Identification of a Phosphorylation-Dependent Nuclear Localization Motif in Interferon Regulatory Factor 2 Binding Protein 2

Background - Interferon regulatory factor 2 binding protein 2 (IRF2BP2) is a muscle-enriched transcription factor required to activate vascular endothelial growth factor-A (VEGFA) expression in muscle. IRF2BP2 is found in the nucleus of cardiac and skeletal muscle cells. During the process of skeletal muscle differentiation, some IRF2BP2 becomes relocated to the cytoplasm, although the functional significance of this relocation and the mechanisms that control nucleocytoplasmic localization of IRF2BP2 are not yet known. // Methodology/Principal Findings - Here, by fusing IRF2BP2 to green fluorescent protein and testing a series of deletion and site-directed mutagenesis constructs, we mapped the nuclear localization signal (NLS) to an evolutionarily conserved sequence 354ARKRKPSP361 in IRF2BP2. This sequence corresponds to a classical nuclear localization motif bearing positively charged arginine and lysine residues. Substitution of arginine and lysine with negatively charged aspartic acid residues blocked nuclear localization. However, these residues were not sufficient because nuclear targeting of IRF2BP2 also required phosphorylation of serine 360 (S360). Many large-scale phosphopeptide proteomic studies had reported previously that serine 360 of IRF2BP2 is phosphorylated in numerous human cell types. Alanine substitution at this site abolished IRF2BP2 nuclear localization in C2C12 myoblasts and CV1 cells. In contrast, substituting serine 360 with aspartic acid forced nuclear retention and prevented cytoplasmic redistribution in differentiated C2C12 muscle cells. As for the effects of these mutations on VEGFA promoter activity, the S360A mutation interfered with VEGFA activation, as expected. Surprisingly, the S360D mutation also interfered with VEGFA activation, suggesting that this mutation, while enforcing nuclear entry, may disrupt an essential activation function of IRF2BP2. // Conclusions/Significance - Nuclear localization of IRF2BP2 depends on phosphorylation near a conserved NLS. Changes in phosphorylation status likely control nucleocytoplasmic localization of IRF2BP2 during muscle differentiation

An Interdisciplinary Approach to Online Genetics Education

This dissertation presents an interdisciplinary approach to online genetics education. Chapter I provides an overview of remote education and a brief history of genetics. While the COVID-19 pandemic prompted a turn to emergency online education, exposure to this form of delivery has led to increased intentional online education. Chapter II focuses on computer and technology use in healthcare by nursing students. This study investigated nursing students’ perspectives using a modified version of the Pretest for Attitudes Toward Computers in Healthcare (PATCH) assessment scale to explore students’ ideas about computers and technology in healthcare delivery. According to the PATCH scale, the majority of the class had “enthusiastic” or “idealistic” views of computers and students supported technologies, such as electronic medical records, for use in practice. In Chapter III, nursing students in an interdisciplinary healthcare genetics class completed eight online learning activities designed using the Community of Inquiry (CoI) framework. A mixed-methods survey measured students’ engagement and rating of the online activities. Students embraced online learning activities which were interesting to them, aided studying, and were applicable to future nursing practice. In bonus results, students who scored the highest affinity for technology in healthcare rated online learning activities significantly higher than others. Chapter IV presents a systematic review of the literature for online education for rare genetic diseases. The review described the studies by subject matter, mode of delivery, and target learners. Although results were limited, studies included quality, interdisciplinary online education developed for both healthcare professionals and the public. Drawing on the results from Chapters II, III, and IV, Chapter V presents a Model for Online Genetics Interdisciplinary Education (MOGIE). The model integrates principles of community learning, online design, and essential topics in biopsychosocial aspects of rare disease. The model can be applied to online/hybrid-designed courses and provides comprehensive education pertaining to a rare disease and related disciplines such as epidemiology, bioethics, genetic counseling, and clinical care. In summary, this dissertation examines online genetics education in terms of student attitudes, engagement, and a review of the literature to culminate in a proposed educational model