The Role of Pulmonary Collectin N-terminal Domains in Surfactant Structure, Function, and Homeostasis in Vivo *

The N-terminal domains of the lung collectins, surfactant proteins A (SP-A) and D (SP-D), are critical for surfactant phospholipid interactions and surfactant homeostasis, respectively. To further assess the importance of lung collectin N-terminal domains in surfactant structure and function, a chimeric SP-D/SP-A (D/A) gene was constructed by substituting nucleotides encoding amino acids Asn1–Ala7 of rat SP-A with the corresponding N-terminal sequences from rat SP-D, Ala1–Asn25. Recombinant D/A migrated as a 35-kDa band on reducing SDS-PAGE and as a ladder of disulfide-linked multimers under nonreducing conditions. The recombinant D/A bound and aggregated phosphatidylcholine containing vesicles as effectively as rat SP-A. Mice in which endogenous pulmonary collectins were replaced with D/A were developed by human SP-C promoter-driven overexpression of the D/A gene in SP-A−/− and SP-D−/−animals. Analysis of lavage fluid from SP-A−/−,D/A mice revealed that glycosylated, oligomeric D/A was secreted into the air spaces at levels that were comparable with the authentic collectins and that the N-terminal interchange converted SP-A from a “bouquet” to a cruciform configuration. Transmission electron microscopy of surfactant from the SP-A−/−,D/A mice revealed atypical tubular myelin containing central “target-like” electron density. Surfactant isolated from SP-A−/−,D/A mice exhibited elevated surface tension both in the presence and absence of plasma inhibitors, but whole lung compliance of the SP-A−/−,D/Aanimals was not different from the SP-A−/− littermates. Lung-specific overexpression of D/A in the SPD−/− mouse resulted in hetero-oligomer formation with mouse SP-A and did not correct the air space dilation or phospholipidosis that occurs in the absence of SP-D. These studies indicate that the N terminus of SP-D 1) can functionally replace the N terminus of SP-A for lipid aggregation and tubular myelin formation, but not for surface tension lowering properties of SP-A, and 2) is not sufficient to reverse the structural and metabolic pulmonary defects in the SP-D−/−mouse.