Human surfactant protein A2 gene mutations impair dimmer/trimer assembly leading to deficiency in protein sialylation and secretion

PLoS One. 2012;7(10):e46559. doi: 10.1371/journal.pone.0046559. Epub 2012 Oct 3.

Abstract

Surfactant protein A2 (SP-A2) plays an essential role in surfactant metabolism and lung host defense. SP-A2 mutations in the carbohydrate recognition domain have been related to familial pulmonary fibrosis and can lead to a recombinant protein secretion deficiency in vitro. In this study, we explored the molecular mechanism of protein secretion deficiency and the subsequent biological effects in CHO-K1 cells expressing both wild-type and several different mutant forms of SP-A2. We demonstrate that the SP-A2 G231V and F198S mutants impair the formation of dimmer/trimer SP-A2 which contributes to the protein secretion defect. A deficiency in sialylation, but not N-linked glycosylation, is critical to the observed dimmer/trimer impairment-induced secretion defect. Furthermore, both mutant forms accumulate in the ER and form NP-40-insoluble aggregates. In addition, the soluble mutant SP-A2 could be partially degraded through the proteasome pathway but not the lysosome or autophagy pathway. Intriguingly, 4-phenylbutyrate acid (4-PBA), a chemical chaperone, alleviates aggregate formation and partially rescued the protein secretion of SP-A2 mutants. In conclusion, SP-A2 G231V and F198S mutants impair the dimmer/trimer assembly, which contributes to the protein sialylation and secretion deficiency. The intracellular protein mutants could be partially degraded through the proteasome pathway and also formed aggregates. The treatment of the cells with 4-PBA resulted in reduced aggregation and rescued the secretion of mutant SP-A2.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Biopolymers / chemistry
  • Biopolymers / genetics*
  • CHO Cells
  • Cricetinae
  • Cricetulus
  • DNA Primers
  • Glycosylation
  • Humans
  • Microscopy, Fluorescence
  • Mutation*
  • N-Acetylneuraminic Acid / chemistry*
  • Polymerase Chain Reaction
  • Pulmonary Surfactant-Associated Protein A / chemistry
  • Pulmonary Surfactant-Associated Protein A / genetics*

Substances

  • Biopolymers
  • DNA Primers
  • Pulmonary Surfactant-Associated Protein A
  • N-Acetylneuraminic Acid

Grants and funding

This work was supported by the Key Project of Chinese Ministry of Education (210159), the Introduction of Talents Program of Institutions of Higher Education of Guangdong Province (http://www.gdhed.edu.cn), “973” National S&T Major Project (2010CB912504, 2011CB503900) and the National Natural Science Foundation of China(30900595, 81170101). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.