not, we need to refocus efforts to explore the detailed similarities and differences among extant and extinct apes, and the diversity of their locomotor adaptations.

Research supported by NSF and LSB Leakey Foundation.
Contribution of polyphenols to the total antioxidant capacity of pomegranate and grape juices
Authors: Bolling Bradley Warren , Blumberg Jeffrey B. , Chen C.‐Y. Oliver
Published: June 2021

Pomegranate juice (PJ) and grape juice (GJ) have comparable ORAC assay values, but PJ is more potent than GJ in other in vitro measures of Total Antioxidant Capacity (TAC). We examined the basis for these differences in the ORAC, FRAP, and DPPH assays. Juices were fractionated into their sugar, organic acid, neutral phenol, and anthocyanin components to assess the contribution of each to their TAC. Polyphenol fractions contributed ≥75% of the TAC for both juices. For PJ, neutral phenols contributed the most to ORAC, FRAP, and DPPH values with 50, 49, and 46% of TAC of the whole juice, respectively; GJ anthocyanins contributed the most with 53, 47, and 49%, respectively. The contribution of PJ polyphenols to FRAP values was 56% greater than GJ. The contribution of PJ polyphenols to ORAC values were 42% less than GJ. The ORAC/FRAP ratio of GJ neutral phenols and anthocyanin fractions were ≥1.5‐fold higher than those of PJ fractions, while ORAC/DPPH ratios were 3.5‐fold larger. Fraction recombination revealed interactions were dependent upon the assay. Sugars and organic acids antagonized PJ polyphenols in the DPPH assay by 50% and the GJ ORAC value by 22%. Therefore, the differential TAC of PJ and GJ can be attributed to the larger intrinsic electron transfer capacity but smaller hydrogen transfer capacity of PJ phenols and anthocyanins than GJ. Supported by the USDA and POM Wonderful LLC. BWB was supported by NIGMS K12GM074869.
Absorption kinetics of polyphenols and alteration of antioxidant performance in older adults after ingesting Angelica keiskei
Authors: Switkowski Karen M , Camacho Camila Renata Correa , Chen Oliver , Rasmussen Helen , Aldini Giancarlo , Chun HK , Cho Soomuk , Park KM , Russell Robert , Saltzman Edward , Blumberg Jeffrey , Yeum Kyung‐Jin
Published: June 2021

Angelica keiskei (AK) is a leafy green vegetable rich in phytochemicals such as carotenoids (C) and flavonoids (F). To determine the absorption kinetics of polyphenols in AK and the alteration of antioxidant performance in vivo, older adults (>60 y) with metabolic syndrome (MetS, n=5) and healthy controls (HC, n=5) consumed 5 g dry AK powder encapsulated in gelatin capsules with a low C & F liquid diet. Plasma samples were collected at baseline, 30 min, 1, 2, 3, 4, 5, 6, 7, and 8 h and analyzed for C and F using HPLC with UV and EC detection, respectively, for Total Antioxidant Performance (TAP) by fluorometry, and for DNA damage using single cell gel electrophoresis (Comet assay). After ingesting AK, increases in plasma quercetin were observed at 1–3 and 6–8 h in the HC and at all time points in the MetS groups compared to baseline (p<0.05). Plasma lutein was elevated in the HC group at all time points and at 6–8 h in the MetS group compared to baseline (p<0.05). Increase in TAP were observed at 1, 3, 6, and 7 h in the HC group and at 1 and 5h in the MetS group compared to baseline (p<0.05). Baseline DNA damage was more than 10% higher in the MetS compared to the HC group (p<0.05). Thus, phytochemicals in AK are bioavailable and can increase antioxidant performance in both HC and MetS subjects. [Supported by USDA ARS #1950‐51000‐065‐08S, USA and BioGreen 21 #20070301034009 RDA, Korea]
Association of age, gender and weight status with willingness to taste fruits and vegetables among elementary school children
Authors: Lakkakula Anantha , Wong Wei‐Ping , Zanovec Michael , Pierce Sarah , Tuuri Georgianna
Published: June 2021

This study examined the association between age, gender, and weight status with willingness to try fruits and vegetables offered during a tasting program. A total of 312 first, third, and fifth graders (51% boys, 34% first graders, and 30% third graders) attending low‐income public elementary schools in southeastern Louisiana participated. Prior to tasting, children were measured for height and weight. Small portions of 4 vegetables (carrots, peas, tomatoes and bell peppers) and 4 fruits (apricots, cantaloupe, peaches and pears) were offered. The children recorded whether they “chewed and swallowed”, “spit it into the napkin” or “did not taste” each item. Scores for vegetables and fruits were examined separately. A logistic regression analysis indicated that first graders were 4.6 times more willing to taste 2 or more vegetables provided when compared to fifth graders (OR = 4.6, 95% CI: 2.31, 9.14; p < 0.01), and third graders were 8.1 times more willing to taste 2 or more vegetables provided when compared to fifth graders (OR = 8.1, 95% CI: 3.65,18.22; p < 0.01). No differences were observed between first and third graders, gender or weight status group and willingness to try vegetables. No associations were found between willingness to try fruits and age, gender or weight group. Younger elementary school children were more willing to try vegetables than older children. Supported by USDA grant #2009‐35200‐05287.

Grant Funding Source: USDA grant #2009‐35200‐05287
Green tea EGCG suppresses specific antigen driven T cell response and affects CD4+ T cell differentiation
Authors: Pae Munkyong , Ren Zhihong , Meydani Simin Nikbin , Wu Dayong
Published: June 2021

T cells, in particular CD4+ T helper (Th) cells, play a key role in mediating many aspects of autoimmune diseases. Th1 and the recently discovered Th17 cells are pro‐inflammatory subsets of Th cells, which are responsible for inducing autoimmunity, whereas regulatory T cells (Treg) have a protective effect. Green tea ingredient epigallocatechin‐3‐gallate (EGCG) has been shown to improve symptoms and reduce pathophysiological changes in some animal models of autoimmune diseases. However, the underlying mechanism is not well elucidated, in particular, involvement of Th17 and Treg has not been studied. In this study we found that EGCG inhibited proliferative response of both normal and autoreactive T cells to their specific antigens. This effect was due to its inhibitory effect on both T cells and antigen presenting cells, the former being more significant. EGCG inhibited Th1 but had no effect on Th2 response. Furthermore, EGCG impeded Th1 and Th17 differentiation but promoted Treg development from naïve CD4+ T cells. In the presence of transforming growth factor‐β, interleukin‐6 promoted Th17 and suppressed Treg development, which was diminished by EGCG. Natural Treg population was not affected by EGCG. Together our results indicate that EGCG inhibits Th1 and Th17 differentiation and has no effect on Th2 cells, while it enhances Treg development. These findings help us understand the underlying mechanisms of EGCG‐afforded protection of autoimmune diseases reported in animal models. Future studies are warranted to delineate the impact of EGCG on the regulatory network that controls CD4+ T cell differentiation. Supported by USDA #58‐1950‐7‐707.
Maturity aggravates sepsis‐associated skeletal muscle catabolism in growing pigs.
Authors: Orellana Renán A , Suryawan Agus , Gazzaneo María C , Wilson Fiona A , Nguyen Hanh V , Fiorotto Marta L , Davis Teresa A
Published: June 2021

Synthesis and accretion of muscle protein is elevated in neonates and decreases with development. During sepsis, muscle protein synthesis is reduced, but the effect of development on the metabolic response to sepsis in skeletal muscle is not well understood. Fasted 7‐ and 26‐d‐old pigs were infused for 8 h with LPS (0 and 10 μg•kg−1•hr−1) and fractional protein synthesis rates (FSR) and translation and degradation signals were determined in longissimus dorsi muscle (n=5–7/group/age). α‐Actin in serum, and FSR, ribosomal abundance, and caspase‐3 abundance in muscle were greater in 7‐ than 26 d‐old pigs, indicating a higher baseline protein turnover in the younger pigs. In muscle, LPS decreased FSR, protein synthetic efficiency, and eIF4G phosphorylation and increased AMPK phosphorylation and MURF‐1 and atrogin‐1 abundance in both age groups. Serum 3‐methylhistidine and α‐actin levels and caspase‐3 abundance in muscle increased in 26‐ but not in 7‐d‐old pigs in response to LPS. The results suggest that the mechanisms regulating protein accretion in skeletal muscle in neonatal pigs during sepsis are influenced by development such that the LPS‐induced muscle catabolism becomes more severe as maturation advances. NIH AR51563, NIH AR44474, and USDA 6250‐51000‐040

Grant Funding Source: NIH AR44474, and USDA 6250‐51000‐040
Glycemic index does not affect overall glucagon like peptide – 1 levels in healthy overweight women
Authors: Casey Shavawn Marie , Horn William , Hembrooke Tara , Keim Nancy
Published: June 2021

Background

Glucagon‐like peptide‐1 (GLP‐1) is secreted in response to food ingestion specifically carbohydrate‐rich food sources. GLP‐1 is involved in a variety of physiologic functions including: regulation of plasma glucose, inhibition of gastric emptying, and decreasing food intake by increasing satiety.

Methods

A randomized cross‐over design was used to measure the incretin response to 2 standardized breakfast meals: a high glycemic index (HGI) meal and a low glycemic index (LGI) meal. Both meals were matched for energy and macronutrient contents (54% C, 15% P, & 31% F). Seven healthy, overweight women participated. Blood samples were obtained before and for 8 hours following consumption of the test meals. Plasma GLP‐1 [(7–36) and (7–37) amide] quantification was done by active ELISA assay.

Results

There were no differences in fasting plasma GLP‐1 or in postprandial GLP‐1 in response to the HGI and LGI breakfast meals.

Grant Funding Source: general funds and the USDA, ARS WHNRC

Grant Funding Source: USDA
Maternal Low Protein Diet Induces ATF3 but not ASNS Expression in Liver of Female Offspring Through Amino Acid Response (AAR) Pathway
Authors: Zhou Dan , Pan Yuan Xiang
Published: June 2021

Objective

To investigate the impact of maternal low protein on programming of AAR pathway.

Method

Sprague‐Dawley rats were fed a control (18% protein) or low protein diet (LP, 9% protein) during gestation. Liver from female offspring was collected at 38 days of age to analyze expression and histone modification of genes in AAR pathway.

Result

expression of ATF3 in LP group increased significantly. ATF4 and p‐eIF2alpha, key upstream factors in AAR pathway, were up‐regulated as well. Chromatin immunoprecipitation assay confirmed that Pol II and ATF4 binding was increased at the ATF3 promoter while there was a decrease of ATF3 and C/EBPbeta binding. On the other hand, promoter of the downstream gene ASNS showed a different profile. Acetylated histone H4, a marker for an open chromatin was increased at the ASNS promoter, along with an increased binding of ATF4, ATF3 and C/EBPbeta. However, tri‐methyl histone H3 at lysine 9, a marker for chromatin silencing, was increased in LP group, possibly leading to no change in Pol II binding and therefore no change in ASNS gene expression level.

Conclusion

Maternal low protein diet programs AAR pathway in liver of female offspring. This programming occurs at upstream of AAR pathway, mainly affecting gene expression of transcription factors, including ATF4 and ATF3. Furthermore this may prime the downstream target genes such as ASNS to respond to low protein environment later.

Grant Funding Source: USDA
How Does the 2007 U.S. Food Supply (FS) Relate to Federal Dietary Guidance?
Authors: Post Robert , Hiza Hazel A.B. , Bente Lisa
Published: June 2021

Per capita FS estimates are useful to monitor the potential of the FS to meet the nutritional needs of Americans. The Dietary Guidelines for Americans (DGA) and MyPyramid help consumers choose diets to improve nutritional status. This study analyzed the amount of food available in the FS for per capita per day consumption for each food group, and compared compliance with the 2005 DGA and MyPyramid. FS data were converted into daily per capita food servings comparable with recommendations in the DGA and the MyPyramid intake patterns using a multistage process. The per capita food available for the grains and meats groups met and exceeded the MyPyramid recommendations for the 1,600 and 2,000 calorie levels. The 2,000 level was 28 and 22%, respectively, for grains and meats. However, the average number of food servings for fruits, vegetables and milk groups fell short by 55, 28, and 40%, respectively, for the 2,000 level. None of the food servings met the 2,800 level. The FS servings measure changes in food consumption over time using MyPyramid as a dietary assessment tool. Federal dietary recommendations are key to Federal efforts to monitor the dietary and nutritional status of the population. Source of support: USDA.
Persistence of Lactobacillus Reuteri DSM17938 in the Human Intestinal Tract
Authors: Smith Tracey J , Anderson Danielle , Sikes Anthony , Margolis Lee M , Young Andrew J.
Published: June 2021

Probiotics may enhance gastrointestinal health and immune function. The efficacy of different dosing strategies on colonization and persistence of probiotics are undefined. We assessed colonization and persistence of Lactobacillus reuteri (L. reuteri) (BioGaia, Stockholm, Sweden) after daily or alternate day dosing. Volunteers ate pudding with L. reuteri (109 CFU) daily (DAILY; n = 9) or on alternate days (ALT; n = 9) over 7 days. Fecal samples were collected on dosing days (D1–7) and after dosing ended (D13–15 and D20–22), and analyzed for L. reuteri. Results are reported in 3‐day increments (D2–4, D5–7, D13–15, and D20–22). L. reuteri count rose in DAILY ([mean ± SD] D2–4: 4 × 104 ± 2 × 104CFU, p < 0.01; D5–7: 10 × 104 ± 9 × 104CFU, p < 0.01) and ALT (D2–4: 21 × 104± 20 × 104CFU, p < 0.01; D5–7: 11 × 104± 15 × 104CFU, P = 0.06), and fell in both groups 1 week after dosing ended (p < 0.01). Total volunteers with detectable L. reuteri 1 and 2 weeks after dosing ended was similar for DAILY (4/9 and 2/9, respectively) and ALT (3/9 and 2/9, respectively). L. reuteri count was higher D2–4 for ALT vs. DAILY (p < 0.05), but similar thereafter. Alternate day probiotic intake achieves equivalent colonization to daily intake, but colonization declines rapidly once dosing stops. Initially, it appears that responsiveness to probiotics may differ between individuals, but those differences do not persist with longer consumption. Funded by MRMC.
The effects of dietary antioxidants and age on hepatic CRP levels in rodents
Authors: Hasan Syeda Tahira , Kong Eun Hee , Guo Weimin , Wu Dayong , Wang Yan‐Lee , Veeramachaneni Sudipta , Wang Xiang‐Dong , Meydani Mohsen
Published: June 2021

Dietary antioxidants may counter inflammation associated with many diseases like CVD and cancer. C‐reactive protein (CRP) is a well‐established marker of systemic inflammation. Using ELISA, we examined CRP levels in liver samples of young and old animals and animals fed with high fat diet or alcohol diet supplemented with antioxidants. Rats treated with high fat liquid (LDC) diet (71% energy from fat) for 6 wks had NASH and increased hepatic CRP levels by 53%, as compared to control (35% energy from fat). Rats fed with liquid (LDC) alcohol diet (36% energy from alcohol) with lycopene supplementation (3.3 mg/Kg BW/d) for 11 wks increased hepatic CRP levels by 2 fold (56.8 ± 22.9 vs. 28.0 ± 7.2 pg/μg protein), as compared with rats fed control diet supplemented with lycopene. In addition, 26 mo old C57BL/6 mice had CRP levels 2.3 fold higher than young (6 mo) mice. Supplementing mice with a mixture of tocopherol/tocotrienol (provided 500 ppm vit. E/kg diet) for 6 wks significantly reduced CRP levels by 30% and 40% in young and old mice, respectively. We conclude that hepatic CRP increase with age and with consumption of high fat diet. While a mixture of tocopherol appears to attenuate CRP levels, high dose of lycopene supplementation augments hepatic CRP in livers of rats fed alcohol. The latter indicates potentially harmful effects of consuming high amounts of both alcohol and lycopene. Supported by USDA agreement # 58‐1950‐7‐707
Basicranial and temporal bone morphology of a sample of early human fossils
Authors: Smith Heather F
Published: June 2021

Extensive debate has centered around the location and timing of the initial speciation events of humans and the subsequent microevolution and migratory routes that lead to their dispersal around the world. Several early human fossils preserve portions of the temporal bone and other aspects of the basicranium, the morphology of which has been found to reflect genetic distances and population history reliably in humans. Thus, the analysis of these regions in early fossil Homo sapiens can inform our interpretations of their relationships to each other and to specific modern human populations.

Eighteen ectocranial landmarks from the temporal bone and basicranium were digitized on 14 early Anatomically Modern Human fossils and casts and 9 Neandertals. A modern human sample of 13 populations was also measured for comparative purposes. Compared to the modern sample, the early humans were more variable in morphology, suggesting that the fossil sample reflects a greater genetic diversity than humans today, who have undergone a genetic bottleneck. In a multidimensional scaling plot, the fossil specimens also do not cluster together morphologically. The Neandertal specimens fell outside the cluster of early and modern Homo sapiens, and did not share closer relationships with the human specimens from the same continent than from elsewhere.

This study was supported by NSF BCS‐0622570 and the Wenner‐Gren Foundation.

Grant Funding Source: n/a
E. E. Just, Johannes Holtfreter, and the origin of certain concepts in embryo morphogenesis
Authors: Byrnes W. Malcolm
Published: June 2021

Ernest E. Just (1883–1941) is best known for his his elucidation of what are known as the fast and slow blocks to polyspermy during fertilization. Just's contemporary Johannes Holtfreter (1901–1992) is known for his work on amphibian morphogenesis, which helped to lay the foundation for modern vertebrate developmental biology. After giving a brief biographical sketch and outlining his scientific contributions, this poster will argue that Just's work and ideas strongly influenced two of the concepts for which Holtfreter is best known: tissue affinity and autoinduction. First, Just's experiments demonstrating developmental stage‐specific changes in the adhesiveness of the blastomeres of cleavage embryos helped lay the foundation for Holtfreter's concept of tissue affinity. Second, Just's notion of the intrinsic irritability of the egg cell strongly informed Holtfreter's concept of the non‐specific induction of neural tissue formation in amphibian gastrula ectoderm explants. Acknowledgment of these contributions by Just in no way diminishes the importance of Holtfreter's groundbreaking work. It does, however, extend the impact of Just's work into the area of embryo morphogenesis. It connects Just to Holtfreter and positions his work as an antecedent to embryo research that continues to this day. Supported by grants from the NIH RCMI and MBRS‐SCORE programs, and the NSF.
A stopped‐flow kinetic analysis of substrate binding and catalysis by Escherichia coli RNA polymerase
Authors: Johnson Ronald Sanders , Strausbauch Mark
Published: June 2021

To test the hypothesis that the binding of the correct NTP to a pre‐insertion site triggers rapid pyrophosphate (PPi) release followed by entry of the NTP into the active site for incorporation, we used the intrinsic protein fluorescence of the core polymerase in well‐defined elongation complexes to monitor nucleotide binding and incorporation. The increase in fluorescence in each case could be fitted by a single exponential. Plots of the observed first order rate constants versus NTP concentration were hyperbolic. This is consistent with a mechanism involving rapid binding followed by a slow conformational change. In the case of UMP incorporation, the value of the rate constant for the conformational change was 590±60 s−1 and the value of the apparent dissociation constant for UTP was 8.6±3.8 μM; in the case of AMP incorporation, the corresponding values were 520±50 s−1 and 15.7±5.6 μM. The rate constant for PPi release in the presence of the next correct nucleotide for incorporation was reported previously to be 587±178 s−1. The coincidence between the rate constants for the conformational change and PPi release is consistent with the postulated mechanism. NSF Grant #211369
Computational approach to investigate the catalytic chemical reaction mechanism of glyoxysomal Malate Dehydrogenase
Authors: Guterres Hugo , Bell Ellis
Published: June 2021

Geometry optimization calculation was used to find the minimum total energy and structure of the transition state during conversion of malate to oxaloacetate. During the reaction proton abstraction to H220 in the active site, and hydride transfer to NAD occur. Our primary goal was to develop a computational approach to examine the effects of active site and second sphere mutations in the activity of gMDH. Gaussian 03W was used to build the structure of the compounds, arrange their symmetry, and run the geometry optimization calculations. A truncated model including Histidine 220, malate and NAD was used to analyze the effect of the catalytic base. The result shows an imaginary frequency which indicates the presence of the transition state. The geometry structure of the transition state confirms the hydride transfer but the proton transfer is disoriented. This suggests that amino acids around the active site play important roles in facilitating the hydride and proton transfer. We have finished the transition state energy for the native gMDH and are working to find the total energy for transition state of mutants gMDH to facilitate the study of second sphere mutations.

This work is supported by NSF Grant MCB 0448905 to EB.
Structure‐function studies of the regulatory bicarbonate binding in H. influenzae carbonic anhydrase: a functional reversion of allostery using multiple site mutations
Authors: Hoffmann Katherine M. , Rowlett Roger S.
Published: June 2021

Type II β‐carbonic anhydrases are metalloenzymes that catalyze the conversion of carbon dioxide to bicarbonate using a characteristic closed four‐side chain coordination sphere for the active site Zn ion. Non‐catalytic bicarbonate binding sites have been discovered in some type II β‐CAs 8 Å away from the active site. A network of seven hydrogen bonds holds the inhibitory bicarbonate in place, and kinetic studies confirm cooperative inhibition by bicarbonate at sufficient levels. A comparison of bicarbonate‐bound HICA with type I β‐CAs lacking the non‐catalytic bicarbonate binding site reveals a number of principal interactions that might be responsible for the allosteric regulation of HICA. W39 provides a direct, stabilizing hydrogen bond to the inhibitory bicarbonate, a one‐residue shift of proline 48 changes an influential backbone orientation of the 44–48 loop, and a G41 residue makes additional room in the allosteric binding site compared to a conserved Ala in non‐allosteric β‐CAs. Individual site mutants of W39 have revealed predictable decreases in cooperativity consistent with the loss of a stabilizing hydrogen bond to the bicarbonate. Structure function studies of compound site mutants, however, abrogate the binding of inhibitory bicarbonate in the crystalline state even at high concentrations.

This work is supported by NSF grant CHE‐0819686 and MCB‐0741396.
In Vitro Reconstitution of a Bacterial Polysaccharide Biosynthetic Pathway
Authors: Zhao Guohui , Woodward Robert , Yi Wen , Li Lei , Eguchi Hironobu , Motari Edwin , Cai Li , Liu Xianwei , Song Jing Katherine , Han Weiqing , Zhang Wenpeng , Wang Peng George
Published: June 2021

Bacterial polysaccharides represent an important pathogenic factor in infectious diseases. However, the complexity of these molecules has hampered efforts to delineate their biosynthesis. In our research, we chemo‐enzymatically synthesize O86 O‐unit (RU‐PP‐Und). We carried out an in vitro reaction to probe polymerization using 3H‐labelled RU‐PP‐Und and partially purified polymerase Wzy. The paper chromatography result demonstrated the formation of higher molecular weight compounds in the presence of Wzy. The radioactive signal on the SDS‐PAGE gel gave a ladder‐like pattern consistent with the formation of multiple repeating units. It is thus evident that repeating unit polymerization has been successfully reconstituted in vitro. We also introduced purified Wzz proteins obtained from two different E. coli O86 strains (B7 and H2) into the polymerization system. Our results demonstrate the in vitro generation of polysaccharides with appropriate chain length modalities through use of purified Wzz proteins. We prepared analogs derived from various lipids. The results indicate that double bond geometry holds more importance in determining the efficiency of polymerization. In conclusion, we have demonstrated the use of a reconstitution system to not only enable the in vitro production of polysaccharides, but also allow for generation of different chain length modalities. This system has provided the groundwork necessary for further underpinning the molecular details of the wzy‐dependent pathway. This research is supported by NCI (R01 CA118208), NIH (R01 GM085267), NSF (CHE‐0616892) and Bill & Melinda Gates Foundation (51946) for financial support.
Tumor necrosis factor is necessary, but not sufficient, for castration induced prostate regression
Authors: Davis Jennifer Sarah , Nastiuk Kent L , Krolewski John J
Published: June 2021

Tumor necrosis factor (TNF) is a pro‐inflammatory cytokine that plays a vital role in immune responses. TNF is also a potent apoptotic stimulus in vitro. However, few reports have demonstrated a physiologic role for TNF induced apoptosis in vivo. Employing high field MRI (magnetic resonance imaging) to measure mouse prostate volume, we demonstrate that the extent of castration induced prostate regression is significantly reduced in mice null for either the TNF or TNFR1 genes. Wild type (WT) mice receiving soluble TNFR2 prior to castration exhibit a nearly identical reduction of prostate regression. Additionally, membrane‐bound TNF protein and TNF mRNA levels increase in the ventral lobe of rat prostate following castration. When injected into the peritoneum of TNF−/− mice at the time of castration, soluble TNF is able to restore normal levels of prostate regression. However, WT mice receiving soluble TNF in the absence of castration do not exhibit prostate regression, indicating that TNF alone is not sufficient, but must act in the context of additional castration induced signals. These findings support a physiologic role for TNF in prostate regression following androgen withdrawal. Understanding this role may lead to novel therapies for prostate cancer.

J.S. Davis is supported by DOD Pre‐doctoral Training Grant #PC073208.
Truncated beta‐helices maintain template‐assisted hemolysis
Authors: Laurent Daniel , Weaver Todd
Published: June 2021

The two‐partner secretion pathway is the most commonly found protein secretion pathway within gram‐negative bacteria. TPS pathways harbor both A (TpsA) and B (TpsB) components. TpsB components are beta‐barrel outer membrane proteins that are responsible for the coupled secretion and activation of their paired TpsA component. TpsA components include adhesions and hemolysins. Recently, we have characterized HpmA265, a truncated TpsA homologue from Proteus mirabilis. Functionally, HpmA has been shown to facilitate biphasic, cooperative, heat stable, and uni‐directional activation of hemolysis. Structurally, HpmA265 was shown to adopt a right‐handed beta‐helix structure and form a dry, dimeric interface between exposed, carboxy‐terminal, on‐edge beta‐strands. To further investigate the importance of exposed on‐edge beta‐strands we constructed a series of truncated mutants that proceeded up the beta‐helix. HpmA243 and HpmA141, which harbor the first 243 and 141 residues, were the focus of this study. The two carboxy‐terminal mutants were expressed, purified and characterized for hemolytic activity. Both carboxy‐terminal truncated forms of HpmA were still able to facilitate the activation of hemolysis. In addition, HpmA141 was able to maintain higher order oligomeric structure even in the presence of anionic detergent and high temperature. The results further support the theory that exposed carboxy‐terminal beta‐edges are utilized to facilitate TpsA folding and activation. This model of activation has similarities to the propagation of beta‐amyloid diseases. These studies were supported by ASBMB Undergraduate Research Award (to DL) and NSF award MCB0744754 (to TW).
Role of fluorine in distortion from planarity of Polycyclic Aromatic Hydrocarbons
Authors: Liebowitz Daniel , Caruso Francesco , Zajc Barbara , Rossi Miriam
Published: June 2021

Polycyclic aromatic hydrocarbons (PAHs) are a large class of compounds that pose a cancer risk to humans. Previously, it has been shown that increased molecular distortion decreases the amount of PAH precursors that are transformed into carcinogenic metabolites through cytochrome P450‐mediated oxidation of Benzo[c]phenanthrene (BcPh) derivates to DNA‐alkylating diol epoxides. The presence of substituent groups in and around the bay or fjord regions has been shown to influence out‐of‐plane distortion. In our studies, we analyzed the effect of the introduction of Fluorine atoms into the angular ring of chrysene and BcPh. Chrysene has a less hindered bay region and is very planar, while BcPh is the smallest PAH with a highly hindered fjord region and has been shown to be distorted 26.7° between the outermost rings. The 3‐D structures of 5‐FluoroBcPh, 6‐FluoroBcPh, 5‐Fluorochrysene, Benzo[g]chrysene and 10‐Fluorobenzo[g]chrysene were determined by single‐crystal X‐ray diffraction with Mo radiation using the Bruker Smart APEX II diffractometer. Results reveal a decrease in distortion from planarity upon the introduction of Fluorine atoms. The angle between the outermost rings was found to be decreased from 26.7° in BcPh to 24.3° and 23.6° in the 6‐Fluoro‐ and 5‐FluoroBcPh respectively. The angle between the outermost rings in 10‐Fluorobenzo[g]chrysene was found to be 33.33°, which is a decrease from 36.04° in its non‐fluorinated precursor. Future studies on these and other similar compounds will clarify the role of substituent groups on the out‐of‐plane distortion and measure the relationship between distortion and DNA alkylating ability. Research is supported by a grant from CNR for Short Term Mobility Funding (FC) and an NSF grant for the Bruker X‐ray diffractometer (CHE 0521237).
MicroRNAs Associated with Environmental Stress in Arabidopsis
Authors: Brown Rebecca Sul Hee , Jeong Dong‐Hoon , Meyers Blake C , Green Pamela J
Published: June 2021

One way plants respond to environmental stress is by modifying their gene expression through the use of microRNAs (miRNAs). miRNAs are noncoding small RNAs that regulate gene expression at the posttranscriptional level by base pairing with complementary messenger RNA (mRNA) molecules, causing either mRNA cleavage or translational inhibition. To elucidate the roles of miRNAs in environmental stress responses, wild type plants and miRNA enriched mutant plants were subjected to various environmental stresses. Small RNA libraries from stress‐treated seedlings and flowers were constructed and sequenced using deep sequencing technologies. Computational analysis revealed differential expression of miRNAs between stress‐treated plants and control plants with some miRNAs upregulated and some downreguled by environmental stress. Additional analysis revealed potentially novel miRNAs in Arabidopsis as well. These results suggest that miRNAs play important roles in stress responses, and that miRNA identification in Arabidopsis has not been saturated. Future work will involve validating highly stress‐regulated miRNAs and prospective new miRNAs, and examining their potential target genes for regulation. Stress‐regulated miRNAs will be incorporated into multi‐network models and their biological roles will be hypothesized and tested through functional studies. R.S.H.B. was supported by the Howard Hughes Medical Institute, and NSF grant MCB#0548569 to P.J.G. provided research support.
Thermodynamic Examination of Conserved Sequences of TAR RNA in HIV‐1
Authors: Hannan Philipp Lorenz , Kresheck Gordon , Grover Neena
Published: June 2021

Folding of ribonucleic acids generates multiple helical and non‐helical regions. Various non‐canonical interactions occur in the bulges and loops formed in the non‐helical regions of RNA to create a diversity structures that are recognized by target molecules. In HIV‐1 TAR RNA the trinucleotide bulge region interacts with an arginine rich domain of the Tat protein, leading to a 100 fold increase in transcription of the viral genome. A base triple between a bulged‐U and a conserved base pair in the upper stem is expected to be important for Tat‐TAR interactions. Previous studies in our laboratory have examined the stability of TAR RNA constructs containing trinucleotide bulges, and have found that uracil‐rich bulged constructs are ~2 kcal/mol more stable than cytosine‐rich bulged constructs. Thermodynamic properties of a bulge containing all purines, a bulge containing methylated sugars, and TAR constructs with altered conserved base pairs above and below the bulge are being examined in this study, along with their arginine binding properties. Thermal denaturation and isothermal calorimetry experiments show that arginine binding to a TAR UCU construct improves in the presence of divalent ions, whereas TAR CCC shows no arginine binding. This work was funded by NSF MCB‐0621509 to NG.
Thermodynamic stablity and divalent ion interactions of RNA containing purine‐rich bulges
Authors: Strom Shane Fancher , Grover Neena
Published: June 2021

Bulges are a common non‐helical motif found in RNA that play an important role in RNA recognition. Small asymmetrical bulges occur in many different RNA including HIV‐RNA and various microRNA. In HIV‐1 TAR RNA, a trinucleotide bulge interacts with the Tat protein of HIV‐1 to increase its rate of transcription one hundred fold. Previous studies in our lab have shown that TAR RNA containing cytosine‐rich trinucleotide bulges are ~2 kcal/mol more stable than those containing uracil‐rich bulges, in 1 M KCl conditions. Magnesium ions stabilize TAR RNA containing uracil‐rich bulges by ~1–2 kcal/mol and cytosine‐rich bulges by ~ 1 kcal/mol, relative to 1 M KCl conditions. In this study, we are examining the differences in RNA stability as the bulge size is varied between one and five nucleotides. Thermal denaturation experiments are being performed on varying concentrations of RNA in buffers containing 1 M KCl or varying concentrations of divalent ions. Altering the nucleotide sequence of the bulge alters RNA stability as well as its ion binding properties. A comparison of thermodynamic properties of purine‐ and pyrimidine‐rich bulges will be presented. This work was funded by NSF MCB‐0621509 to NG.
Protein interaction and phosphorylation of diaphanous 1 regulate mitochondria movement in H295R human adrenocortical cells
Authors: Li Donghui , Sewer Marion B
Published: June 2021

Our lab has previously shown that mitochondrial movement regulates production of cortisol and adrenal androgen by adrenocorticotropin (ACTH) and dibutyryl cAMP (Bt2cAMP) stimulation. This movement is dependent on phosphorylation of RhoA and the RhoA effector protein diaphanous 1 (DIAPH1). DIAPH1 promotes the polymerization, and nucleation and branching of actin filaments. Based on these data, we hypothesized that ACTH/Bt2cAMP‐stimulated mitochondria movement requires the formation of a macromolecular complex containing DIAPH1. To test this hypothesis, we used mass spectrometry to identify proteins bound to DIAPH1. We found that DIAPH1 binds to several proteins, including microtubule‐associated protein 4, dynamin‐1, kinesin, β‐actin, and g‐actin. Further, we also identified that DIAPH1 is phosphorylated at multiple sites. Using time‐lapsed video microscopy, we found that overexpression of DIAPH1wt‐GFP but not mutants (DIAPH1S1067D–GFPand DIAPH1Y1097F‐GFP) increases mitochondria movement. These results suggest that protein interaction and phosphorylation of DIAPH1 play a key role in the regulating the rate of mitochondria movement. This study is supported by MCB0818119 from the NSF.
When Reduction Leads to Expansion: A Simple Native PAGE Gel Enzyme Assay Experiment as a Starting Point for Other Structure‐Function Investigations
Authors: Hazzard James Taylor , Pasalic Dario , Edwards Sarah , Friedrich Kenneth
Published: June 2021

A simple assay on native PAGE gel for two E. coli enzymes, purified by undergraduates in a biochemistry laboratory, provides an excellent springboard for more sophisticated investigations on protein structure. They perform colorimetric assays on b‐galactosidase (b‐gal) using ortho‐nitrophenyl‐b‐galactosidase in a buffer containing b‐mercaptoethanol (BME) and on alkaline phosphatase (AP) using FAST RED TR/Naphthol AS‐MX reagent. Half of the class assays for b‐gal first and AP second; the other half of the class reverses the order. Results differ because the former group reduces two disulfide bonds which are essential for AP activity, only observing activity for b‐gal, while the latter sees activity for both enzymes.

This simple PAGE experiment leads the students to the question: Is loss of activity due to protein denaturation? Answering this question raises more sophisticated investigations, including BME dialysis, intrinsic fluorescence spectroscopy of AP tryptophan residues, ANS fluorescence spectroscopy, and circular dichroism. These can be offered as special projects to laboratory groups who report their findings to the entire class, providing a holistic approach to investigating the relationship between loss of activity and protein denaturation. Initial data and results from our laboratory course will be presented. This work supported in part by a grant from NSF (DUE 0837398 to JTH).