Karina Meiri - US grants

The significance of the neuronal growth cone in the formation of specific, connections during the development of the nervous system has been well established. This proposal describes experiments designed to investigate how one of the major components of growth cone membranes, the growth-associated 'protein GAP-43, is regulated in the growth cone, with the eventual goal of understanding how the growth cone is able to respond to extracellular signals and transduce them into intracellular events. The protein GAP-43 is regulated in two ways; its amount is regulated so that it is induced in neurons that are growing axons during development or regeneration, but interestingly it cannot be re-induced in the injured axons of neurons which are unable to regenerate, implying that it may perform a function intimately connected with axon growth. Second, its phosphorylation is regulated by other kinases and by autophosphorylation. These experiments use both primary neuronal tissue culture and a preparation of isolated growth cones to ask the following questions. Firstly, is GAP-43 phosphorylated by more than one kinase, and is that phosphorylation restricted to particular areas of the neuron? Second what are the extracellular signals that stimulate these kinases and does phosphorylation of GAP-43 regulate its subcellular distribution? Finally, GAP-43 is able to autophosphorylate; does this ability regulate its subcellular distribution or association with other proteins? or is GAP-43 able to phosphorylate other growth cone proteins, and so regulate their behavior. GAP-43 is associated with membranes even though it is not hydrophobic. These experiments will the nature of GAP-43 association with membranes and also the nature of a small group of three membrane proteins with which it is closely associated. Finally GAP-43 is a component of membrane that copurifies with actin microfilaments. The nature of this interaction and its regulation will be investigated. Answers to these questions will increase our understanding of the role of GAP- 43 in the growth cone, but also, because the experiments will develop a model system, will enable us to investigate more general questions about the role of signal transduction in the growth cone, with the eventual goal of using this knowledge to design strategies which circumvent the failure of some neurons (such as mammalian CNS) to regenerate successfully.

Several features of the nervous system specific 'growth-associated' protein GAP-43 suggest that it plays a fundamental role in cell biological processes common to axonogenesis and presynaptic function: It has been highly conserved during vertebrate evolution and is expressed in all neurons during development: Levels of GAP-43 are dramatically induced during axon outgrowth but, significantly, are not re-induced in injured axons of neurons that do not regenerate successfully: It is highly enriched in the growth cone where it associates with the membrane skeleton, the structure responsible for regulating the shape changes that are a crucial response to extracellular guidance cues: Finally, GAP-43 is a target for several Ca2+-dependent enzymes, including those responsive to extracellular signals, and levels of phosphorylated GAP-43 correlate with functional states of the growth cone. All of these features are consistent with a role for GAP-43 in the integration of extracellular signals within the growth cone, and suggest that understanding the molecular basis for GAP-43 function will further our understanding of the processes that direct axon growth. One powerful way to investigate GAP-43 function at the molecular level is to create a homozygous null mutant cell line in which transcription of GAP-43 is prevented, and then to compare its phenotype with cells that have further been transfected with GAP-43 cDNAs mutated at specific functional sites. Toward this end we have isolated and characterized a murine genomic clone for GAP-43 and used it to construct a replacement vector to knock out the GAP-43 gene at its native locus by homologous recombination. We will introduce the construct into the pluripotent embryonal carcinoma cell line P19 which differentiates into cholinergic-like neurons on treatment with retinoic acid, and will select lines homozygous fop disruption of GAP-43 transcription by their resistance to G418, a neomycin analog. Then, using a series of specific assays we have developed to investigate the role of GAP-43 in growth cone function, we will characterize the phenotype of the null mutant P19 cells with respect to attachment to laminin, regulation of neurite outgrowth and organization of the membrane skeleton and cytoskeleton. Finally we will transfect the P19 cells with GAP-43 cDNAs that have been mutated so that phosphorylation and calmodulin binding will be abnormal, and use these transfected cells to assess the relative contributions of phosphorylated GAP-43 and calmodulin to neurite outgrowth. The results generated by these experiments will provide information fundamentally important to our understanding of how axonogenesis is regulated during development and regeneration, and will enable us to begin to address the question of to what extent an axon needs to be structurally normal in order to form functional synapses.

tissue /cell culture

DESCRIPTION (provided by applicant) A fundamental aspect of developmental neurobiology that we still do not fully understand is how positional information coordinates with local regulation of neurogenetic programs to establish specific patterns in the brain. In vivo, the issue is experimentally accessible in the cerebellum. Inbred mice strains show complex yet invariant patterns of cerebellar foliation that is underpinned by a simple laminar structure comprised of a limited number of defined neuronal subtypes, facilitating characterization of the underlying mechanisms regulating neurogenesis. Examining defects in patterning of cerebellar cortex using mutant mouse models has confirmed that cerebellar patterning is intimately tied to the differentiation program of the neuroblasts, but how regulation occurs is still unclear. Our preliminary results suggest that certain molecules crucial for regulating membrane/cytoskeletal interactions during axon guidance also play important roles in regulating the neurogenic response to patterning information. One of these is the nervous system-specific protein GAP-43. We already know that GAP-43 is required in order for differentiated neurons to respond to signals that give rise to patterning in the CNS. For example, our GAP-43 knockout mouse fails to form either topographic maps in cortex, or telencephalic commissures, because in both cases GAP-43 (-/-) neurons are unable to respond to immunoglobulin superfamily (Ig-SF) mediated axon outgrowth and guidance signals. However the GAP-43 (-/-) mouse also has severe defects in cerebellar patterning that are evident before axonogenesis but during the time that neurogenesis is being regulated by extracellular patterning programs. The objective of this FRICA grant is to understand the molecular mechanisms underlying the disruption of patterning in the cerebellum that occurs when GAP-43 is absent. We describe 3 experiments: First, to determine the earliest stage of cerebellar development that requires GAP-43 function. Second to investigate whether 2 known neurogenic regulators in the cerebellum (bFGF and BDNF) can function when GAP-43 is absent - we already know that bFGF requires GAP-43 and that BDNF can stimulate GAP-43 phosphorylation in growing axons. Finally we will characterize how absence of GAP-43 affects transduction of lg-SF signals that regulate the cell cycle in differentiating cerebellar neurons. This research will be performed primarily in India as an extension on NIH grant# RO1 NS33118.

DESCRIPTION (provided by applicant): Engaging students in the biomedical sciences while they are still in high school is a critical first step toward educating a scientifically literate citizenry as well as initiating the pipeline that will eventually result in increased numbers of biomedical and health related professionals. This proposal is a collaboration between biomedical scientists at Tufts University School of Medicine, biology teachers at the Madison Park Technical and Vocational High School, the Boston Latin Academy and the Boston Latin School, all inner city Boston public high schools, and members of the Wright Center for Innovation in Science Education, also at Tufts. Its goal is to engage the imagination of 11th and 12th grade students who do not see the science of their real world experiences mirrored in the classroom. It accomplishes this goal by developing and disseminating a novel inquiry-based high school biology curriculum that focuses on biomedical research in the context of five 'great diseases'that challenge global health - infectious, neurological, cardiovascular, cancer and diabetes. A key element of the proposal is addressing the challenge of aligning content and process in the classroom to enable teachers to create the knowledge-centered classrooms that are critical for learning transfer. Accordingly the first aim is to first develop a learning community that will interactively guide the in-depth professional development underlying each of the 5 curricular modules. The learning community will then collaborate on the second aim;to develop curricular content and to generate deliverables for implementation of the curriculum in the classroom. This material, which will include a syllabus and web-based virtual and real interactive inquiry-base laboratory exercises, will be disseminated into the Boston Public School system and the broader educational community as part of the third aim. The fourth aim is to design and implement an evaluation strategy that will allow for ongoing revision during the project to ensure optimization of outcomes. The involvement of Tufts medical and biomedical graduate schools ensures that the program is highly sustainable and our final goal is to establish it as a model of how medical schools and school districts can interact to disseminate understanding of the biomedical research underlying disease at the high school level. PUBLIC HEALTH RELEVANCE (provided by applicant): Disappointingly, even after significant efforts at pedagogical reform, research continues to document adolescents steadily losing interest in science throughout high school. The economic impact of this disenchantment is significant and the life science/biomedical sector is growing slowly due to lack of a qualified work force, particularly in mid-level jobs. There is some cause for optimism: the same studies that demonstrate adolescent disenchantment with 'school'science show their interest in 'real world'science remaining stable over the same time frame. We postulate that students will only become engaged and able to visualize themselves as science professionals when they see the science of their real world experiences mirrored in the classroom. We address this hypothesis by generating an innovative curriculum designed to capture adolescent imaginations by focusing on the cutting edge research underlying 5 'great diseases'that impact global public health. We design the curriculum using a learning community consisting of practitioners of science and teachers of science whose goal is to collaborate to generate the content knowledge that will facilitate learning transfer in a knowledge-centered classroom.

The usefulness of any model experimental system depends on the extent to which it is able to mimic normal in vivo responses. A case in point is models that use controlled tissue culture environments to investigate how neurons grow. Most researchers fail to account for all key influences that neurons encounter in the embryo, particularly the mechanical behavior of embryonic tissue. This omission turns out to be critical. Evidence from the P.I.'s laboratory shows that unless the tissue culture environment mimics the mechanical status of the embryo, certain neurons are unable to respond normally to critical guidance cues. Very little is known about how mechanical signals impact neuronal responses to molecular guidance cues. These experiments address that deficit using a tissue culture model developed by the P.I. to account for the mechanical status of the embryo. They focus on a protein that plays a key regulatory role in how neurons respond to both mechanical and molecular signals. With the aim of unraveling mechanisms that coordinate neuronal responses to molecular guidance cues in the embryo, these investigators will use biochemical and molecular approaches to manipulate the mechanical environment. Their ultimate goal is to construct a tissue culture model with maximum in vivo relevance to both neurons and other mechanosensitive cells. The work has broader impact by virtue of two K-12 outreach programs established by the P.I. The first has sponsored over 200 inner city students to perform inquiry-based projects in research laboratories. The second has established professional development for their teachers. This award will expand opportunities for both students and their teachers to participate in authentic inquiry-based research, reinforcing the STEM educational pipeline.

DESCRIPTION (provided by applicant): The infectious disease (ID) burden has doubled in the US since its all-time low in 1980, in large part because of changes in human behavior. As a consequence, how ID will impact us in the future critically depends on how successful we will be in educating a literate citizenry able to make lifestyle choices that limit the impact of infection Despite this, ID rarely focuses in the high school classroom, in part because few teachers have been exposed to the topic, but also because the structure of teacher professional development conspires to segregate teachers from ID content experts, most of whom are in medical schools. To address this need we partnered with teachers to produce a comprehensive curriculum designed to engage high school students in the impact of ID on global society. Successful implementation of novel curricula requires intensive teacher support. In response we developed a professional development model in which near-peer content expert mentors and master teachers collaborate to model how to effectively teach complex concepts. In this way teachers learn strategies to confidently implement new material. We call this approach Modeling for fidelity. Here we propose to leverage this model and our ID curriculum as graduate-level courses that will disseminate the materials nationally, both face-to-face and online. The curriculum addresses the most recent national science education standards in biology and in literacy using an innovative framework focused on five core questions about ID - Why should I care about ID?; What does it mean to have an ID?; How does a microbe become a pathogen?; How do pathogens make us sick? How do I get better? It uses Socratic and Case- based approaches that foster active learning together with interactive activities and extended literacy components. It is specifically designed to bring inquiry based learning to schools with few resources or scheduling that precludes extensive hands-on experience. It is also flexible enough to incorporate any existing activities that further our learning objectives. Critically our graduat courses will also teach teachers skills to identify, locate and evaluate the reliable resources the need to remain current and keep the curriculum dynamic, as well as strategies to effectively adapt the curriculum for their specific classroom needs. Teacher participants in the courses will receive intensive support to pilot the curriculum in the classroom with diverse students. Gains in content knowledge and fidelity of implementation will be rigorously evaluated and best practices for dissemination established. The face-to-face graduate courses will prepare approximately 120 teachers from across the US, while the virtual learning platform will capture material from these courses and disseminate it freely on line. Crucially the virtual learning platform will also involve a direct mentoring component. Together they have the potential to reach significant numbers of teachers and students each year. Our approach meets the need for high-quality, integrated curricula designed for 21st Century learners while helping students build the deep understanding needed to make informed choices about their health.

CBET 1506883
Meiri, Karina F.

RAPID: Effectively Communicating EBOLA Information to Vulnerable Populations

This proposal is a RAPID response to improve education and communication about preventive measures related to Ebola. It targets 7th-12th grade middle and high school students and their teachers. Concerns about how the response to a disseminated outbreak of Ebola in the US would be managed are legitimate given that even a cursory examination of the Internet reveals the extent that misinformation is driving public awareness. Much of the US population is inadequately equipped to evaluate information provided by the media: More than half has difficulty in understanding and using health information, another segment is inherently suspicious of "official" information and a final segment has limited English language skills. We clearly need alternative routes to communicate critical information that: (a) are managed by scientists and clinicians who understand the nuances of infection and public health and who are able to explain clearly why information and directions can evolve over time, and that (b) occur in interactive settings where misinformation can be directly addressed. The high school classroom is the last place where entire cohorts of the US population gather to learn at the same time without regard to their health literacy, ideology or English language status, providing an ideal venue. This project will be developed by an established curriculum design team of biomedical scientists and Boston Public School teachers who have created a successful curriculum "The Great Diseases" that provides 10th-12th grade students with opportunities to acquire and apply problem-solving skills through collaborative inquiry. We will leverage this experience to generate a stand-alone suite of lessons (1 for middle school and 3 for high school) to address key scientific and public health questions related to Ebola. The project has two key elements: first to provide teachers with a complete platform of educative materials and interactive online support to enable them to implement the lessons in the classroom. Second, to involve families in the learning process so that students can act as a conduit for legitimate information. The resources developed will be freely available and accessible online.

The intellectual merit of the project relates to both students and teachers: 1) It provides students will a curated range of up-to-date information within a structure that enables them use active inquiry and problem solving to understand key issues, while teaching them the characteristics of reliable information and how to seek it out. Importantly, we will leverage our previous work with the Great Diseases curriculum that demonstrated how students that are engaged in the material communicate more with their families about the topics they are learning to structure the lessons to include family participation, asking families to evaluate an informational brochure students have created in the language spoken at home. 2) It provides teachers with resources and support in a familiar format, enabling them to effectively transfer this complex, dynamic information to their classrooms. The impact of the project relates to its significant potential for national and international impact. We have actively worked with over 150 teachers and about 12,000 students throughout the US and our approach to teaching and learning has successfully fostered student engagement in disease-related topics, knowledge of the health science underlying disease and healthy behaviors and critical thinking abilities and the improved self-efficacy that is important for continued independent learning. The largest gains were seen in inner city urban public schools and were independent of gender, ethnicity or English language learner status. We have also reached many more: Since 2013 people in 42 US states and 41 countries have accessed our freely available resources. More than 45,000 of these views lasted > 2 minutes, an indication of interest. We will leverage the breadth of this impact by providing an interactive blog for FAQs and by asking teachers to upload representative public health brochures to create a freely accessible, vetted, multilingual repository

? DESCRIPTION (provided by applicant): In 2014 many US adults lack key competencies in thinking analytically, evaluating and solving STEM-related problems, hence many STEM jobs particularly in life sciences and health go unfilled and more than 45% of the US population particularly from underserved communities, cannot manage their health care effectively. High school, where entire cohorts of the US population gather to learn together for the last time, is an obvious venue in which to develop these capacities. In response we partnered with teachers to develop a curriculum `The Great Diseases' for 10th-12th graders that targets these skills. No curriculum can be successful unless teachers have the skills to teach it, so we also developed a program in which scientists mentor teachers how to translate the science content of the curriculum into their classrooms. Teacher participants significantly increased student engagement and analytical and problem-solving abilities, and fostered confidence in learning about this kind of material - a critical element of health literacy. Importantly these gains occurred in a large number of diverse schools and were independent of teacher and also the gender and ethnicity of their students. In this project we aim to capitalize on the partnerships and evaluation tools we have already used successfully to create the curriculum and professional development model, to expand the scope of the curriculum by creating new activities more directly focused on the critical thinking skills we want to foster, and by expandin into new target audiences. We will therefore accomplish the following goals: (1) We will promote the analytical skills required for workforce preparation and health care management by developing three extensions to the core GD curriculum that will help teachers: address complex topic; stimulate claims evaluation related to health science research and identify valid new material they can incorporate into lessons to keep their curriculum current. (2) We will expand dissemination of the curriculum to pre-service teachers in collaboration with Bridgewater State University, the major provider of STEM and health education in Massachusetts by developing graduate level courses that will contextualize the content to classroom practice. (3) We will promote dissemination of the curriculum to in-service teachers nationwide by combining the curriculum and mentorship into a web-based course that combines online learning and virtual interactions between teachers and mentors. Then, in collaboration with Teach for America we will provide the course to teachers who work in challenging environments in urban or rural areas. We will evaluate the effectiveness of the program as well as knowledge gains and critical thinking abilities in program participants (teachers and their students). The proposed project is significant because teachers gain the skills to promote STEM competencies in their students, fostering workforce preparation and health literacy. It is innovative because The Great Diseases program provides a model for how medical school scientists can interact with teachers to influence curriculum and teacher development and because the web-based resource promotes nationwide dissemination.

This project will advance efforts of the Innovative Technology Experiences for Students and Teachers (ITEST) program to better understand and promote practices that increase students' motivations and capacities to pursue careers in fields of science, technology, engineering, or mathematics (STEM) by developing and testing a novel technology-rich high school bioscience curriculum and teacher professional development model. The new curriculum and associated online tools will promote development of critical competencies among students in the context of bioscience career-related investigations. The project will also develop a web-based application that will enable remotely located classrooms to participate in the new curriculum, develop new professional development approaches for high school teachers to support implementation of the new curriculum, and produce research data that will expand the knowledge base on factors influencing career selection and persistence among students with respect to bioscience careers. The curriculum will engage students in simulations that involve designing experiments, predicting results, and interpreting data. Video vignettes of bioscience professionals will be embedded in the curriculum to raise awareness of the wide range of careers related to the bioscience simulations. The associated teacher professional development will engage teachers in co-designing learning activities, piloting activities with students during summer activities, and an ongoing mentoring process with both in-person and virtual components.

This project is a collaborative effort involving scientists, educational researchers, and computer programmers from the Center for Translational Science Education at the Tufts University School of Medicine and teachers and biotech researchers in the Boston metropolitan area. The curricular approach being implemented and tested combines four strategies that have been shown to increase active learning, self-efficacy, and career awareness among students, and to effectively support teacher implementation of new curricula: a) Collaborative problem-based learning strategies that build career awareness by modeling how workforce teams collaborate in inquiry to solve authentic problems; b) Use of collaborative learning technologies that engage students and teachers in modeling, social persuasion, and problem-solving activities to foster teamwork and shared decision-making; c) A collaborative curriculum design process that engages teacher in developing curricula; and d) A teacher professional development approach that progresses through a series of in-person and virtual experiences that support teacher learning before, during, and after implementation of the new curriculum. A mixed methods research approach will be used to examine student outcomes related to disciplinary content knowledge and practices, student self-efficacy and outcome expectations, interests, and career goals. Pretest data and posttest data from innovation and comparison groups will be analyzed using multivariate techniques to examine relationships among outcome measures.