Xiaohong Gu: Accumulation and Cellular Clearance of IAPP and Proteotoxicity of ATTR in Drosophila Models

Abstract

Proteins’ functions are dependent on their three-dimensional (3D) structure. Under certain circumstances, proteins misfold and form aggregates, sometimes leading to amyloidosis. 

Islet amyloid polypeptide (IAPP) builds up amyloid in the pancreatic islet of patients with type 2 diabetes (T2D). We utilized the GAL4-UAS system to express human proIAPP (hproIAPP), human IAPP (hIAPP) and mouse IAPP (mIAPP) in the brain of Drosophila melanogaster. In transmission electron microscope (TEM), we observed an accumulation of non-fibrillar aggregates in fat body tissue surrounding the brain. TEM tomography used for ultrastructure analysis revealed a spherical shape and a 5-fold twinning structure composed of two crystal packings:  the body centered tetragonal (BCT) structure and the triclinic structure.

Our biological systems have developed the ability to assist in folding and removing non-functional proteins. We directed the expression of hproIAPP and hIAPP to the 16 ventral lateral neurons (LNvs) and monitored intracellular responses to protein aggregation. We observed that overexpression of hproIAPP and hIAPP significantly reduced the number of LNvs over time. Further studies showed that expression of hproIAPP and hIAPP did not trigger ER stress and apoptosis but resulted in an accumulation of ubiquitinated aggregates, autophagosomes, and lysosomes, indicative of activation of aggrephagy. Overexpression of hproIAPP/hIAPP in flies with the Gstd-ROS reporter results in ROS generation, which can contribute to cell death. 

Systemic amyloidosis is a rare condition where amyloid deposits occur in multiple organ systems. Deposits of wild type transthyretin (TTRwt) cause the most prevalent form of systemic amyloidosis, while TTR mutations can result in familial forms of the disease. The clinical profile in hereditary ATTR amyloidosis differs in the age of onset, tissue distribution, symptoms, disease penetrance, and prognosis. The Gal4-UAS system was applied to produce flies expressing human TTRwt and single mutant TTR. The Hand-C-Gal4 driver directs the TTR expression in the cardioblasts of the heart. We found the expression of TTRV30L, TTRV30M, TTRA109S, and TTRL111M had a significant impact on cardiac parameters. The Nrv2-Gal4 driver directs the TTR expression to the central and peripheral nerve cells. Expression of TTRV30L, TTRV30M, TTRL55P, and TTR L111M by Nrv2-Gal4 altered the activity or circadian rhythm in the fly. The results showed that different mutations give rise to different phenotypes. 

In summary, our Drosophila melanogaster models provide valuable insights into amyloidosis and allow for cellular and organ analysis.