bluuebunny/crossref_metadata_2025_split huggingface dataset:
Functional Analyses of Myc and β‐catenin during Echinoderm Intestinal Regeneration using RNAi
Authors: Alicea‐Delgado Miosotis , Salem‐Hernández Joseph , Garcia‐Arraras Jose E 
Published: June 2021

Molecular approaches have been widely used to understand and characterize regenerative processes in different animal systems. Nevertheless, characterizing the genetic component of echinoderms using RNAi techniques represent a tremendous challenge. We have now designed and implemented the use of Dicer‐substrate RNAi technology (DsiRNA) to knockdown the expression of Myc and β‐catenin transcripts on tissue explants of the sea cucumber Holothuria glaberrima. Quantification of Myc and β‐catenin transcripts showed a decrease following electroporation of specific DsiRNAs when compared to explants electroporated with control DsiRNA. Explants were then assayed for the effects on cell proliferation, cell dedifferentiation and apoptosis. No effect on cellular dedifferentiation was observed. In contrast, the number of apoptotic cells was significantly different in explants electroporated with DsiRNAs‐targeting Myc and β‐catenin when compared to control DsiRNA. Finally, a decrease in cell proliferation was observed following the reduction of Myc transcript, but not following the reduction of β‐catenin. These results suggest that Myc has a significant role on cell proliferation and apoptosis, but not on cell dedifferentiation during the regeneration of the intestine. Similar to Myc, β‐catenin has a significant role on apoptosis, but not on cell proliferation and cell dedifferentiation. Moreover, our results show that DsiRNAs and intestinal tissue explants are a valuable tool to manipulate and explore the role of genes of interest in echinoderms.Support or Funding InformationFunded by NIH R15GM124595, NSF IOS‐1252679 and the University of Puerto Rico.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Role of TRPV4‐TAZ signaling axis in epithelial‐mesenchymal transition
Authors: Rahaman Shaik , Sharma Shweta , Goswami Rishov 
Published: June 2021

Epithelial to mesenchymal transition (EMT) is a critical event in development, fibrogenesis, and oncogenesis. Emerging data support a role for both a mechanical signal, and a biochemical signal, e.g., transforming growth factor β1 (TGFβ1) in EMT. Here, we report evidence that transient receptor potential vanilloid 4 (TRPV4), a mechanosensitive channel sensitive to both matrix stiffness and TGFβ1 stimuli, is a likely mediator of EMT. We found that antagonism of TRPV4 blocked both matrix stiffness‐ and TGFβ1‐induced EMT in normal human primary epidermal keratinocytes (NHEKs), as determined by changes in morphology and alterations in expression of EMT markers including E‐cadherin, N‐cadherin, and α‐smooth muscle actin (α‐SMA). In a murine skin fibrosis model, TRPV4 deletion resulted in decreased expression of the mesenchymal marker, α‐SMA, and increased expression of epithelial marker, E‐cadherin. Mechanistically, we found that: i) TRPV4 activity was essential for the nuclear translocation of TAZ (transcriptional coactivator with PDZ‐binding motif) in response to matrix stiffness and TGFβ1; ii) Antagonism of TRPV4 activity inhibited both matrix stiffness‐induced and TGFβ1‐induced expression of TAZ proteins; and iii) TRPV4 antagonism suppressed both matrix stiffness‐induced and TGFβ1‐induced activation of Smad2/3, but not of AKT. Altogether, these data identify a novel role for TRPV4 in regulating EMT in response to both matrix stiffness and TGFβ1.Support or Funding InformationAHA (13SDG17310007), Startup grant from University of Maryland, NIH (1R01EB024556‐01), and NSF (CMMI‐1662776) grants to Shaik RahamanThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Hof1 of Aspergillus nidulans is Essential for Cytokinesis and Septum Formation
Authors: Rowland Lauren , Brookover Zoe , Ward Rachel , Hill Terry , Jackson‐Hayes Loretta 
Published: June 2021

Cytokinesis is an essential component of cellular development in filamentous fungi. The process parallels that of cell division in animal cells in requiring assembly and constriction of a contractile actomyosin ring (CAR) in the cell periphery. However, the process differs from that in animals by involving the simultaneous construction of a chitin‐rich cross wall termed a septum, which separates semi‐independent compartments of the fungal hypha. Through randomized mutations, we have generated a temperature‐sensitive mutant defective in septum formation, designated strain RCH59. Mendelian crossing confirmed that the temperature sensitive phenotype results from a single locus mutation. Next Generation Whole Genome Sequencing identified a mutation that is located in gene AN4566 encoding the homolog of Saccharomyces cerevisiae's Hof1. The mutation in Aspergillus nidulans’ version of Hof1 (AnHof1) is at base 2436 out of 5536 and is a transition from cytosine to thymine. The mutation is predicted to remove an Arginine and introduce a premature stop codon after residue 738. The missing 342 C‐terminal residues includes the entire SH3 domain of AnHof1. We cloned the wildtype version of AnHof1 into plasmid pRG3 and complemented the phenotype in RCH59, confirming that the mutation occurs in AnHof1. We also deleted AnHof1 to demonstrate that a AnHof1 minus strain has an aseptate phenotype. In the yeasts (S. cerevisiae and S. pombe), Hof1 forms a ring structure that co‐localizes with the CAR and is involved with mediating the cytoskeletal rearrangements necessary for cytokinesis. We GFP tagged AnHof1 to demonstrate that the protein product localizes to sites of septum formation and co‐localizes with actin during at least part of CAR assembly and constriction. In further work, we have generated double‐mutant strains in which a GFP‐tagged septation‐related protein is expressed in the RCH59 strain to demonstrate a network of genetics interactions between AnHof1 and other proteins involved in septation.Support or Funding InformationThis research was supported by NSF grant RUI‐0742907 to TWH and LJH and NSF grant RUI‐1615192 to LJH and TWH.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Inositol Pyrophosphates and Phosphate Sensing in Plants
Authors: Gillaspy Glenda , Adepoju Olusegun , Land Eric , Williams Sarah Phoebe , Donahue Janet , Cridland Caitlin , Freed Catherine , Perera Imara 
Published: June 2021

Phosphate (Pi) is an essential nutrient for plants, required for plant growth and seed viability. Under Pi stress, plants undergo dynamic changes to leverage available Pi. One class of signaling molecules implicated in the Pi sensing pathway is the inositol pyrophosphates (PPx‐InsPs). PPx‐InsPs have high energy bonds, and have been linked to maintaining Pi and energy homeostasis in plants, yeast, and humans. In plants, PPx‐InsPs are thought to function in sensing Pi changes by binding to proteins containing an SPX domain. One particular SPX protein binds to PPx‐InsPs which prevents a transcription factor from activating transcription of the so‐called phosphate starvation response (PSR) genes. Plants are known to accumulate two common PPx‐InsPs, PP‐InsP5 (called InsP7) and PP2‐InsP4 (called InsP8), but the synthesis pathway has not yet been elucidated. We have identified and characterized the plant VIP kinases, enzymes that likely function as PP‐InsP5 kinases. The identity of a plant inositol kinase that can phosphorylate InsP6, resulting in PP‐InsP5, however, has remained elusive. We will describe our recent results in characterizing this pathway, and genetic mutants key for understanding inositol kinase gene function in plants. Our results support a model in which InsP7 and InsP8 function to turn off the PSR in plants.Support or Funding InformationThis work was supported by an award from the NSF to GG and IP:MCB 1615953This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Transgenic analysis of maize endosperm metabolism
Authors: Juhl Emily , Myers Alan M. , Hennen‐Bierwagen Tracie A. 
Published: June 2021

Maize seeds provide a major component of the calories in the human diet, are critical as a feed source in animal agriculture, and are an important source of biorenewable raw materials. Endosperm tissue accounts for approximately 80% of seed weight, and is the site where carbohydrate and protein storage end products are produced in this net anabolic system. Sucrose supplied from photosynthetic tissues of the maternal plant is transported to growing seeds where it is divided between catabolic pathways that generate ATP and anabolic biosynthetic processes that utilize ATP to generate starch and storage proteins. Considering the high degree of ATP requirement, the fact that maize endosperm is an extremely hypoxic environment raises questions about how energy is generated, and the net efficiency of the system for product formation. This project tested the hypothesis that fully oxidative metabolism is not required for normal kernel development, and accordingly that ATP is generated exclusively or substantially through fermentative processes. To do so, RNAi technology was utilized to inactivate the 2‐oxoglutarate dehydrogenase (ODH) step of the citric acid cycle. T‐DNA vectors were constructed that expressed a double stranded RNA structure containing sequences homologous to both of the maize mRNAs encoding the E2 subunit of this enzyme. Highly active, tissue specific promoters were used to restrict the effects of the transgene exclusively to endosperm tissue. Plants were generated containing the anti‐ODH transgene, and subsequent crosses have established segregating populations of sibling kernels that either contain or lack that element. ODH activity and protein abundance in endosperm tissue will be measured to verify that the transgene is effective in inactivating the targeted enzyme activity. Subsequently, segregating kernel populations will be compared regarding growth rate over the course of development, mature kernel weight and composition, and global metabolite profiles. These studies will reveal the extent to which ODH is required for starch and storage protein synthesis to proceed in developing maize endosperm. This will reveal whether normal ATP generation in this tissue requires oxidative processes or, alternatively, whether fermentative processes can be sufficient to provide the energy required for the massive amount of starch and protein synthesis that makes maize such an important agronomic resource. Answering this question will enable future efforts to improve maize yields and grain quality.Support or Funding InformationThis research was supported by NSF award 1517256 to A.M.M. and T.A.H.‐B.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Data Mining Scientific Literature Demonstrates Use of Biological and Medical Data Across Scientific Disciplines
Authors: Verdiguel Natalie , Feng Zukang , Westbrook John , Zardecki Christine 
Published: June 2021

The Protein Data Bank (PDB) was established as the 1st open access digital data resource in biology and medicine. Today, the PDB archive contains over 140,000 atomic‐level biomolecular structures determined by crystallography, NMR spectroscopy, and 3D electron microscopy. About 20 new structures are deposited daily. PDB data is used in basic and applied research, patent applications, discovery of lifesaving drugs, product innovation, and education. About 80% of all PDB structures deposited have a “primary citation,” which is the first paper to describe the molecule and its structure and function. In turn, these primary citations have contributed to more than 1 million publications. An initial study by Clarivate Analytics on the Nucleic Acids Research publication demonstrated that PDB data drive high‐impact research in diverse scientific fields. To build on this study, we used Python programming to perform several data mining studies to understand the impact of these structures in fundamental biology, biomedicine, and energy in the scientific literature. The scientific literature database, Web of Science, tracks scientific article metrics such as the number of times cited, subject category, year published, funding source, and keywords. Web of Science data for PDB primary citations was compiled as used as the data set. Two main investigations were performed. In the first, we examined the number of citations per article (data overall and by individual subject categories) to develop “top 10 cited structures” lists and analyzed for trends. In the second investigation, we extended our data mining analysis to consider for each main subject category, the papers with the most citations and the structures they cite, and the most frequently occuring keywords. We have observed different aspects of biological data utility in various subject areas, at times in ways unanticipated from the structures' primary citation. This research demonstrates the value of biological data from the PDB by showing the diverse ways in which the data are being used.Support or Funding InformationRCSB PDB is funded by a grant (DBI‐1338415) from the National Science Foundation, the National Institutes of Health, and the US Department of Energy.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Our Unique Cell Lines Derived Serially After Injection With Human T24 HRAS Transformed NIH/3T3 Mouse Cells (GhrasT‐NIH/3T3) Showed The Highly Tumorigenic T1‐A Cells Cultured From The First Primary Tumor Had A Six‐Fold Higher LD50 For Resveratrol Killing Than The LD50 For Highly Metastatic Lung T4‐PA Cells Derived After Three Subsequent In Vivo / In Vitro Serial Manipulations
Authors: Ray Durwood Barber , Jones David H. 
Published: June 2021

Our new cell lines demonstrating progressively increasingly metastatic behavior as they were derived in a series of in vivo/in vitro manipulations (Figure 1) provide the opportunity to evaluate in vivo and in vitro compounds and methods to kill primary tumor cells and various forms of increasingly metastatic cells derived from the same primary tumor (Figure 2) (Ray, D.B., et. al., Experimental Cell Research, 2016 Jan; 340 (1): pp. 1–11). We determined the dose of the anti‐cancer natural phytoalexin resveratrol to kill 50% (LD50s) of our various cells that represent progressive cancer stages. Cell counts were recorded daily, over the course of 3 to 6 days, using gridded plates and photomicrographs of six pre‐determined 2 mm square sections of each plate. An average percent change in cell number from each individual grid was then calculated. P‐values were determined between each treatment time and dose and significance was determined. The NIH Swiss and NIH/3T3 cells were not significantly affected by 3–300 μM after 3 days. After 3 or 4 days the LD50s for the tumorigenic cells were GhrasT‐NIH/3T3 cells: LD50 = 3.5+/− 1.0 μM; T1‐A cells: LD50 = 45 +/− 5.0 μM; T4‐PA cells: LD50 = 7.0 +/− 1.5 μM. Thus, unexpectantly, the highly tumorigenic T1‐A cells were significantly more difficult to kill than the highly metastatic T4‐PA cells. This pattern may mimic the epithelial to mesenchymal transition (EMT) seen in breast and other cancer progression. EMT enhances mobility, invasion, and resistance to apoptotic stimuli as tumor cells acquire stem cell properties and exhibit marked therapeutic resistance. RNA‐Seq, genome sequencing (WGS) and spectral karyotyping (SKY) analysis of these cells demonstrating increasingly aggressive metastatic cancer tumorigenesis that progressed from a single nuclear genotype and mitochondrial haplotype should help contribute to the understanding of cancer progression. These cells are available by contacting our laboratory.Support or Funding InformationSupported in part by the National Science Foundation grant # 8602787, the Grove City College Research Fund, the Grove City College Swezey Research Fund and the Grove City College Janicki Foundation Research Fund.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Weight loss and fitness intervention increase markers of hepatic bile acid (BA) synthesis, while reducing serum total BA concentrations in sedentary, obese insulin resistant women
Authors: Mercer Kelly E , Pack Lindsay M , Lou Xiangyang , Thyfault John , Campbell Caitlin , Fiehn Oliver , Chandler Carol J , Burnett Dustin J , Souza Elaine C , Casazza Gretchen A , Keim Nancy , Newman John W , Hunter Gary R , Fernandez Jose R , Garvey W Timothy , Harper Mary‐Ellen , Hoppel Charles L , Adams Sean H 
Published: June 2021

Beyond adipose and muscle, excerise has profound effects in other metabolically‐important tissues including the liver. Bile acids (BAs) are synthesized from cholesterol via the rate‐limiting enzyme cholesterol 7 alpha‐hydroxylase (CYP7A1) and contribute to the solubilization and absorption of lipid soluble nutrients, and serve as important signaling molecules capable of systemic endocrine function. Circulating BA are known to be higher in obesity and insulin resistance, but effects following exercise and diet‐induced weight loss are unknown. To test if improved fitness and weight loss influence bile acid metabolism and homeostasis, we measured fasting serum concentrations of total BAs (conjugated and free forms of primary and secondary BAs) in sedentary, obese insulin‐resistant women (N=12) before and after a ~14 wk weight loss and exercise intervention [Campell et al. 2014, PLoS One, 9: e84260]. In addition, serum FGF19 (a regulator of BA synthesis) and 7‐alpha‐hydroxy‐cholesten‐3‐one (C4, a verified marker of CYP7A1 enzymatic acitivity) were measured by ELISA and LC‐MS, respectively. Using mixed‐model analyses and the change in relative VO2peak (mL/min/kg) as a covariate, we observed a significant decrease (~30%) in total serum BAs post‐intervention relative to pre‐intervention samples. Corresponding to the decrease in total BAs, fasting serum concentrations of C4 were 55% higher post‐intervention relative to the matched pre‐intervention samples (P=0.004). Interestingly, we observed no changes in serum FGF19. These data suggest that weight loss and/or improved fitness altered BA metabolism marked by increased hepatic BA synthesis and reduced serum BA. The underlying mechanisms remain to be determined, but work from our preclinical rodent models supports the idea that increased BA metabolic flux is linked to enhanced hepatic mitochondrial function.Support or Funding InformationThis work was supported by USDA Projects 6026‐51000‐010‐05S and 2032‐51530‐022‐00D, and NIH‐NIDDK R01DK078328 and NIDDK U24 DK097154.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Evaluating Physiological Interactions between the Electrogenic Na/HCO3 Transporter NBCe1‐B and its Cytosolic Binding Partner IRBIT
Authors: Lee Seong‐Ki , Boron Walter F. 
Published: June 2021

The electrogenic Na/HCO3 transporter NBCe1 is expressed in numerous organs, where it plays many important roles, including trans‐epithelial HCO3− transport (e.g., pancreatic ducts) and maintenance of intracellular pH (e.g., cells in CNS). Presently, five NBCe1 variants are known: the A variant, mainly in kidney; B, ubiquitous; C, CNS; and D and E, murine reproductive organs. The extreme NH2‐terminus (Nt) of each variant is a strong determinant of transport activity. A and D have a unique Nt of 41 amino acids (aa) that, at least in A, confers high transport activity. B, C, and E have a unique Nt of 85 aa that, at least in B and C, confers low transport activity. However, B interacts with IRBIT [inositol trisphosphate (IP3)‐receptor binding protein released with IP3], which binds to the unique Nt of NBCe1‐B, and markedly enhances the transport activity of NBCe1‐B. Previously, others examined the interaction by fusing a fragment of the unique Nt of NBCe1‐B/C/E to maltose‐binding protein (MBP) for use in a MBP pull‐down assay, and found that NBCe1‐B (e1B) residues 1–62 are necessary for the interaction. In a recent study, we showed that the cationic cluster (residues 40–48 inclusive) is an essential element of the autoinhibitory domain (AID) that confers low activity on e1B. The purpose of the present study is to refine the structural requirements of the IRBIT‐e1B interaction under physiological conditions. Here, we co‐express 7 constructs of e1B (one at a time) together with super‐IRBIT (sIRBIT, which lacks the PP1 binding site) in Xenopus oocytes, followed by two‐electrode voltage‐clamp, extracellular‐surface‐biotinylation, and NeutrAvidin pull‐down of plasma‐membrane proteins. We find that sIRBIT copurifies, as assayed by western blot, with biotinylated e1B protein, but only for low‐activity e1B constructs (i.e., AID intact) for which sIRBIT enhances transport activity. sIRBIT does not copurify with the Δ940–48 e1B construct, which lacks the cationic cluster (i.e., AID disrupted). Thus, the cationic cluster is not only essential for autoinhibition, but also for sIRBIT binding. Our approach will be useful for elucidating other motifs that may be necessary for the interaction of IRBIT with NBCe1‐B, ‐C, or ‐E.Support or Funding InformationSupported by grants from the Office of Naval Research, N00014‐15‐1‐2060, N00014‐16‐1‐2535This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.