bluuebunny/crossref_metadata_2025_split huggingface dataset:
Transmembrane (TMEM)‐163 protein is a novel zinc transporter
Authors: Cantrell Quinlan W , Silva Joshua , Nguyen Cathleen , Hildebrand Leanne D , Rivas Tiffany , Shoemaker Ryan , Rojas America , Cuajungco Math P 
Published: November 2023

A mutation or deletion of the MCOLN1 gene which codes for the Mucolipin‐1 (TRPML1) ion channel protein causes the human lysosomal storage disorder called Mucolipidosis IV (MLIV). An elevation of chelatable zinc, particularly in lysosomes, is observed in cells affected by MLIV compared to normal cells. The mechanism behind the zinc buildup in MLIV cells remains elusive; however, evidence suggests that it is due to the failure of non‐functional TRPML1 to mediate the release of zinc from the lysosomes or failure in lysosomal exocytosis. We recently identified TMEM163 as an interacting partner of TRPML1. We hypothesize that TMEM163 is a transporter that facilitates zinc flux in normal cells, but that the loss of TRPML1 function in MLIV cells leads to abnormal TRPML1‐TMEM163 protein interaction such that the cells uncontrollably accumulate zinc. To infer function as a zinc transporter, we first studied the membrane topology of heterologously expressed TMEM163 protein. Contrary to its predicted orientation, we found that both the N‐ and C‐terminus regions of TMEM163 lie at the extracellular rather than intracellular milieu. To test functional similarities between TMEM163 and several known zinc transporters (ZNTs), we cloned the cDNA of TMEM163 and specific ZNTs into the pBI‐CMV dual‐expression construct (containing a minimal CMV promoter vector). Single cDNA constructs were created as controls. The single gene and dual gene constructs were then heterologously expressed in human embryonic kidney (HEK)‐293 cells. Twenty‐four hours post‐transfection, treated and control HEK‐293 cells were then exposed to zinc chloride (ZnCl2, 100 mM) for 3 and 5 hours. Spectrofluorometric end‐point assay using the zinc‐specific Fluozin‐3 AM dye showed that the single TMEM163 expression in cells produces intracellular zinc accumulation upon 5 hours of ZnCl2 exposure, while co‐expression of TMEM163 with ZNT4, but not other ZNTs, showed a significant synergistic effect on intracellular zinc levels at both time points tested. Overall, these results suggest that TMEM163 may serve a similar role as those of the ZNTs in transporting zinc, but that it is an influxer. Future studies relating to TMEM163‐TRPML1 protein interaction should provide new research opportunities to further understand the involvement of zinc in the underlying neuropathology and progressive cell degeneration observed in MLIV disease.Support or Funding InformationNIH R15‐NS070774, NSF MCB‐0920127, and NIH T34‐GM008612‐20
Structural Determination of the BBSome Core Complex
Authors: Aoba Takuma 
Published: November 2023

Bardet‐Biedl syndrome (BBS) stems from failed assembly of BBS proteins into a functional protein complex called the BBSome. The BBSome consists of eight BBS proteins (1–2, 4–5, 7–9 and 18) and constitutes a key component of intraflagellar transport system in eukaryotic primary cilia. Lack of a functional BBSome results in cilium‐related symptoms such as retinal degradation, polydactyly, obesity, kidney dysfunction, and other disorders. To better understand mechanisms of BBSome assembly and function, we have analyzed the structure of a BBSome assembly intermediate, the BBSome core complex (BBS2, 7, and 9), by a combination of cryo‐electron microscopy and crosslinking coupled with mass spectrometry (XL‐MS). The structure of the BBSome core complex revealed a parallel arrangement of BBS2 and 7 through a coiled‐coil interaction and a perpendicular arrangement of BBS2 and 9 through interactions in their α‐helical domains. Unnatural amino acid cross‐linking and mutagenesis identified specific residues at the binding interfaces between the core complex components as well as BBS‐linked mutations that disrupt the complex. Thus, these structural studies not only visualize the BBSome core complex and reveal its assembly process, but they also provide insight into the molecular basis of Bardet‐Biedl syndrome.Support or Funding InformationBarry M. Willardson
Dysregulation of V‐ATPase Function Impairs Lysosomal Acidification in a Mouse Model of a Rare Lysosomal Storage Disease, INCL
Authors: Bagh Maria B , Peng Shiyong , Chandra Goutam , Zhang Zhongjian , Mukherjee Anil B 
Published: November 2023

Lysosomes are the major organelles for intracellular digestion. They contain >50 hydrolases that require acidic pH for optimal function. The acidification of the intracellular organelles, including the lysosome, is a fundamental biological process of pivotal importance to eukaryotic cells. Lysosomal acidification is regulated by v‐ATPase, a multi‐subunit protein, which is localized to the lysosomal membrane. The cytosolic V1‐sector hydrolyzes ATP generating energy for the membrane‐anchored V0‐sector, which imports protons from the cytoplasm to the lysosomal lumen for acidification. Lysosomal acidification is defective in virtually all lysosomal storage disorders (LSDs) in which neurodegeration is a devastating manifestation and in common neurodegenerative disorders like Alzheimer's and Parkinson's. Although it is known that v‐ATPase maintains the endosomal/lysosomal acidic pH, the mechanism(s) underlying defective lysosomal acidification in these neurodegenerative disorders remains largely unknown.Palmitoylation (also known as S‐acylation) is the only reversible lipid modification of proteins. In this process, a 16‐carbon fatty acid (predominantly palmitate) is attached to polypeptides via thioester linkage. Dynamic palmitoylation (palmitoylation‐depalmitoylation) regulates the function of many proteins, especially in the central nervous system. It requires the catalytic actions of palmitoyl‐acyltransferases (PATs), which catalyze palmitoylation and those of palmitoyl‐protein thioesterases (PPTs), which catalyze depalmitoylation respectively. Inactivating mutations in the CLN1 gene encoding palmitoyl‐protein thioesterase‐1 (PPT1), a lysosomal depalmitoylating enzyme, underlie a devastating neurodegenerative LSD, called infantile neuronal ceroid lipofuscinosis (INCL). Using Cln1−/− mice, which mimic INCL, as a model, we uncovered that dynamic palmitoylation of V0a1 subunit of v‐ATPase promotes its interaction with the adaptor proteins for trafficking from the ER (where it is synthesized) to the lysosomal membrane. Like a relay race, V0a1 is transferred from one adaptor to the other for its sequential transport from the ER‐Golgi to the lysosomes. In Cln1−/− mice, lack of Ppt1 causes mistargeting of V0a1 to the plasma membrane instead of the lysosomal membrane, dysregulating lysosomal acidification, which contributes to neuropathology. Our findings reveal an unanticipated role of Cln1/Ppt1 in regulating lysosomal acidification and suggest that varying factors suppressing v‐ATPase activity may contribute to pathogenesis of other LSDs and some common neurodegenerative diseases.Support or Funding InformationIntramural program of the Eunice Kennedy Shriver National Institute of Child Health and Human Development of the National Institutes of Health (NIH).
Students’ Understandings of their Ethical Obligations to their Cadavers after their First Anatomy Lab: A Qualitative Study
Authors: Fortunato John , Wasserman Jason , Hankin Mark 
Published: November 2023

First year medical students completed video‐recorded reflections as an assignment for their ethics course. They were prompted to reflect on both their initial feelings as they started their first dissection and their ethical responsibilities to their cadavers. These reflections were analyzed using a schema akin to Grounded Theory, a qualitative methodology that allows data driven findings to systematically emerge from narrative data without specifying hypothesis in advance. This approach sheds light on themes related to complex social and emotional experiences that quantitative research methodologies (like surveys) have difficulty capturing because they necessarily reduce students’ holistic experiences to a narrower set of generalizable constructs measured with predetermined multiple‐choice options.This research showed that initial reactions to first dissection tend to follow one of two patterns: Some students showed an initial sense of concern, worry, or uneasiness as they entered the cadaver lab and began the dissection. However, the technical aspects of dissection eased their concerns as time went on. In contrast, other students report being focused on the technical demands and learning objectives such that they had no reservations upon entering the lab or beginning the dissection. However, at some point later on, an awareness of cadaveric humanity and the gravity of donation came upon them. In regards to ethical obligations, a responsibility to “respect” the cadaver, though for various underlying reasons, was nearly unanimous amongst students. Additionally, an obligation to maintain donor privacy, an obligation to treat the cadaver “like a patient,” and reverence toward the donor's family members were among the more commonly referenced reasons for respect of the cadaver. Other commonly cited ethical responsibilities included a duty to be academically prepared for the anatomy dissection in an effort to learn as much as possible from the gift of cadaveric donation. These reported ethical obligations, when understood against the broader backdrop of a number of other potential ethical duties, suggests that students’ ethical relationship with the cadaver is developmental and could be encouraged in its growth and maturity.This analysis of student narrative provides unique insight into the first‐year medical student's sense of ethical obligation and emotional response in the cadaver lab. Moving forward, anatomical sciences faculty can use this research in an effort to tailor the methodology in which they address medical students’ sensitivities and emotional awareness within anatomy programs.Support or Funding Informationnone
Measurement of branched chain amino acids (BCAA) in milks and protein supplements
Authors: Hubble Kyle , Swanson Derek , Keenan Kelly 
Published: November 2023

The amount of money spent on nutritional supplements by Americans is in the billions per year. Protein supplements, which are often made from whey in milk, boast of high levels of BCAA. There are mixed results to suggest that BCAA play a role in muscle recovery but they remain a popular supplement. The goal of this project was to develop an undergraduate laboratory experiment in which levels of BCAA are measured in various milks and supplements. These milks and supplements were hydrolyzed to release the amino acids and BCAA were measured using the leucine dehydrogenase enzyme assay which specifically measures the sum of these three amino acids. In addition, the level of arginine was measured using the Sakaguchi assay. In order to confirm these results, the levels of these amino acids was tested using reverse phase high performance liquid chromatography (RP HPLC). It was necessary to quantify the BCAAs and the quality of the data was assessed by spiking the samples with one of the BCAA and measuring the % recovery. The results are consistent by both methods which demonstrates that the leucine dehydrogenase is a valid method to measure BCAA. While the protein levels for the supplements are similar to the claims of manufacturer's, the level of BCAAs are markedly lower. This experiment is a suitable undergraduate biochemistry project.Support or Funding InformationThis project was supported by an REU grant from Stockton University.
Active learning in a flipped classroom: implementation and assessment
Authors: Barral Ana Maria , Ardi Veronica C 
Published: November 2023

The inverted (“flipped”) classroom approach uses class time to discuss difficult topics, clarify misconceptions, and have students apply the concepts through active learning, the act of doing. Active learning approaches have shown to enhance learning of the sciences, particularly in biology. In a flipped classroom, lecture materials are given in recorded videos, allowing passive learning to take place outside the classroom. This study examined whether there was an increase in learning and retention in students who completed active learning activities in the classroom within a flipped classroom design, compared to the traditional lecture approach, in a General Biology for Majors course. From the course learning outcomes, we identified a number of basic biology concepts for assessment purposes.To measure learning, pre‐ and post‐tests with 15–20 questions were administered to students. Questions in the tests were scored on the Bloom's Taxonomy scale by three independent researchers. The majority (93%) of the questions involved knowledge and comprehension, with approximately 33% of the questions also requiring higher cognitive skills such as application and analysis. The percent of correct responses was calculated for both flipped (n=2, 15–21 students per class) and control (n=2, 15–28 students per class) classrooms. The means of correct percentages were higher in students in the flipped classroom as compared to traditional classroom, with a statistically significant difference in the post‐test results. Students in the flipped classrooms performed better (mean = 58.3% correct) than those from the traditional classroom (mean = 33.0%) in the higher cognitive questions. Currently, we are deepening the assessment of student learning by using qualitative “Think Aloud” methodology to evaluate students’ application of concepts to problem solving. We also administered surveys to examine students’ perceptions of the study materials and in‐class activities. Most students (n=32) agreed that the videos and learning activities were helpful for their understanding of the material (3.97 and 4.09 on a Likert scale of 1–5, with 5 being most helpful). As for the individual in‐class activities, students overwhelmingly agreed that 1) they understood the concept better, 2) they believed the activities would help them retain the concepts in the future, 3) the activities sparked their interest in biology, and 4) the activities aided in critical thinking. The average response ranged between 4.00–4.67 for the 7 activities, using a 1–5 Likert scale with 5= strongly agree. Altogether these data suggest that students gain knowledge by active learning and flipped classroom.Support or Funding InformationNational University COLS Teaching Grant
Mentor‐Mentee Interaction and Laboratory Social Environment: Do They Matter in Doctoral Students’ Publication Productivity?
Authors: Ynalvez Marcus Antonius , Ynalvez Ruby , Ramirez Enrique 
Published: November 2023

We explored the social shaping of science at the micro‐level reality of face‐to‐face interaction in one of the traditional places for scientific activities ‐‐ the scientific lab. We examined how doctoral students’ perception of their: (i) interaction with doctoral mentors (MMI), and (ii) lab social environment (LSE) influence productivity. Construed as the production of peer‐reviewed articles, we measured productivity using total number of articles (TOTAL) and number of articles in top journals (TJ). Face‐to‐face survey data were obtained from n=210 molecular biology Ph.D. students in selected universities in Japan, Singapore, and Taiwan. To summarize the original 13 MMI and 13 LSE semantic‐differential items that we used to measure students’ perceptions, a principal component (PC) analysis was performed. The result was a parsimonious set of 4 MMI PCs and 4 LSE PCs. To identify which PCs influenced publication counts, we performed a Poisson regression analysis. Although perceived MMI was not linked to productivity, perceived LSE was linked: Students who perceived their LSE as intellectually stimulating and as having privacy reported high levels of productivity. Our findings not only highlight how students’ perception of their training environment factors in the production of scientific output, they also carry implications for improving mentoring programs in science.Support or Funding InformationNational Science Foundation Social Behavioral and Economic Sciences (Award#0830109)
Improving undergraduate biochemistry curriculum using threshold concepts: an analysis of student understanding
Authors: Freese Samantha , James Kristen , Lewis Jennifer , Loertscher Jennifer , Minderhout Vicky 
Published: November 2023

Threshold concepts (TCs) are concepts that, when mastered, represent a transformed understanding of a discipline without which the learner cannot progress. TCs have been identified in many fields with the goal of redesigning curricula to best benefit the learning of the student. Student data was collected for three of the five biochemistry TCs (steady state, the physical basis of interactions, and the thermodynamics of macromolecular structure formation). Responses were analyzed to determine ubiquitous and nuanced themes in student understanding. Integrative themes including electrostatic properties of molecules, entropic and enthalpic contributions, free‐energy, and kinetics of regulatory systems were investigated due to their importance in biochemistry learning. Our analysis reveals that students often exhibit limited understanding of these concepts, thus they lack the depth of knowledge to fully integrate ideas and apply them to varied biochemical contexts.Support or Funding InformationNSF‐DUE‐1224868
Activatable COX‐2 Probes in Optical Imaging of Cancer
Authors: Uddin Md. Jashim , Crews Brenda C , Ghebreselasie Kebreab , Daniel Cristina K , Marnett Lawrence J. 
Published: November 2023

Conventional imaging methods, such as angiography, computed tomography, magnetic resonance imaging and radionuclide imaging, rely on contrast agents that are “always on”. A unique aspect of optical imaging is that fluorescent probes can be designed to be activatable, so that they are only “turned on” under certain conditions. Probes that are activatable and also accumulate in the tumor in a targeted fashion could be ideal for detection of superficial tumors or tumors that can be accessed by endoscopy. We developed fluorocoxib A, the 1st COX‐2 targeted optical imaging probe, which displayed a high degree of selectivity of uptake in the COX‐2‐positive tumors over COX‐2‐negative tumors. Having an established proof‐of‐concept compound for COX‐2‐targeted imaging, we sought to expand the applicability of this approach by designing activatable pro‐fluorescent nitroxide probes that are capable of visualization of cancer with high signal‐to‐noise ratios. We synthesized a series of nitroxide derivatives of fluorocoxib A and B, containing a carboxy‐X‐rhodamine (ROX) fluorophore closely linked to a TEMPO or PROXYL nitroxide moiety. These probes exhibited selective COX‐2 inhibition in purified enzymes and intact cells, and showed significantly reduced fluorescence due to efficient quenching of the excited electronic state of the fluorophore by the nitroxide radical. Upon radical trapping in cancer cells, these COX‐2‐targeted probes demonstrated enhanced fluorescent emission, making them effective in detecting COX‐2 in tumors in vivo. Thus, this novel strategy allows target‐specific delivery of quenched agents into tumor cells and subsequent on‐site fluorescence activation by radical trapping allowing selective visualization of tumor in vivo. This technology can be widely adapted for background free detection of inflammatory and neoplastic diseases in preclinical and clinical settings.Support or Funding InformationThis work was supported by research grants from the National Institutes of Health number CA89450 (LJM).
How Well Do Nigerian Heath Staff Interpret Weight Trajectory On Growth Charts In Children Under 6 Months?
Authors: Wright Charlotte M , Ezeofor Ifeyinwa , Garcia Ada 
Published: November 2023

Growth charts are widely used in developing countries to assess nutritional status over time in young infants, but little is known about how well they are interpreted.Objectivesto test the extent to which health staff consider the pattern of weight gain over time, or just current weight centile and how this is affected by use of the usual Road to Health (RTH) chart or the new World Health Organization (WHO) chart format.MethodChild Health staff in two teaching hospitals and 4 government health centres in Enugu, Nigeria completed a questionnaire on chart use, plotted sample weights on RTH and WHO charts and answered questions on 4 plotted charts, 2 in RTH and 2 WHO format. A factorial design was used so that each respondent rated 4 of 12 permutated charts, presented in one of 3 versions of the questionnaire. The permutatations comprised: 3 different weight trajectories between birth and age 6 months (slow=−2SD fall; average= SD unchanged; fast=1–2 SD rise) towards two attained weights (average=50th centile; small= 2nd–3rd centile) each presented on RTH and WHO format. No respondent viewed the same growth pattern more than once in either format. Responders were asked to rate each the weight pattern shown as slow, steady or rapid and whether it would cause them to increase monitoring or refer out (concern).ResultsOf 233 staff approached, 222 (95%) completed questionnaires. Most were hospital‐based (78%); 32% medical doctors, 13% nutritionists and 54% nurses; 46% had more than 10 years experience. Most respondents used charts often and confidently. Plotting accuracy was generally good and best on the more familiar RTH chart.Respondents rated 856 charts. Rating of weight gain was generally poor, but varied significantly with size and chart type. On the RTH chart respondents were more likely to recognize slow weight gain in a small (35%) than an average child (19%) and more likely to recognize rapid weight gain in an average (53%) than a small child (18%), but overall their responses were not better than chance (p= 0.052 and 0.15).On the WHO charts slow weight gain was better recognised, though still more in a small (65%; p<0.001) than an average (40%; p=0.002) child. Recognition of fast weight gain was still poor: 53% also rated a rapidly gaining small child as having slow weight gain. In a binary logistic regression model, final size more strongly predicted a slow weight gain rating (OR=2.4; p<0.001) than an actual slow weight gain pattern (OR 1.8; p<0.001). Only a minority felt that any of the weight gain patterns merited concern; this was significantly related to the actual weight gain pattern on WHO, but not on the RTH charts.ConclusionMany health staff seemed unable to interpret weight trajectory or recognize slow weight gain and were more influenced by current weight than actual weight gain pattern, though the new WHO format improved recognition.Support or Funding InformationIfeyinwa Ezeofor was supported by a Ford Foundation scholarship