PROJECT SUMMARY The goal of this research is to examine how gene networks are re-used and specialized to generate repeated organs in the body, such as hairs, neurons, or muscles. Gene regulatory networks are assemblages of genes that transcriptionally regulate one another. These networks govern which proteins are produced in a cell to derive its own unique physical properties such as size, shape, or color. We have known for some time that many networks are re-used during development to generate various types of repeated organs across the body. This makes sense, as it would be much faster to re-use an existing network rather than completely build one from scratch. However, we have very few model systems in which the specifics of gene regulatory network re-usage can be studied. Experimental systems in which the usage of a network can be induced are particularly powerful for investigating how this process works. A second related problem is that of specialization. While repeated organs share many commonalities, it is the differences between them that we would often like to understand. How is a network that is used in hundreds of locations across the body individualized to generate a position-specific morphology? Here, we will examine a gene regulatory network that has been re-used to generate a new set of hair organs in Drosophila. Of particular interest, these hairs are highly specialized and are far larger than the other hairs that have been documented in this species. We have identified a gene that was critical to the re-use of this hair-forming network, and have made the remarkable discovery that we can induce hairs in species that lack these structures by simply activating this gene in the right developmental tissue. This finding led us to propose an exploratory set of experiments that will rapidly illuminate how this gene regulatory network was re-used and specialized: (A) We will examine the expression of genes known to be part of the ancestral larval hair network in the species which possesses these new highly specialized hairs. (B) We will seek out new genes that contribute to the spectrum of diversity of hair morphologies we have observed. (C) Using CRISPR/Cas9 and gene mis-expression technologies, we will determine whether genes identified to be important for generating specialized hairs are required or sufficient to do so. This research will inform the how processes of network re-use and specialization molecularly operate, providing insights into how specific genes and cellular processes may play roles in the regulation of organ size and shape.