Mutations in the SFTPC gene associated with interstitial lung disease in human patients result in misfolding, endoplasmic reticulum (ER) retention, and degradation of the encoded surfactant protein C (SP-C) proprotein. In this study, genes specifically induced in response to transient expression of two disease-associated mutations were identified by microarray analyses. Immunoglobulin heavy chain binding protein (BiP) and two heat shock protein 40 family members, endoplasmic reticulum-localized DnaJ homologues ERdj4 and ERdj5, were significantly elevated and exhibited prolonged and specific association with the misfolded proprotein; in contrast, ERdj3 interacted with BiP, but it did not associate with either wild-type or mutant SP-C. Misfolded SP-C, ERdj4, and ERdj5 coprecipitated with p97/VCP indicating that the cochaperones remain associated with the misfolded proprotein until it is dislocated to the cytosol. Knockdown of ERdj4 and ERdj5 expression increased ER retention and inhibited degradation of misfolded SP-C, but it had little effect on the wild-type protein. Transient expression of ERdj4 and ERdj5 in X-box binding protein 1 ؊/؊ mouse embryonic fibroblasts substantially restored rapid degradation of mutant SP-C proprotein, whereas transfection of HPD mutants failed to rescue SP-C endoplasmic reticulum-associated protein degradation. ERdj4 and ERdj5 promote turnover of misfolded SP-C and this activity is dependent on their ability to stimulate BiP ATPase activity.

The BiP cochaperone ERdj4 removes misfolded proteins from the ER lumen by associating with ERAD machinery. Global deficiency of ERdj4 results in widespread constitutive ER stress, decreased survival, and metabolic derangements in mice. These findings indicate that the chaperone activity of ERdj4 is important for ER homeostasis in vivo.

Surfactant protein B (SP-B)1 is a critical component of pulmonary surfactant, a lipid-protein mixture which forms a film along the surface of the alveolar epithelium, and is absolutely required for maintenance of alveolar stability at low lung volumes. Surfactant phospholipid mixtures lacking the hydrophobic proteins SP-B and SP-C have poor surface film forming properties, whereas surfactants containing SP-B as the sole protein component rapidly form a stable phospholipid film in vitro and restore lung function in surfactant-deficient preterm animals (1-3). Genetic ablation of the murine SP-B locus leads to acute respiratory distress syndrome at birth resulting in death within minutes (4). Likewise, mutations resulting in SP-B deficiency in human infants lead to severe respiratory distress and death in the neonatal period (5, 6). Intratracheal administration of exogenous SP-B to infants with hereditary SP-B deficiency failed to restore lung function, suggesting that SP-B may have functions in addition to promoting formation of a stable surface film in the alveolus (7). Consistent with this hypothesis, SP-B deficiency in mice and human infants was associated with failure to form lamellar bodies and altered pro-SP-C processing (4, 8).Expression of SP-B is restricted to alveolar Type II cells and nonciliated bronchiolar epithelial (Clara) cells of the pulmonary epithelium (9, 10). In Type II cells, proteolytic processing of the SP-B proprotein is initiated in the multivesicular body with cleavage of an NH 2 -terminal propeptide to generate a processing intermediate of 25 kDa (11,12). Subsequent cleavage of a COOH-terminal propeptide results in liberation of the hydrophobic mature peptide which forms homodimers of 18 kDa. Mature SP-B is stored with surfactant phospholipids in lamellar bodies, the contents of which are released into the alveolar airspace via basal and stimulus-induced secretion (reviewed in (13)).In contrast to the well characterized SP-B biosynthetic pathway in Type II cells, little is known about synthesis and processing of SP-B in Clara cells. Technical problems in the isolation of pure populations of these cells has made study of SP-B processing by Clara cells difficult. Clara cells are the most abundant cell type within the conducting airways of the murine lung, comprising more than 50% of the epithelial cells lining the terminal bronchioles (14,15). Expression of SP-B in cells at the terminal airway/alveolar junction suggests that under certain circumstances, Clara cells may contribute SP-B to the alveolar surfactant pool. In addition to a putative alveolar surfactant function, Clara cell SP-B may also promote formation of a surfactant film in the terminal bronchioles which may be important for maintaining the patency of small conducting airways. Finally, given the widespread distribution of Clara cells in the murine airway, it is possible that SP-B has function(s) that are independent of surfactant activity. In order to better understand the role of SP-B in Type II cells and Clara cells, we ha...