Buildup Arsenal for Functional Study of Huntingtin in Drosophila. Zhen Xu¹, Yanning Rui¹, Zhihua Chen¹, Dongsheng Chen¹, Antonio Tito¹, Yamin Sun¹, Sheng Zhang^1,2,3. 1) Center for Metabolic & Degenerative Diseases , the Brown Foundation Institute of Molecular Medicine; 2) the Graduate School of Biomedical Sciences (GSBS); 3) Department of Neurobiology and Anatomy, The University of Texas Health Science Center (UTHEALTH) at Houston.

   Although Huntingtin gene has been extensive studied since its identification in 1993, with many proposed cellular roles, its normal function remains not well-defined. Characterizing an Htt homolog in Drosophila (dhtt), a simple yet genetically tractable system will complement the established mammalian models for Huntingtin studies. We had established a null-mutant (dhtt-ko) and performed preliminary characterization of its phenotypes. Further, we found that expression of human Htt could rescue the dhtt mutant phenotypes, suggesting the evolutionally conserved Htt functions and also supporting the use of fruit fly to study human Htt. To further take advantage of the abundant tools available in Drosophila for functional investigation of Huntingtin, we decided to develop a set of toolkit that would allow convenient detection and isolation of endogenous dHtt protein. Pacman method, the recently developed genome-tagging technique in Drosophila, allows efficient tagging and transformation of a large gene in its genomic DNA context, with the expression of the tagged gene still under the control of its native regulatory elements, thus ensuing the normal pattern and expression level of the tagged protein. Using this approach, we have successfully generated a set of genome dhtt transgenic lines with different fluorescence and epitope tags that allow convenient tracking and isolation of endogenous dHtt protein. This set of tools will greatly facilitate in vivo analysis and manipulation of this large protein in Drosophila. Establishing dhtt-ko mutant and a collection of genome-tagged dhtt lines enable us to carry out a detailed functional study of dHtt protein, which in turn should provide critical insights into the normal function of Htt and help decipher the mechanisms underlying HD.