Zebrafish are an ideal system to discover genes that are essential for hematopoietic development in vivo. The spectrum of blood cell types and the genes known to regulate normal hematopoiesis are well conserved between zebrafish and mammals, suggesting that the regulatory mechanisms directing lineage specification and differentiation are similar across vertebrate species. The advantages to using zebrafish include external fertilization of eggs that enables easy access to the translucent embryos, rapid embryonic development, high fecundity, and ease of mutagenesis. Our lab exploits these characteristics by performing forward genetic screens to identify mutants with abnormal hematopoietic development. Mutations are induced by random insertion of a transposable gene trap element into the genome. The beauty of this technique is the transposon contains a RFP reporter, thus tagging cells in vivo and providing us with the dual benefit of being able to visualize and collect the RFP-positive cells as well as identify genes that are necessary for hematopoiesis. View diagrams explaining this technique.
Genetic instability is a hallmark of many cancers, including those of hematopoietic origin. We have identified a zebrafish mutant with pleiotropic developmental defects including widespread cell death, DNA damage, and abnormal numbers of chromosomes or rereplicated, unsegregated chromosomes. The rapidly dividing cells of the embryonic hematopoietic system are very sensitive to these events and mutants have severely decreased numbers of mature blood cells, with the residual cells being large in size. The gene mutated in this line is highly conserved between zebrafish and humans, making it an attractive tumor suppressor candidate. However, it remains to be seen whether this factor is a bona fide tumor suppressor in hematopoietic malignancies or how mutation of this gene results in genomic instability. Our laboratory is using a variety of methods to examine these questions.Top
transgenic zebrafish
Our goal is to create zebrafish harboring transgenic cell tracers, such as GFP, that can be used to track a specific population of cells during development and throughout adulthood. These transgenic lines are an extremely versatile tool because they allow for the live, real-time analysis of cells during development and purification of these cells, which can lead to cytological, RNA expression, and transplantation or cell tracing experiments.Top
The current paradigm of leukemogenesis is that multiple genetic insults cooperate to give rise to a malignant cell. The genetic composition of malignant cells often includes clonal chromosomal deletions, which contribute to transformation by deleting one or more tumor suppressor genes. However, the deleted regions are often very large making the identification of the critical genes difficult. Interestingly, many of the zebrafish genes that we have found to be essential for embryonic hematopoiesis have human orthologs located within commonly deleted regions. Working in concert with our clinical collaborators, we are examining adult hematopoiesis in zebrafish carrying single or combinatorial genetic alterations, including mutations or transgenic oncogene expression. Our goal is to establish models that mimic human diseases, to reveal novel pathways that are relevant to human cancer and gain insights into the pathogenesis of leukemia. They may also open perspectives for targeted therapies.Top