Accumulation of Mutant α1-Antitrypsin Z in the Endoplasmic Reticulum Activates Caspases-4 and -12, NFκB, and BAP31 but Not the Unfolded Protein Response

Abstract

In ␣ 1 -antitrypsin (␣1AT) deficiency, a polymerogenic mutant form of the secretory glycoprotein ␣1AT, ␣1ATZ, is retained in the endoplasmic reticulum (ER) of liver cells. It is not yet known how this results in liver injury in a subgroup of deficient individuals and how the remainder of deficient individuals escapes liver disease. One possible explanation is that the "susceptible" subgroup is unable to mount the appropriate protective cellular responses. Here we examined the effect of mutant ␣1ATZ on several potential protective signaling pathways by using cell lines with inducible expression of mutant ␣1AT as well as liver from transgenic mice with liver-specific inducible expression of mutant ␣1AT. The results show that ER retention of polymerogenic mutant ␣1ATZ does not result in an unfolded protein response (UPR). The UPR can be induced in the presence of ␣1ATZ by tunicamycin excluding the possibility that the pathway has been disabled. In striking contrast, ER retention of nonpolymerogenic ␣1AT mutants does induce the UPR. These results indicate that the machinery responsible for activation of the UPR can distinguish the physical characteristics of proteins that accumulate in the ER in such a way that it can respond to misfolded but not relatively ordered polymeric structures. Accumulation of mutant ␣1ATZ does activate specific signaling pathways, including caspase-12 in mouse, caspase-4 in human, NFB, and BAP31, a profile that was distinct from that activated by nonpolymerogenic ␣1AT mutants.In the classical form of ␣1AT 2 deficiency, the mutant ␣1ATZ molecule is retained in the endoplasmic reticulum (ER) of liver cells rather than secreted. There is an 85-90% reduction in ␣1AT levels in the blood and body fluids. This deficiency affects ϳ1 in 1800 live births and results in the premature development of pulmonary emphysema in adult life. Chronic liver disease develops in a subgroup of homozygotes, usually becoming evident during childhood. There is also an increased incidence of hepatocellular carcinoma later in life. Emphysema is caused by a loss-of-function mechanism whereby lack of ␣1AT in the lung allows proteolytic destruction of the connective tissue matrix. In contrast, liver injury appears to involve a gain-of-toxic-function mechanism whereby the accumulation of mutant ␣1ATZ in the ER damages liver cells (1).Nevertheless, relatively little is known about the factors that predispose the "susceptible" subpopulation of PIZZ individuals to liver disease and/or protect the remainder of the PIZZ population from liver disease. By using skin fibroblast cell lines from PIZZ individuals with or without liver disease engineered for expression of ␣1ATZ, we have shown previously that there is a lag in ER degradation of mutant ␣1ATZ in cells from PIZZ individuals with liver disease (2). These results provided evidence that the response of cells to the accumulation of this mutant protein in the ER, particularly the degradative machinery, could play a role in determining the susceptibility to liver disease among ...

Alpha-1-antitrypsin mutant Z protein content in individual hepatocytes correlates with cell death in a mouse model

“…Several previous studies have shown that the intracellular accumulation of the a1AT mutant Z protein is associated with caspase activation and other markers of apoptosis in in vitro systems. 4,5,8,9 Furthermore, when in vivo model systems are examined, an increased rate of hepatocellular proliferation is present, suggesting a compensatory response to increased hepatocellular death. 3 However, when either a homozygous ZZ human liver or a model transgenic mouse liver has been examined, widespread evidence of hepatocellular death has not been readily apparent.…”

“…[3][4][5] Our laboratory and others have reported a series of studies that have begun to examine the mechanism of liver cell injury in a1AT deficiency. [3][4][5][6][7][8][9] The data suggest that an …”

“…[3][4][5]11 Immunoblots were performed as previously described in triplicate with antibodies previously noted to be useful in these systems. 4,5,8 For the Jo2 antibody treatment, mice were given a single intraperitoneal injection of 6 m of anti-Jo2 antibody (DB Bioscience). Indomethacin was also used, as noted previously.…”

“…[3][4][5] Our laboratory and others have reported a series of studies that have begun to examine the mechanism of liver cell injury in a1AT deficiency. [3][4][5][6][7][8][9] The data suggest that an accumulation of the a1AT mutant Z protein in the ER of hepatocytes activates autophagy, causes mitochondrial injury and mitochondrial autophagy, and is associated with caspase activation and apoptosis. We have further shown that experimentally increasing the quantity of the polymerized a1AT mutant Z protein in the liver is directly related to increased injury.…”

Intracellular Processing of 1-Antitrypsin

“…However, several reports have convincingly demonstrated that the UPR is not activated either in numerous cultured cell lines (15,45,46) or in transgenic mice (47). The exact mechanism(s) by which the protein-accumulated protein leads to cellular injury is still unknown, but is currently under intense investigation.…”