Dysfunctional autophagy in Hermansky-Pudlak syndrome associated lung fibrosis

Abstract

Hermansky-Pudlak syndrome (HPS) is a rare autosomal recessive disorder and HPS types 1, 2 and 4develop pulmonary fibrosis referred to as Hermansky-Pudlak syndrome associated interstitial pneumonia (HPSIP). We have previously reported spontaneous lung fibrosis in HPS1/2 double mutant mice primarily associated with surfactant accumulation, severe lysosomal stress and alveolar epithelial cell type II (AECII) apoptosis. In the current study we hypothesized that autophagy, a lysosome mediated degradation pathway might be altered, resulting in the excessive lysosomal stress that is reported under HPSIP conditions. Hence we analyzed the key autophagyproteins in the lungs of HPS1/2 mice and HPS1 patients and studied the effects of HPS1 knockdown in vitro in A549 cells. We observed a marked increase in LC3B lipidation and other key autophagy proteins in the lungs of HPS1/2 mice. However, immunohistochemistry on serial lung sections demonstrated a weak staining for LC3B within the AECII of HPS1/2 mice and in human HPS1 patient lungs. Immunogold labeling for LC3B confirmed these findings. Knockdown of HPS1 in vitro revealed an increase in LC3B lipidation with a simultaneous accumulation of p62, associated with an increase in pro apoptotic caspases (cleaved caspase3 and caspase 8). Overexpression of LC3B decreased the HPS1 knockdown induced p62 and cleaved caspase 8 accumulation while rapamycin treatment did not show the same effect. We conclude that loss of HPS1 protein results in impaired autophagy and apoptosis that are restored by exogenous LC3B and that dysfunctional autophagy might play a critical role in the development and progression of HPSIP.