Inherited pathogenic variants of surfactant protein C (SFTPC) can cause familial pulmonary fibrosis. The most common, SFTPC-I73T, aberrantly localises to the plasma membrane and causes alveolar type 2 (AT2) dysfunction via a poorly understood toxic gain-of-function. Understanding the mechanisms of SFTPC trafficking and their interactors in health and disease is crucial to defining how pathogenic variants induce cellular dysfunction.

To investigate trafficking components required for SFTPC trafficking that are perturbed by the I73T variant, I undertook SFTPC-WT and -I73T proximity labelling using TurboID and identified labelled proteins by mass spectrometry. I discovered >100 proteins preferentially labelled by SFTPC-WT and therefore likely to be required for normal SFTPC trafficking. I undertook validation of biologically relevant hits by CRISPR/Cas9 deletion of candidates involved in membrane trafficking, ubiquitination and lysosomal acidification. I found that many induced misprocessing and relocalisation of SFTPC-WT to the plasma membrane, phenocopying the I73T mutation. I undertook further validation of top hits in a novel CRISPR interference AT2 organoid model.

I also undertook an exploration of the biology of MPZL1, a poorly characterised Ig-superfamily member which differentially interacts with, and is relocalised by, SFTPC-WT and I73T. This suggests that the normal function of MPZL1 may require interaction with SFTPC-WT. I defined novel aspects of its expression, cleavage and activation and showed from RNA sequencing that MPZL1 activation upregulates genes involved in cell migration; this may be crucial for AT2 function in alveolar repair.

Together, this work has identified novel interactors involved in SFTPC trafficking and explored the biology of MPZL1, offering further insights into how SFTPC-I73T contributes to the development of pulmonary fibrosis.