RESEARCH
The study of development integrates and synthesizes multiple fields in biology: molecular and cell biology, physiology and metabolism; evolutionary biology. It also integrates several fields in medicine including regenerative medicine and cancer. We primarily use the model organism Drosophila melanogaster as our experimental system for asking fundamental questions in developmental biology. The unprecedented richness of knowledge about Drosophila plus the unrivalled toolkit of experimental methods available make this model system, arguably, the most powerful one in biomedical research. We focus on two classes of molecules whose existence only emerged recently: non-coding RNAs and microproteins. We ask questions about the unique and special functions these molecules have on development. Fundamental knowledge about these molecules will provide new opportunities for disease diagnosis and treatment.
NON-CODING RNAS
A large portion of the transcriptome is comprised of RNAs that do not code for proteins and yet they regulate gene expression. These include short interfering RNAs (siRNAs), microRNAs (miRNAs), piwi interacting RNAs (piRNAs), and long noncoding RNAs (lncRNAs). For over 25 years, we have been working to understand the molecular mechanisms by which these non-coding RNAs function in Drosophila development. Starting with our discovery that Drosophila were able to perform RNAi, our early efforts focused on siRNAs and miRNAs. More recently, our efforts have been focusing on lncRNAs, of which over 2,000 distinct genes synthesize these RNAs in Drosophila. We have also returned to studying siRNAs by exploring their possible connection to the neurodegenerative disease Amyotrophic Lateral Sclerosis or ALS. This disease involves several key RNA-binding proteins such as TDP-43 that alter functions in motor neurons. We are looking at how the RNAi pathway may interact with this system during disease onset.
MICROPROTEINS
Microproteins are a new class of proteins heretofore overlooked in proteomic annotations owing to their small size (< 100 amino acids). This led to the misclassification by genome projects of microprotein-coding mRNAs as being lncRNAs. Nicknamed the "Dark Proteome”, microproteins are believed to number in the 100’s to 1,000’s depending on the species, with mammals having among the most abundant numbers. Virtually nothing is known about their biochemical activities and organismal functions. As part of our efforts to study lncRNAs, we have found a minority of those annotated as lncRNAs encode for microproteins. One such microprotein, Dafcin, is a 21 amino acid amphipathic helix that structurally is most similar to the fusion peptide in the hemagglutinin protein coating the surface of influenza virus. HA peptide induces negative curvature of host endosomal lipid membranes as part of the mechanism by which the virus escapes the endosome of host cells. We have found that Dafcin also induces negative curvature in oocyte lysosomes, not to penetrate, but rather to limit lysosome growth.
GENOMIC TOOLS FOR GENE DISCOVERY
A cornerstone of modern biology is the continual development of new and robust experimental technologies for gene discovery. We have been active in technology development for almost 30 years, beginning with the development of RNAi as a genetic tool in Drosophila. More recently, we have worked to make CRISPR/Cas9 mediated genome editing a more reliable and efficient method in Drosophila. We have also adapted a self-cleaving ribozyme from Schistosoma to use as a gene knockout tool in Drosophila. When the DNA fragment encoding this 83 nt catalytic RNA is inserted into a gene, it effectively prevents accumulation of the gene's mRNA since the ribozyme cleaves the mRNA in two. The ribozyme is effective at destroying noncoding RNAs as well as isoform-specific RNAs. We are currently developing the method for high-throughput screens as well as making the system able to turn off genes in a targeted manner.
