Establishment, Growth and Regeneration of Zebrafish Fins

In vertebrates, the adult form emerges from the embryo by mobilization of precursors, also known as adult stem cells. What different cell types these precursors give rise to, how many precursors establish the tissue or organ, and how they divide to establish and maintain the adult form remain largely unknown. In this thesis work, I examined how the zebrafish establish and maintain one of its adult organs, the adult fin. In the first part of my thesis, I focused on one specific type of cells, the pigmented cells in the fin, to address questions concerning their establishment and maintenance, with a variety of clonal and lineage analyses. Early embryonic labeling with lineage-marker bearing transposons showed that all classes of fin melanocytes (ontogenetic, regeneration and kit-independent melanocytes) and xanthophores arise from the same melanocyte-producing founding stem cells (mFSCs), while iridophores arise from distinct precursors. Additionally, these experiments showed that on average, 6 and 9 mFSCs colonize the caudal and anal fin primordia, and daughters of different mFSCs always intercalate to form the adult pattern. Labeled clones are arrayed along the proximal-distal axis of the fin, and melanocyte time-of-differentiation lineage assays showed that while most of the pigment pattern growth is at the distal edge of the fin, significant growth also occurs proximally. This suggested that leading edge Melanocyte Stem Cells (MSCs) divide both asymmetrically to generate new melanocytes, and symmetrically to expand the MSCs and leave quiescent MSCs in their wake. Clonal labeling in adult stages confirmed this and reveals different contributions of MSCs and transient melanoblasts during growth. These analyses build a comprehensive picture for how MSCs are established and grow to form the pigment stripes of the adult zebrafish fins. In the second part of this thesis, I examined the lineages other than the pigmented cells in the fin for their establishment and maintenance. Again, I used transposon-based clonal analysis to identify the lineage classes that collaborate to make the adult zebrafish caudal fin, an organ that emerges during the larval-to-adult metamorphosis. We identified 9 distinct lineage classes, including, epidermis, melanocyte/xanthophore, iridophore, intra-ray glia, lateral line, osteoblast, dermal fibroblast, vascular endothelium, and resident blood. These lineage classes argue for distinct progenitors, or organ founding stem cells (FSCs), for each lineage. Furthermore, they show that lineages retain fate restriction throughout growth of the fin. Thus, these results show distinct FSCs for the 4 neuroectoderm lineages, and disprove the notion that the dermal fibroblasts are progenitors for fin ray osteoblasts. Similarly, the shared lineage for artery and vein argues against two distinct origins for the fin vasculature from the dorsal aorta or cardinal vein. Zebrafish fins share with salamander limbs the remarkable ability to regenerate following partial amputation. Like in salamander limbs, transdifferentiation of cells or lineages in zebrafish fin regeneration blastema is often postulated. Regeneration of lineages derived from single progenitors or FSC showed that there is no transfating or transdifferentiation between these lineages in the regenerating fin. This result shows that, the same as in growth, lineages retain fate restriction when passed through the regeneration blastem