PROJECT SUMMARY Tandem duplicate genes are a core part of human genome structure. At the same time, variations in gene copy number occur frequently in humans, leading to diseases such as autism, schizophrenia, and bleeding disorders, as well as variation in individual traits. Despite this growing relevance, biologists have a limited understanding of the direct effects of tandem gene duplication. A simple prediction is that a tandem duplication of an entire gene would double gene expression. Instead, recent studies have found that young tandem duplicate genes are more than twice as active as single copy genes, producing an excess of RNA and protein over the expected 2-fold level. The tandem structure of the gene is responsible for the excess, not the change in copy number, suggesting that gene proximity and/or specific regulatory sequences may be responsible. Population-level studies indicate that such large changes in gene expression from tandem duplicates are often highly detrimental. However, despite the obvious relevance for cellular and organismal function, it is not clear how this excess occurs and if it is dependent on a specific aspect of gene structure. The purpose of this proposal is to identify genetic factors that cause excess expression from tandem duplicate genes. Aim 1 will test the hypothesis that divergence between duplicate gene copies does not eliminate excess expression. Aim 2 will test the hypothesis that the excess expression of tandem duplicates is distance-dependent, as predicted by a computer simulation of gene transcription. Aim 3 will test the hypothesis that the excess expression is mediated by insulator sequences, genetic factors with roles in other kinds of neighbor interactions. To pursue these aims, a team primarily composed of undergraduate students will apply innovative gene- construction techniques and precise quantitative expression assays in Drosophila flies to provide new insight into how tandem genes work.