Carnegie Science | Fall 2018 12 New Tool for Female Reproductive Genetics There are several different types of vectors, molecules that ferry new genetic material into an organism. This simplified diagram shows a viral vector carrying a new gene into a cell nucleus where it can be replicated. Image reprinted with permission from yourgenome.org, Genome Research Limited www.yourgenome.org/facts/what-is-gene-therapy Steven DeLuca is a Howard Hughes Medical Institute postdoctoral fellow in the Spradling lab. SUPPORT: The Helen Hay Whitney Foundation and Howard Hughes Medical Institute, in addition to Carnegie, supported this work. he fruit fly Drosophila melanogaster is a powerful model organism for studying animal and human development and disease, and there are many tools to genetically modify its cells. One tool is called the Gal4/UAS two-component activation system. This tool is a biochemical method used to study gene expression—the process of turning on a gene—and gene function. Although it has been a mainstay of Drosophila genetics for twenty-five years, it only functions effectively in nonreproductive cells, not in egg-producing cells. It has not been known why. Now, Carnegie’s Steven DeLuca and Allan Spradling have discovered why, and they have developed a new tool that can work in both cell types. Genetics published the research in June 2018. The GAL4 gene is a transcription factor. Transcription factors encode proteins that turn on genes. The Gal4 protein recognizes an activator (called UAS), which can activate a gene of interest. A special version of UAS (called UASp) was made at the Department of Embryology in 1998 to work during egg-cell development. But the fact that different tools are needed for nonreproductive cells and egg-forming cells has been a major limitation. The original UAS, called pUASt is a vector—a molecule that ferries foreign genetic material into another cell—and it contains a promotor called Hsp70. As the name suggests, promotors are bits of DNA that initiate or promote gene activation. Researchers have developed several varieties to improve its expression. The promotor is a member of a family of proteins in most organisms that are important for protein folding and for protecting cells from stress. Protein- folding mechanisms are vital to life and to understanding diseases. DeLuca and Spradling studied the differences between the UASp and UASt promoters. Their research agreed with previous reports that UASt worked better than UASp in all nonreproductive tissues while UASp worked better in the female egg-producing system. They also looked at why UASt was weakly expressed in the female reproductive system. The evidence indicated that noncoding RNA molecules (called piRNA) orchestrated the suppression that limited UASt. They then looked at where these UASt-piRNAs originated and what was happening and then attempted to create a new version of the UAS vector that works well in both the nonreproductive cells and the egg-producing system. DeLuca explained, “We hypothesized that Hsp70 piRNAs might recognize UASt RNA to initiate piRNA silencing. To prevent Hsp70 piRNAs from recognizing UASt RNA, we trimmed down the UASt vector’s nucleotides—basic units of DNA and RNA—to be shorter than a single piRNA. We went from 213 nucleotides to 19 nucleotides. We named this shortened variant ‘UASz,’ because we hoped it would be the last one anyone would make!” The scientists found that UASz was expressed about four times higher than UASp at all stages in the egg-producing system.  CytoPlasm Vector binds to cell Virus vector NEW GENE Vector Packaged into Vesicle Vesicle Breaks down Releasing VeCtor Nucleus New Gene Injected into nucleus