Intracellular Inclusions Containing Mutant α1-Antitrypsin Z Are Propagated in the Absence of Autophagic Activity

Abstract

Mutant ␣ 1 -antitrypsin Z (␣ 1 -ATZ) protein, which has a tendency to form aggregated polymers as it accumulates within the endoplasmic reticulum of the liver cells, is associated with the development of chronic liver injury and hepatocellular carcinoma in hereditary ␣ 1 -antitrypsin (␣ 1 -AT) deficiency. Previous studies have suggested that efficient intracellular degradation of ␣ 1 -ATZ is correlated with protection from liver disease in ␣ 1 -AT deficiency and that the ubiquitin-proteasome system accounts for a major route, but not the sole route, of ␣ 1 -ATZ disposal. Yet another intracellular degradation system, autophagy, has also been implicated in the pathophysiology of ␣ 1 -AT deficiency. To provide genetic evidence for autophagy-mediated disposal of ␣ 1 -ATZ, here we used cell lines deleted for the Atg5 gene that is necessary for initiation of autophagy. In the absence of autophagy, the degradation of ␣ 1 -ATZ was retarded, and the characteristic cellular inclusions of ␣ 1 -ATZ accumulated. In wild-type cells, colocalization of the autophagosomal membrane marker GFP-LC3 and ␣ 1 -ATZ was observed, and this colocalization was enhanced when clearance of autophagosomes was prevented by inhibiting fusion between autophagosome and lysosome. By using a transgenic mouse with liver-specific inducible expression of ␣ 1 -ATZ mated to the GFP-LC3 mouse, we also found that expression of ␣ 1 -ATZ in the liver in vivo is sufficient to induce autophagy. These data provide definitive evidence that autophagy can participate in the quality control/degradative pathway for ␣ 1 -ATZ and suggest that autophagic degradation plays a fundamental role in preventing toxic accumulation of ␣ 1 -ATZ.3 a monomeric 394-amino acid glycoprotein, is synthesized and secreted primarily by liver cells. It is a prototypic member of serine protease inhibitor (serpin) superfamily proteins and the most abundant of the circulating serpins. The principal role of ␣ 1 -AT in serum is to protect lung tissues from destructive proteases (elastase, cathepsin G, and proteinase 3) released by neutrophils during inflammation. Some genetic alterations in ␣ 1 -AT are responsible for defective secretion and thus cause serum ␣ 1 -AT deficiency (1-3). The most common causal mutation found in Caucasian populations is the replacement of Glu-342 by Lys that characterizes the Z mutant of ␣ 1 -AT (␣ 1 -ATZ). This substitution is sufficient to cause an abnormality in folding early in the secretory pathway with retention of the mutant ␣ 1 -ATZ molecule in the ER of liver cells. Homozygotes for the ␣ 1 -ATZ mutation (PIZZ) are characterized by serum levels of ␣ 1 -AT that are ϳ10 -15% of those in the general population and are susceptible to two major target organ injuries. Destructive lung disease/emphysema in adults is due to a loss-of-function mechanism. Chronic liver disease often first discovered in childhood, but also affecting adults, is due to a gain-of-toxic-function mechanism in which liver cell injury results from the hepatotoxic effects of retained ␣ 1 -ATZ...

Autophagic disposal of the aggregation-prone protein that causes liver inflammation and carcinogenesis in α-1-antitrypsin deficiency

“…28 The results demonstrated a significant delay in disposal of ATZ in the atg5-null background compared with the wild-type background. There was progressive accumulation of insoluble ATZ with very large inclusions extending from the ER into the cytoplasm in many of the atg5-null cells.…”

Endoplasmic Reticulum Stress: A New Pathway to Induce Autophagy

“…To prevent the toxicity, cell degrades accumulated ATZ by ERAD. Recently, proteins involved in autophagy were shown to participate in the removal of ATZ aggregates, indicating that autophagy also is utilized to degrade this protein; [8][9][10] however, the relationship between autophagy and ER stress had not been clear. We recently found that autophagy is indeed induced in response to ER stress.…”

Autophagy in the liver

“…58 In cells overexpressing ATZ, autophagosomes seem to be closely juxtaposed with the ATZ aggregates, and suppressing autophagy retards the removal of ATZ. 59 Furthermore, in the yeast model of ATZ degradation, it has also been found that aggregated ATZ is targeted to the autophagy pathway for degradation. 60 It is thus conceivable that autophagy can play an important role in the liver pathogenesis of AT deficiency.…”