Grp78, Grp94, and Grp170 interact with α1-antitrypsin mutants that are retained in the endoplasmic reticulum

In alpha(1)-antitrypsin (alpha(1)-AT) deficiency, a mutant form of alpha(1)-AT polymerizes in the endoplasmic reticulum ( ER) of liver cells resulting in chronic hepatitis and hepatocellular carcinoma by a gain of toxic function mechanism. Although some aspects of the cellular response to mutant alpha(1)-AT Z have been partially characterized, including the involvement of several proteasomal and nonproteasomal mechanisms for disposal, other parts of the cellular response pathways, particularly the chaperones with which it interacts and the signal transduction pathways that are activated, are still not completely elucidated. The alpha(1)-AT Z molecule is known to interact with calnexin, but, according to one study, it does not interact with Grp78. To carry out a systematic search for the chaperones with which alpha(1)-AT Z interacts in the ER, we used chemical cross-linking of several different genetically engineered cell systems. Mutant alpha(1)-AT Z was cross-linked with Grp78, Grp94, calnexin, Grp170, UDP-glucose glycoprotein: glucosyltransferase, and two unknown proteins of similar to 110-130 kDa. Sequential immunoprecipitation/immunoblot analysis and coimmunoprecipitation techniques demonstrated each of these interactions without chemical crosslinking. The same chaperones were found to interact with two nonpolymerogenic alpha(1)-AT mutants that are retained in the ER, indicating that these interactions are not specific for the alpha(1)-AT Z mutant. Moreover, sucrose density gradient centrifugation studies suggest that similar to 85% of alpha(1)-AT Z exists in heterogeneous soluble complexes with multiple chaperones and similar to 15% in extremely large polymers/aggregates devoid of chaperones. Agents that perturb the synthesis and/or activity of ER chaperones such as tunicamycin and calcium ionophore A23187, have different effects on the solubility and degradation of alpha(1)-AT Z as well as on its residual secretion.