Introduction: The role of the androgen receptor (AR) in disease pathology is determined by its mutational status and androgen hormone-dependent activity. There is a class of mutations in the AR gene that results in an expansion of a polymorphic trinucleotide-CAG repeat, coding for a polyglutamine tract in exon 1 of the AR gene. Normally, the CAG tract is 20 repeats, whereas in the disease known as spinal and bulbar muscular atrophy (SBMA) the repeat is expanded to >37 repeats. SBMA is a motor neuron disease directly linked to an androgen-dependent toxic gain of function caused by the polyglutamine expansion in the AR. The characteristics of SBMA are weakness, atrophy, and fasciculations of bulbar, facial and limb muscles that are attributable to degeneration of lower motor neurons in the spinal cord and brainstem. Understanding the molecular pathological mechanisms the AR polyglutamine polymorphism disorder could yield potential candidate therapeutic targets.
Aims of the Study: The aim of this project is to reveal mechanistic pathways involved SBMA pathogenesis. We employed an AR transgenic Drosophila genetic interaction screen of homologues of protein interactors of the expanded polyglutamine AR (polyQ-AR).
Results: A proteomics-coupled systems biology approach was used to characterize the proteome of wild-type (WT)-AR vs. polyQ-AR complexes. Through this process we have identified a number of androgen-dependent protein interactors known to be involved in RNA splicing. To investigate the role of polyQ-AR in RNA splicing, we undertook a genetic interaction screen using an UAS-polyQ-AR and UAS-WT-AR humanized-Drosophila transgenic lines and crossing them to UAS-RNAi or enhancer trap lines of fly homologues to our polyQ-AR protein interactors, in the presence or absences of androgen hormone ligand. The genetic interaction screen has identified a number of suppressors of the androgen-dependent polyQ-AR phenotype.Most interestingly, we identified HNRNPA2B1gene to be the strongest genetic interactor. No observable phenotypes have been noticed in WT-AR files, supporting the novel gain-of-function properties of the polyQ-AR in RNA splicing.
Conclusions: The proteomics and systems biology approach has allowed us understand AR functionality beyond its classical role as a transcription factor. Using Drosophila as a genetic interaction screening tool, we have robustly explored a novel mechanistic property of polyQ-AR. Through these investigations, it is now clear that the polyQ-AR is involved in RNA splicing pathways and has provided new dimensions to our understanding of SBMA disease and potential therapeutic avenues.