2004 — 2008 |
Keiper, Brett |
N/AActivity Code Description: No activity code was retrieved: click on the grant title for more information |
Function of Tissue-Specific Eif4e Isoforms in Caenorhabditis Elegans @ East Carolina University
The critical question in embryonic development is, "How do nondescript (undifferentiated) cells change in character to become the very specialized cells that make up the vital organs of a fetus?". It is already known that the production of new and unique proteins in individual cells very early in embryo development contributes to their taking on the character of adult tissue types. Processes that introduce (synthesize) new proteins ultimately direct cells to form distinct organ systems such as brain, muscle, gonad and gut. It is this regulation of new protein synthesis in embryos that is the focus of this research. The soil-dwelling nematode worm, Caenorhabditis elegans, develops from a fertilized egg to an organized multicellular embryo in a manner very similar to that of higher animals. Fortunately these worms have a far simpler body plan made up of just a few muscles, neurons, digestive and reproductive organs. They also grow to adulthood in just one day. More to the point, the critical genes and gene products expressed at the earliest stages of development are very similar in this simple model system. Among these products are messenger RNAs (mRNA) that are stored in egg or embryo. These mRNAs are recruited to ribosomes (the protein synthesizing machines of the cell) at just the right moment to be used as a template for the synthesis of regulatory or structural proteins. The mRNA recruiting factors have been studied for many years, and were thought to be identical in every type of cell in the body. However, Dr. Keiper's laboratory has recently shown that unique forms (isoforms) of these factors are found in certain tissue types, most notably those that are undergoing developmental changes in embryos. The current project addresses the role of specialized protein synthesis factors in selecting which mRNAs are used in embryonic cells. An understanding of their unique activities will begin to uncover how different cells begin to express a unique set of proteins. This cell-specific translational control is part of the internal program that takes embryos from a uniform mass of cells to a complex adult organism. Students and postdoctoral researchers involved in this project will learn how informational biomolecules (DNA, RNA and protein) elaborate the changes that make embryogenesis a fascinating dynamic process.
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0.915 |
2009 — 2014 |
Keiper, Brett |
N/AActivity Code Description: No activity code was retrieved: click on the grant title for more information |
Translational Control of Growth and Apoptosis in C. Elegans Development by Initiation Factor Isoforms @ East Carolina University
New gene expression is required for cells to begin to grow and divide (proliferation) or to alter their fate (differentiation). The development of egg and sperm (gametes) likewise involves crucial periods of gene expression and differentiation. Early cells divide and then differentiate into either sperm or oocytes. Gene transcription is active early in gametogenesis, but becomes silenced as cells enter meiosis, and thus mRNAs that have been stored during the proliferative phase become the sole means to produce new proteins. Developmentally important mRNAs become translated into protein on ribosomes in the oocyte, spermatocyte or embryo, and the new proteins direct either continued differentiation or cell death (apoptosis). This research project is focused on proteins called eIF4 factors, which contact the mRNAs to be translated as the first step in recruiting them to the protein synthesis machinery. Specifically, the activities of eIF4 factors that lead to new protein synthesis in gametes and embryos will be investigated. Animal model systems such as the soil-dwelling nematode worm, Caenorhabditis elegans, are essential in this research because they are generally simpler than human embryos. The C. elegans embryo develops from a fertilized egg to an organized multi-cellular embryo in a manner very similar to that of higher animals. Fortunately these worms have a far simpler body plan made up of just a few muscles, neurons, digestive and reproductive organs. More importantly in the present context, the critical genes required are very similar in this simple model system. Because protein synthesis mechanisms are well conserved in animals, research findings using C. elegans will shed light on gene expression in vertebrate gametes and embryos, where such methods are not feasible or practical. Broader Impact and Educational Benefit: This project directly impacts the education of a minority and a female graduate student who are completing Ph.D. thesis research in Dr. Keiper's laboratory, as well as Masters and undergraduate students from ECU's Biology and Chemistry departments. As in the past granting period, the project also involves collaborators and students from other universities and high schools with an interest in molecular gene expression during development. The research capitalizes on the lab's experience in the biochemistry of mRNA translation and provides broad laboratory training in molecular techniques, genetics and transgenesis to young scientists. These students find the C. elegans system both tractable and significantly more accessible intellectually than mammalian systems. They receive the greatest educational benefit from first-hand involvement in new discoveries as well as opportunities to present their findings and publish their accomplishments in scientific journals. In recent years, several students who participated in this project have taken positions in the biotechnology industry or entered graduate research/medical professional academic programs.
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0.915 |
2017 — 2020 |
Keiper, Brett Lee, Myon Hee Henderson, Melissa (co-PI) [⬀] |
N/AActivity Code Description: No activity code was retrieved: click on the grant title for more information |
Mrna Selection by Eif4e Isoforms and Their Sequestering Factors. @ East Carolina University
This project explores how "undecided" stem cells progress to a defined cellular identity by changing the proteins they make within the cell. During the early development of an organism, cells must decide their path to become an organ that performs a specialized function. The messenger RNAs (mRNAs) within each cell are blueprints for the proteins to be made (translation). Such mRNAs are used selectively to introduce only the appropriate specialized functions. This project will define the mechanisms that translation factors use to carefully select mRNAs to decode at appropriate times and places, yielding useful information about how cells determine their final fate. High school, undergraduate, Masters, and PhD students will gain hands-on experience and knowledge of the most contemporary bioinformatics, recombinant, molecular and biochemical techniques, making them competitive for entering the workforce while pursuing science careers.
Protein synthesis is highly regulated in early animal development, occurring differently in each cell/tissue type to create functional organs with appropriate architecture and cellular activities. Transcriptional regulation of genes during development is well studied by many labs. However, transcriptional patterns often don't match the spatial and temporal protein requirements. The actual appearance of proteins is dictated largely by mRNA selection/translational control, whose mechanisms are more obscure. Over many years the Keiper lab has studied mRNA selectivity by unique forms of the eIF4 translation factors. The hypothesis is that isoforms of eIF4E and eIF4G positively and selectively recruit dormant mRNPs to ribosomes for efficient protein synthesis. This project focusses on two germ line eIF4E's (IFE-1 and IFE-3) in C. elegans, a simple worm that is an ideal genetic/transgenic model for reproductive development. The first goal of this project is to use resolved polysome RNA Seq, a technology developed in the PIs lab, to identify all RNAs that rely on individual IFEs for efficient translation. The second uses CRISPR/Cas9 to fluorescently tag each IFE to determine its localization in vivo, and allow characterization of its storage and retrieval complexes by MALDI-tof proteomics. The result: a blueprint of dynamic and "whole genomic" protein synthesis, which promotes fertility, reproduction, embryo and organ development
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0.915 |
2021 — 2024 |
Keiper, Brett |
N/AActivity Code Description: No activity code was retrieved: click on the grant title for more information |
McA: Post-Nuclear Granules Traffic Mrnas Through Helicases and Initiation Factors to Set Their Translational Fates @ East Carolina University
This project investigates how animal cells specialize to give rise to gametes (eggs and sperm). This Mid-Career Award will enable the PI, who is an expert in biochemistry, to gain expertise in molecular genetics and microscopy by working with a collaborator at a different institution. The project will also train students in classical RNA-protein biochemistry approaches as part of summer lab courses co-hosted by the PI and collaborator.
Transcriptional regulation of genes during development is insufficient to account for the observed temporal and spatial patterns in proteins required for differentiation and development. Instead, many mRNAs critical for development are also regulated at the level of translation initiation. Translation initiation is controlled by initiation factors such as eukaryotic initiation factor 4 (eIF4). In previous work, the PI’s lab studied how unique forms of the eIF4 protein selectively translate mRNA during the development of Caenorhabditis elegans, a simple nematode. The hypothesis for this proposal is that isoforms of eIF4E known as IFE-1 and IFE-3 selectively recruit dormant mRNAs to ribosomes for efficient protein synthesis. The first goal of this project is to use resolved polysome RNA Seq, a technology developed in the PIs lab, to identify all RNAs that rely on IFE-1 or IFE-3 for efficient translation. The second uses CRISPR/Cas9 technology to fluorescently tag each IFE to determine its localization in vivo, and allow characterization of its storage and retrieval complexes by proteomics.
This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.
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0.915 |