bluuebunny/crossref_metadata_2025_split huggingface dataset:
Investigation of New Chemical Probes for Detection of Citrulline Using Small Molecule Model Systems
Authors: Delaney Brian , Laufenberg Brandon , Kadnikov Dmitry V. 
Published: June 2021

Citrulline is an amino acid formed via post‐translational modification of arginine. Citrullination of proteins underpins development of such health problems as rheumatoid arthritis, multiple sclerosis, other autoimmune diseases, and perhaps other diseases such as Alzheimer's disease. Current probes based on phenylglyoxal exist but are lacking in stability, are difficult to synthesize, and are expensive to produce. A more viable probe that could properly detect concentrations of citrulline in the body could help with early diagnosis of patients affected by these diseases. The goal of this project is to create new, more stable probes for the detection of citrulline in the body. Currently, new reactive heads based on protected phenylglyoxal have been synthesized and their reactions with small molecules mimicking citrulline are being investigated.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effects of Differential Acylation on Aberrant Growth Signaling by Overexpressed Gα13
Authors: White Nicholas F. , McDiarmid Gray R. , Tagliatela Alicia C. , Meigs Thomas E. 
Published: June 2021

Overexpression of Gα13, a member of the G12/13 subfamily of heterotrimeric G proteins, has been implicated in cell transformation and the progression of several cancer types. The mechanism through which wildtype, overexpressed Gα13 drives aberrant growth signaling is not known. G protein α subunits are modified post‐translationally by acylation at the N‐terminus, and this addition of a palmitoyl and/or myristoyl group occurs for all mammalian Gα proteins. Gα13 was previously engineered to mutate two cysteine residues necessary for dual palmitoylation. This modification abrogated signaling to serum response factor by Gα13 in both its wildtype and constitutively activated forms. To determine whether myristoylation restores signaling to non‐acylated wildtype Gα13, the protein was modified to harbor the 6 N‐terminal amino acids from GαT, which is myristoylated but not palmitoylated. Loss of growth signaling by non‐palmitoylated, wildtype G13 was rescued by introduction of this myristoylated sequence. In addition, we used cellular fractionation to assess intracellular distribution of these Gα13 variants. Overexpressed, wildtype Gα13 showed a shift from a membrane associated fraction to a soluble pool, which correlated with a sharp increase in serum response factor signaling. Gα13 lacking any acylation sites localized entirely to the soluble fraction but exhibited no growth signaling. We have also examined the role of an N‐terminal polybasic motif in overexpressed wildtype Gα13 signaling, through amino acid substitutions in this region. To query the nucleotide bound state of the soluble Gα13 pool, we combined cell fractionation with trypsin digestion assays. Surprisingly, overexpressed wildtype Gα13 in the soluble fraction was fully degraded, suggesting lack of GTP binding. These constructs have facilitated co‐precipitation experiments to determine whether change in acylation state affects Gα13 binding to specific target proteins.Support or Funding InformationWe acknowledge support from the North Carolina GlaxoSmithKline Foundation, the C.D. Spangler Foundation, UNC Lineberger Comprehensive Cancer Center, and the UNC‐Asheville Undergraduate Research Program.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Identification of Novel Fibroblast Growth Factor Receptor Signaling Components
Authors: Eichelberger Eric , Webb Jason C. , Stefinko Mariya , Stern Michael J. , Voisine Cindy , Lo Te‐Wen 
Published: June 2021

Fibroblast Growth Factor Receptors (FGFRs) are a type of receptor tyrosine kinase (RTK) that phosphorylate precise tyrosine residues. FGFRs play a role in diverse developmental pathways that mediate cell proliferation, cell migration, and cell survival. We use C. elegans as a model to better understand FGFR signaling specificity. In C. elegans, the EGL‐15 FGFR is imperative for sex myoblast migration and fluid homeostasis. Defects in the processes mediated by the C. elegans EGL‐15 FGFR result in striking phenotypes that can be used to discover components of the EGL‐15 signaling pathway. For example, hyperactivation of EGL‐15 results in the excessive accumulation of clear fluid within the worm's body (the Clr phenotype). The isolation of suppressor of Clr (soc) mutants has led to the identification of many of the core components of EGL‐15 signaling. A previous soc screen identified the SEM‐5 adaptor protein that links RTK activation to the activation of the RAS/MAPK pathway.An egl‐15 mutation, n1457 (a truncation mutation), that eliminates the known SEM‐5 binding sites (Y1009 and Y1087) on EGL‐15 does not confer a Soc phenotype indicating that a key component that links activated EGL‐15 to SEM‐5 has yet to be identified. To identify these missing components, we conducted a modified, “enhancer” soc screen in an egl‐15(n1457) background. Using CRISPR/Cas9, we are introducing mutations in the known SEM‐5 binding sites (Y1009 and Y1087) to determine if the Soc phenotype in newly identified genes is solely dependent on these binding sites. We are also using RNAi to verify the identity of a novel soc gene, cca‐1. Additional genetic analyses and whole‐genome sequencing will be used to identify the molecular identities of additional suppressors identified in our enhancer screen.Support or Funding InformationNIH R15 (GM122001)This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
O‐GlcNAcylation of the Human Kinome
Authors: Liu Xin , Han Guanghui , Renuse Santosh , Pandey Akhilesh , Zhu Heng , Hart Gerald 
Published: June 2021

O‐GlcNAc, first characterized more than 30 years ago, is an O‐linked β‐N‐acetylglucosamine moiety attached to the side chain hydroxyl of a serine or threonine residue. O‐GlcNAc has been found on over 4,000 cytoplasmic and nuclear proteins. The addition of O‐GlcNAc to proteins is catalyzed by O‐GlcNAc transferase (OGT) while its removal is catalyzed by O‐linked N‐acetyl‐β‐D‐glucosaminidase (OGA). This dynamic and reversible modification is emerging as a key regulator of various cellular processes, such as signal transduction, transcription, cell cycle progression and protein‐protein interactions. O‐GlcNAc plays important roles in human diseases, such as cancer, diabetes and neurological disorders. Protein phosphorylation is a post‐translational modification that serves as a rapid and reversible means to modulate protein activity and transduce signals, the regulation of phosphorylation is a central mechanism in cell health and disease. The addition and removal of phosphorylation is through kinases and phosphates respectively, but little is known about O‐GlcNAcylation of kinases.Kinase microarray which has 370 kinases printed in identical pair spots indicated that 297 kinases are substrates for OGT. In addition, 10 kinases were picked from kinase array for overexpression and immunopreciptation to map O‐GlcNAc site. To further confirm the results in vivo, home‐made kinase inhibitor beads were used to pull down the kinases in rat brain. 223 kinases were identified by mass spectrometry and more than 20 kinases have been identified as O‐GlcNAcylated proteins.Support or Funding InformationSupported by NIH P01HL107153, R01DK61671 and R01GM116891. Dr. Hart receives a share of royalty received by the university on sales of the CTD 110.6 antibody, which are managed by JHUThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Developing a physical model of O‐GlcNAc transferase (OGT) in complex with TAB1
Authors: Hernandez Viviana Perez , Nguyen Sophia , Chabbra Alesa , Schmitt Lavin Emily F. 
Published: June 2021

O‐GlcNAcylation is a post‐translational modification similar in importance to the mechanism of phosphorylation in its ability to affect signal transduction. This process is mediated by the enzyme, O‐GlcNAc transferase (OGT). OGT catalyzes the addition of the sugar, N‐acetylglucosamine (GlcNAc) from the carrier molecule Uridine Diphosphate N‐acetylglucosamine (UDP‐GlcNAc), to certain serine or threonine residues in target substrate proteins. The mechanisms by which OGT recognizes its protein substrates, and the details of the molecular interactions between the substrate and enzyme, are still poorly understood. The CREST (Connecting Researchers, Educators, and STudents) team at Nova Southeastern University (NSU) created a physical model of the OGT protein in complex with the fused substrate peptide from TAB1 (a transforming growth factor‐beta‐activated kinase 1 binding protein involved in a variety of signaling cascades including pro‐inflammatory pathways). Details of this protein and its fusion with a segment of the substrate peptide from TAB1 were found in the Protein Data Bank file, 5LVV, and imported into Jmol, a protein visualization software. Specific program commands were utilized in Jmol to manipulate the original file into a format that was later able to be 3‐D printed to create an instructional molecular model. Various features of the OGT fused with the TAB1 substrate peptide were highlighted. Such highlighted features include the regions upstream and downstream of the of the substrate binding sequence. The region upstream is a His6‐tag, while the region downstream is a 3xGly linker segment. Additionally, the fused substrate peptide from TAB1 was highlighted as well as the specific serine that functions as the O‐GlcNAc binding site on the TAB1 substrate. The N and C termini of the OGT were colored blue and red respectively. The overall focus of this project was to explore protein structure and design and build a physical model that illustrates key functional features of the protein. It was funded in part by NSF‐DUE 1725940 for the CREST (Connecting Researchers, Educators, and STudents) Project. Developing 3‐D molecular models in this way is a relatively inexpensive process to visually represent important biological relationships that can be useful for students, professors, and others trying to understand and explain complex molecular pathways.Support or Funding InformationThe work was funded in part by NSF‐DUE 1725940 for the CREST (Connecting Researchers, Educators, and STudents) Project.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Functional Characterization of 2QRU Based on Structural and Enzymatic Analysis
Authors: Pyne Alexandra , Sheikh Sana , D'Ausilio Morgan E. 
Published: June 2021

Beginning in 2000, the National Institutes of Health's Protein Structure Initiative was a fifteen‐year program that worked to solve the structures of many proteins with known DNA sequences. The successful initiative yielded the publication of more than 500 previously uncharacterized proteins. The present study analyzes one such protein, known as 2QRU, a predicted alpha/beta hydrolase found in Enterococcus faecalis. Using modules outlined by BASIL (Biochemistry Authentic Scientific Inquiry Lab) and resources available in a high school molecular biology teaching laboratory, the specific objective of this study is to functionally characterize 2QRU based on structural analysis of the protein. In silico enzyme characterization for 2QRU was carried out using BLAST, Pfam, and Dali servers, followed by active site analysis using the PyMOL plug‐in ProMOL. ProMOL generated a list of proteins with active sites that were structurally similar to 2QRU. For each of the hits with a Levenshtein distance of zero, the predicted catalytic triad for 2QRU was composed of a Ser‐102, Asp‐219, and His‐247. Secondary structure alignments using PROMALS3D and structural alignments in PyMOL were used to compare 2QRU to the active site matches identified using ProMOL. This structural analysis suggests PDB entries 3LIP and 1YSC ‐ a triacylglycerol lipase and serine carboxypeptidase, respectively ‐ are most similar to 2QRU and share a conserved catalytic triad. However, serine carboxypeptidases have yet to be discovered in bacteria, indicating 2QRU may function as a lipase. To investigate the lipase activity of 2QRU in vitro, recombinant his‐tagged protein was isolated from E. coli. Subsequent enzymatic assays using spectrophotometric analysis will be carried out based on methods described for the characterization of Pseudomonas cepacia lipase (PCL) (PDB ID 3LIP) using the substrate p‐Nitrophenyl palmitate, an ester of palmitic acid. As a research project in a secondary school molecular biology research course, this study not only contributes to the field of bacterial lipase research, but also offers students an authentic research experience at an early stage in their scientific careers.Support or Funding InformationThis work was funded by The Pingry School.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Use of a Synchronous Collaborative Learning Environment in a Face‐to‐Face Molecular Biology Classroom
Authors: Clauson‐Kozina Cheryl , Kunzweiler Greg 
Published: June 2021

As stated in “Vision and Change”, biology is an increasingly collaborative endeavor, requiring scientists to work well in teams and across disciplines. Yet, too often our classrooms are designed to be lecture‐based and don't provide sufficient opportunity for students to practice collaboration and teamwork. Here we describe a synchronous collaborative learning tool, MeTL, wherein students in face‐to‐face classrooms work together on problem sets and case studies. Rather than submitting a paper assignment at the conclusion of class time, the students instead work in an online environment that is accessible even after class has ended. Group members can see each other's work in real time and can thus provide assistance in creation of the group submission. Feedback provided to the students is viewable instantaneously, by all group members, and can be synced with Learning Management Systems. Students receive a grade for the work that was completed in addition to a scaled grade based on how well their peers felt they contributed to the activity of the day. We anticipate MeTL being useful to classrooms in many disciplines, but especially the molecular biology classroom.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Sustained teaching mentoring works and benefits mentors as well as those mentored: The Promoting Active Learning and Mentoring (PALM) Network
Authors: Wolyniak Michael J. , Wick Sue , Prunuske Amy J. , Peifer Mark 
Published: June 2021

Many instructors and instructors‐to‐be have heard of the value of using active learning to guide students to deep learning of course material. Some have attended workshops or several‐day immersion experiences to learn how to employ effective active learning activities. In spite of best intentions, plans to actually implement active learning techniques often collapse once an instructor gets caught up in the academic year. To address this situation, ASCB, in cooperation with other professional societies and academic groups and with NSF funding, established the Promoting Active Learning and Mentoring (PALM) Network. This program provides PALM Fellows with a one‐on‐one teaching mentoring relationship for at least one semester with a mentor experienced in active learning. Participants to date have come from various partner societies within the Network. Analysis of teaching behavior before and after mentoring provides evidence that Fellows have been able to increase their use of active learning in accord with the specific objectives they had identified. Interviews with participants indicate that Fellows gain more confidence in their ability to apply active learning principles in future coursework, and mentors appreciate the opportunity to reflect on, refine, and expand their own practice of active learning approaches.Support or Funding InformationPALM is supported through National Science Foundation RCN‐UBE #1624200.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Opening the Doors for Nuclear Gln3 Entry In Saccharomyces cerevisiae
Authors: Tate Jennifer J. , Rai Rajendra , Cooper Terrance G. 
Published: June 2021

The Gln3 and Gat1 GATA transcription activators are responsible for Nitrogen Catabolite Repression‐sensitive transcription in the yeast Saccharomyces cerevisiae. In the presence of excess, usable nitrogen, Gln3 is cytoplasmic and NCR‐sensitive transcription repressed. When cells are grown in nitrogen‐poor medium, treated with the TorC1 inhibitor, rapamycin, or glutamine synthetase inhibitor, methionine sulfoximine (Msx), Gln3 becomes highly nuclear and NCR‐sensitive transcription derepressed. In previous reports, nuclear Gln3 localization was concluded to be mediated by a single nuclear localization sequence, NLS1. Here we show that nuclear Gln3‐Myc13 localization is more complex than previously appreciated. We have identified three Gln3 sequences, other than NLS1 that are almost absolutely required for nuclear Gln3‐Myc13 localization. Two of these sequences exhibit characteristics of monopartite (K/R‐Rich NLS) and bipartite (S/R NLS) NLSs, respectively. Mutations altering these sequences are partially epistatic to a ure2Δ. In contrast, a third sequence, the Ure2 Relief Sequence (URS), exhibits no predicted NLS homology and is only necessary when Ure2 is present. Substitution of the basic amino acid repeats in the Ure2 relief sequence or phosphomimetic aspartate substitutions for the serine residues between those basic residues abolish nuclear Gln3‐Myc13 localization in response to both limiting nitrogen and rapamycin treatment. In contrast, Gln3 responses are normal in parallel serine to alanine substitution mutants. These observations suggest that Gln3 responses to specific nitrogen environments likely occur in multiple, genetically separable steps. At least one general step, i.e., that associated with the Ure2 relief sequence, appears to be prerequisite for Gln3 responses to specific stimuli, e.g., growth in poor nitrogen sources and rapamycin inhibition of TorC1.Support or Funding InformationNational Institutes of Health General Medical Sciences GM‐35642‐27This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Resveratrol and CXCR4 Mediated Migration of Brest Cancer Cell Lines
Authors: Cruz Kevin , Padilla Luis , Trossi Geraline , Muñoz Kevin , Robles Jose , Arroyo Gerardo , Suarez Edu , Ruiz Abigail 
Published: June 2021

In the United States, breast cancer is one of the most common types of cancer among women, being the second cause of cancer‐related deaths. Despite early detection methods and available treatments, breast cancer incidence is increasing worldwide. Some malignant cells have the potential to migrate and invade, metastasizing to distant organs and tissues. Current treatments include surgery, chemotherapy, and radiotherapy. These treatments usually produce side effects that have a negative influence in the quality of life of the patients. Additional studies are necessary to identify candidate natural compounds targeting specific molecules and receptors expressed in breast cancer cells. For example, CXCR4 is a G protein‐coupled receptor known to be one of the major proteins involved in metastasis of breast cancer. It mediates migration towards tissues expressing its specific ligand CXCL12. Natural compounds such as resveratrol, a plant polyphenol commonly found in grapes, peanuts and berries, among other, has shown anti‐proliferative, anti‐estrogenic, and antioxidant properties. We aimed to study the effect of resveratrol on CXCR4‐mediated cell migration. We hypothesize resveratrol will inhibit the ligand‐receptor relation between CXCR4 and CXCL12 decreasing the migration. To accomplish this, we conducted a wound healing assays, in Hs578t breast carcinoma‐cell line and MCF 7 breast adenocarcinoma‐cell lines treated with 25, 50, 100, and 200 μM of resveratrol in the presence of 100 ng/mL of CXCL12. As an internal control we used AMD3100, an antagonist of CXCR4. The results on Hs578t carcinoma cells did not show significant differences in the migration compared to the controls. However, this preliminary data suggest that there is a decrease in migration in the cells treated with 25 and 100 μM of resveratrol, compared to the AMD3100. Regarding MCF‐7 adenocarcinoma cells, we did not observe significant differences in cell migration. Additional experimental replicates are necessary to validate these results. In addition, we are currently studying other malignant and non‐malignant cell lines, including: CCD‐1097 and CCD‐1074 to identify cell specific responses to resveratrol, mediated by the CXCR4‐CXCL12 signaling pathway.Support or Funding InformationNIH‐NIGMS #2R25GM096955This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Non‐Canonical Wnt/Calcium Pathway Regulates the Migration and Function of Primary Mesenchyme Cells
Authors: George Alexander , McCann Tyler , Song Jia 
Published: June 2021

The highly conserved non‐canonical Wnt/Calcium Signaling Pathway (ncWnt/Ca2+) has been shown to regulate cell motility and epithelial to mesenchymal transition, implicating its role in various developmental processes and cancer invasiveness. We use the sea urchin Primary Mesenchyme Cells (PMCs), the only skeletogenic cells in developing sea urchin embryos, to examine directed cell migration. The working hypothesis is that PMC migration and PMC skeletal function are in part regulated by the ncWnt/Ca2+ signaling pathway. We have shown that disruption of elements downstream of the ncWnt/Ca2+ pathway using perturbative drugs decreased embryonic skeletal length as well as dramatically altered PMC migration patterns. In addition to producing similar skeletal and migratory defects, activation of Protein Kinase C (PKC) downstream of the ncWnt/Ca2+ pathway by Phorbol 12‐myristate 13‐acetate (PMA) also resulted in altered spatial expression of transcripts known to regulate PMC migration and function, as well as changed the expression levels of biomineralization genes. Similar phenotypes were observed in PMA treated embryos and the microRNA‐1 (miR‐1) knockdown embryos. MicroRNAs (miRNAs) are short, non‐coding RNAs that suppress gene expression by binding to the 3′UTR of target mRNA transcripts to repress translation. MiR‐1 is bioinformatically identified to bind to PKC and cdc42 of the ncWnt/Ca2+ signaling cascade, indicating its potential role in regulating the ncWnt/Ca2+ signaling pathway. This study identifies the molecular mechanisms of how the ncWnt/Ca2+ pathway impacts PMC directed migration and developmental function which may potentially contribute to our understanding of birth defects and cancer metastasis.Support or Funding Information

‐Senior Thesis Winter Session Scholars Award (Fall 2017)
‐Undergraduate Research Supplies‐And‐Expense Grant (Fall 2016, Fall 2017)
‐Science and Engineering Summer Scholar Award (Summer 2017)
This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.