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Secreted and transmembrane mucins inhibit gene transfer with AAV4 more efficiently than AAV5
Robert W Walters1, Joseph M Pilewski, John A Chiorini
1Department of Internal Medicine, University of Iowa College of Medicine, Iowa City, Iowa 52242, USA.
Abstract:
Adeno-associated virus (AAV) is a promising vector for gene transfer in cystic fibrosis. AAV4 and AAV5 both bind to the apical surface of differentiated human airway epithelia, but only AAV5 infects. Both AAV4 and AAV5 require 2,3-linked sialic acid for binding. However, AAV5 interacts with sialic acid on N-linked carbohydrates, whereas AAV4 interacts with sialic acid on O-linked carbohydrates. Because mucin is decorated with O-linked carbohydrates, we hypothesized that mucin binds AAV4 and inhibits gene transfer. To evaluate the effect of secreted mucin, we studied mucin binding and gene transfer to COS cells and the basolateral membrane of well differentiated human airway epithelia. AAV4 bound mucin more efficiently than AAV5, and mucin inhibited gene transfer with AAV4. Moreover, O-glycosidase-pretreated mucin did not block gene transfer with AAV4. Similar to secreted mucin, the transmembrane mucin MUC1 inhibited gene transfer with AAV4 but not AAV5. MUC1 inhibited AAV4 by blocking internalization of the virus. Thus, O-linked carbohydrates of mucin are potent inhibitors of AAV4. Furthermore, whereas mucin plays an important role in innate host defense, its activity is specific; some vectors or pathogens are more resistant to its effects.
Insights
Mucin, a component of airway mucus, inhibits gene transfer using adeno-associated virus serotype 4 (AAV4) by binding to its O-linked carbohydrates. This specificity impacts AAV vector efficacy in cystic fibrosis gene therapy.
Area of Science:
- Gene therapy
- Virology
- Respiratory medicine
Background:
- Adeno-associated virus (AAV) vectors are promising for cystic fibrosis gene therapy.
- AAV4 and AAV5 bind to human airway epithelia via sialic acid, but only AAV5 infects.
- AAV4 binds O-linked carbohydrates, while AAV5 binds N-linked carbohydrates on sialic acid.
Purpose of the Study:
- To investigate the hypothesis that mucin binds AAV4 and inhibits gene transfer.
- To evaluate the effect of secreted mucin and transmembrane mucin MUC1 on AAV4 and AAV5 gene transfer.
Main Methods:
- Studied mucin binding and gene transfer to COS cells and human airway epithelia.
- Utilized O-glycosidase treatment to assess the role of O-linked carbohydrates.
- Compared AAV4 and AAV5 binding and inhibition by mucin and MUC1.
Main Results:
- AAV4 bound mucin more efficiently than AAV5.
- Secreted mucin and MUC1 inhibited AAV4 gene transfer but not AAV5.
- O-glycosidase pretreatment abolished mucin's inhibition of AAV4 gene transfer.
- MUC1 inhibited AAV4 by blocking viral internalization.
Conclusions:
- O-linked carbohydrates on mucin potently inhibit AAV4 gene transfer.
- Mucin's inhibitory activity is specific, affecting AAV4 more than AAV5.
- This specificity has implications for AAV vector selection in cystic fibrosis gene therapy.