Fibroblast-lineage cell state transitions during zebrafish fin regeneration - PROJECT SUMMARY The applicant proposes two years of fellowship support to complete their Ph.D. training in developmental biology. Research and training will focus on learning and using computational and quantitative imaging approaches to understand mechanisms of cell state transitions underlying organ size control during zebrafish fin regeneration. Adult zebrafish fins, including their bony ray skeletons, regenerate to the original size and shape irrespective of the extent or type of injury. Injury activates intra-ray fibroblasts that migrate distally to form the blastemal mesenchyme. The distal mesenchyme, or “niche” cells, then upregulates the transcription factor Dachshund (Dach) and Wnts that promote regenerative outgrowth. Niche cells gradually deplete over the course of regeneration, slowing outgrowth. Outgrowth stops as niche cells deplete below a critical level, coincident as the fin regains its initial size and shape. Size restoration is thus dependent on the number of fibroblasts activated upon injury, which may simply reflect the volumetric capacity of tapering bony rays. Long-finned zebrafish models show that fibroblast-lineage bioelectric signaling regulates niche depletion by an unknown mechanism. Dynamic Dach and Snail mesenchymal marker expression, single cell transcriptomics, and photoconvertible lineage tracing suggest that niche cells transition back to a mature fibroblast state to repopulate the intra-ray space. However, epithelial marker expression and proliferation studies suggest there may be distinct proliferative and non-proliferative niche populations. This observation indicates that quiescent distal niche cells may transition through a transit-amplifying state and then finally to a mature fibroblast state. The applicant will (1) use existing single cell RNA-sequencing data to identify fibroblast-lineage cell states, (2) use mathematical modeling to determine whether membrane voltage acts as a bioelectric signal to control proliferation of transit amplifying cells, and (3) probe whether quiescent cells transition into transit-amplifying cells through cell labeling and progeny analysis. The applicant will pursue these studies as a member of the Stankunas laboratory at the University of Oregon, a leading center for developmental biology and zebrafish research. A leading external expert will facilitate rigorous mathematical modeling and training. Specialized advanced coursework, formal research presentations, participation within the Developmental Biology Training Program, remote and in-person interactions with expert mathematical modelers, and targeted conference attendance will complement research- based training. These combined efforts will support the applicant’s transition to a postdoctoral position integrating computational and wet lab approaches to developmental biology questions. Long term, this training will support the applicant’s career goal of becoming an independent researcher while facilitating the inclusion of disabled persons in research.
