Figure 1. Cellular and histopathological phenotype of the SP-C^I73T-Neo founder line.

(A) Schematic of the HA-tagged Sftpc^I73T knockin allele showing the PGK-Neo cassette in intron 1. (B) qRT-PCR analysis for Sftpc mRNA expression in 12- to 28-week-old WT and homozygous SP-C^{I73T-Neo/I73T-Neo} (SP-C I73T) mice. (C) Western blot of AT2 lysates for proSP-C (20 μg protein/lane). SP-C^{I73T-Neo/I73T-Neo} mice accumulate an HA-tagged primary translation product (arrowheads) and misprocessed isoforms (arrows, right bracket). In WT/WT mice, both the primary translation product (arrowheads) and major processing intermediate (left bracket) were detected. (D) Immunohistochemistry for proSP-C of lung sections from 8-week-old WT and SP-C^{I73T-Neo/I73T-Neo} mice revealed proSP-C^I73T expression on AT2 cell plasma membranes (arrowheads); proSP-C^WT is expressed in cytosolic vesicles of AT2 cells (arrows). (E) proSP-C Western blot of subcellular fractions enriched in ER or lamellar bodies (LB) from 8-week-old SP-C^WT/WT and SP-C^{I73T-Neo/I73T-Neo} mice. ER from each line contained the corresponding proSP-C primary translation product (arrowheads). The major proSP-C^WT intermediate (M_r, 6,000) was enriched in LB (left bracket). All aberrantly processed proSP-C^I73T isoforms were excluded from SP-C^I73T-Neo LB (arrows, right bracket). (F) Western blot of surfactant showing decreased mature SP-C in SP-C^{I73T-Neo/I73T-Neo} mice. (G) H&E-stained sections from 16-week-old mice (scale bars: 2 mm (upper panels); 200 μm (lower panels). Quantitative morphometry using ImageJ expressed as airspace/tissue ratio. (H) Total soluble collagen content in the left lobe from 32-week-old mice. Data represent mean ± SEM with dot plot overlays. *P < 0.05 versus SP-C^WT by 2-tailed t test.