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Efficient Production and Purification of Recombinant Murine Kindlin-3 from Insect Cells for Biophysical Studies
Published on: March 19, 2014
Interaction of kindlin-3 and β2-integrins differentially regulates neutrophil recruitment and NET release in mice
Zhen Xu1, Jiayi Cai2, Juan Gao3
1Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, WI; Collaborative Research Program for Cell Adhesion Molecules, Shanghai University School of Life Sciences, Shanghai, China;
Abstract:
Kindlin-3 essentially supports integrin activation in blood cells. Absence of kindlin-3 in humans causes leukocyte adhesion deficiency-III characterized with severe bleeding disorder and recurrent infections. Previously, we generated kindlin-3 knock-in (K3KI) mice carrying an integrin-interaction disrupting mutation in kindlin-3 and verified the functional significance of the binding of kindlin-3 to integrin αIIbβ3 in platelets. Here, using K3KI mice, we functionally evaluate the crosstalk between kindlin-3 and β2-integrins in neutrophils. Although the kindlin-3 mutant in K3KI neutrophils is normally expressed, its binding ability to β2-integrins in neutrophils is disabled. In vitro and in vivo analyses disclose that β2-integrin-mediated K3KI neutrophil adhesion and recruitment are significantly suppressed. Interestingly, the ability of releasing neutrophil extracellular traps (NETs) from K3KI neutrophils is also compromised. Substantially, a peptide derived from the integrin β2 cytoplasmic tail that can inhibit the interaction between kindlin-3 and β2-inegrins significantly jeopardizes NET release without affecting neutrophil adhesion and recruitment under the experimental conditions. These findings suggest that crosstalk between kindlin-3 and β2-integrins in neutrophils is required for supporting both neutrophil recruitment and NET release, but the involved regulatory mechanisms in these two cellular events might be differential, thus providing a novel therapeutic concept to treat innate immune-related diseases.
Insights
Kindlin-3 is crucial for neutrophil function. Disrupting its interaction with β2-integrins impairs neutrophil adhesion, recruitment, and NET release, offering new therapeutic avenues for immune diseases.
Area of Science:
- Immunology
- Cell Biology
- Hematology
Background:
- Kindlin-3 deficiency causes leukocyte adhesion deficiency-III, a severe bleeding and infection disorder.
- Kindlin-3 plays a vital role in integrin activation in blood cells, particularly in platelets.
- The interaction between kindlin-3 and integrins is essential for proper immune cell function.
Purpose of the Study:
- To investigate the functional crosstalk between kindlin-3 and β2-integrins in neutrophils.
- To evaluate the impact of disrupted kindlin-3 binding on neutrophil adhesion, recruitment, and NET release.
- To explore potential therapeutic strategies for innate immune-related diseases based on this interaction.
Main Methods:
- Utilized kindlin-3 knock-in (K3KI) mice with a mutation disrupting kindlin-3 and integrin binding.
- Performed in vitro and in vivo analyses of K3KI neutrophil adhesion and recruitment.
- Assessed neutrophil extracellular trap (NET) release in K3KI neutrophils and using a specific inhibitory peptide.
Main Results:
- K3KI neutrophils exhibited significantly suppressed β2-integrin-mediated adhesion and recruitment.
- The ability of K3KI neutrophils to release NETs was compromised.
- An inhibitory peptide targeting the kindlin-3/β2-integrin interaction impaired NET release but not adhesion/recruitment.
Conclusions:
- Crosstalk between kindlin-3 and β2-integrins is essential for both neutrophil recruitment and NET release.
- Distinct regulatory mechanisms may govern kindlin-3's role in neutrophil adhesion versus NET release.
- This interaction presents a novel therapeutic target for innate immune-related diseases.
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