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Fluorescence Activated Cell Sorting System

open

NIGMS - National Institute of General Medical Sciences

This proposal outlines a request to acquire a BD Biosciences FACSMelody Cell Sorter at Florida A&M University (FAMU), a state-controlled public institution. The BD FACSMelody has standard forward scatter (FSC) and side scatter (SSC) detection with nine fluorescence channels and the ability to sort 34,000 drops per second with up to 4-way sorting, enabling analysis and sorting of cell populations based on fluorescently labeled phenotypic markers and reporter gene expressions. The state-of-the-art equipment will greatly enhance research capability and capacity at FAMU, aiding several on-going NIH projects including: 1) engineering liver organoids for in vivo tissue restoration and ex-vivo drug screening, 2) enhancing efficacy of nanoparticles in pancreatic patient-derived xenografts (PDX) models, and 3) development of organoid systems tailored to probe lung cancer. The shared use equipment will improve research and biomedical activities at FAMU and promote collaborations with other biomedical researchers in Tallahassee. Aims of this project are to: (1) Acquire the BD Biosciences FACSMelody System at FAMU, (2) combine the instrument into research programs to promote and expand collaborations across disciplines between FAMU and other research-intensive institutions in Tallahassee and north Florida. This equipment item has been highly requested by the faculty, students, and staff members engaged in the areas of biomedical engineering, biology, pharmaceutical sciences, and food systems. The advanced functions of the FACSMelody are currently unavailable to FAMU researchers. Acquisition of the state-of-the-art system will foster long-term shared instrumentation use and promote collaboration between research groups across campus. As FAMU's west campus in Tallahassee's Innovation Park is currently without fluorescence-activated cell sorting (FACS) capabilities, the requested system would fill a major need to advance biomedical activities at the institution. To facilitate usage, the instrument will be housed in a newly constructed research building at the college of engineering that has a collaborative concept design. Graduate and undergraduate students in biomedical engineering, biological systems engineering, and pharmaceutics will incorporate use of equipment in their undergraduate research theses and Ph.D. dissertations. The requested system will improve workforce development, while also increasing success of external research funding, further improving the research ability of the FAMU faculty.

Up to $248K
2027-06-30
health research

Free to search & build · $99 one-time to unlock the application pack · No subscription

FOXE3 in Creating Lens Chromatin Architecture and Transcriptional Identity

open

NEI - National Eye Institute

The ocular lens serves as an excellent model for studying cell fate determination, as it consists of two distinct cell types derived from the surface ectoderm: lens epithelial and fiber cells. Lens development requires suppression of pro-neurogenic pathways, distinguishing it from other anterior placodes, such as the olfactory placode, which gives rise to neurons. Mutations in key transcription factors (TFs), including AP-2α, FOXE3, and PAX6, lead to overlapping developmental defects such as aphakia, incomplete lens separation, and fiber cell abnormalities, suggesting shared transcriptional targets and regulatory mechanisms. However, a major gap remains in understanding how FOXE3 functions as a transcriptional regulator and how its activity influences chromatin architecture to suppress pro-neurogenic gene expression during lens formation. Preliminary studies using CUT&RUN identified a de novo FOXE3-binding motif, with strong similarity to FOXD3, and revealed that FOXE3 is required to repress neurogenic gene expression in early lens cells. Additionally, RNA-seq analyses of Foxe3 mutant lenses demonstrated significant differential gene expression linked to pro-neurogenic pathways. This led to the hypothesis that FOXE3 functions as both a transcriptional activator and repressor, coordinating chromatin remodeling to establish and maintain lens cell identity. To test this, state-of-the-art chromatin and transcriptomic technologies, including single- nucleus multiomics (ATAC-seq + RNA-seq), CUT&RUN, and functional assays in chick embryos, to dissect FOXE3’s molecular mechanisms will be conducted. Aim 1 will identify FOXE3 transcriptional targets and determine how chromatin accessibility (DARs) and FOXE3-bound enhancers and promoters are altered in Foxe3 mutant lenses. This will be accomplished through the integration of single-nucleus multiomics and CUT&RUN data to map the FOXE3-dependent regulatory landscape. Aim 2 will determine the functional role of FOXE3 in cell fate decisions by (1) gain and loss of function assays in chick lens cells, (2) testing its ability to suppress neurogenesis in the developing chick retina, and (3) identifying FOXE3-interacting proteins via immunoprecipitation and mass spectrometry. The completion of these aims will provide mechanistic insights into FOXE3’s role in lens development, establish a FOXE3 DNA-binding motif, and identify targets of FOXE3 regulation. These studies aim to reveal how FOXE3 mutations contribute to congenital eye disorders such as aphakia, cataracts, microphthalmia, coloboma, and Peters anomaly. By elucidating the chromatin regulatory mechanisms underlying lens cell identity, this work has the potential to inform therapeutic strategies for human eye diseases linked to transcriptional dysregulation.

Up to $468K
2031-03-31
health research

Free to search & build · $99 one-time to unlock the application pack · No subscription

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