The neutrophil glycoprotein Mo1 is an integral membrane protein of plasma membranes and specific granules
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K B Stevenson1, W M Nauseef, R A Clark
1Department of Medicine, Veterans Administration Medical Center, Iowa City, IA.
Abstract:
The glycoprotein Mo1 has previously been demonstrated to be on the cell surface and in the specific granule fraction of neutrophils and to be translocated to the cell surface during degranulation. It is not known, however, whether Mo1 is an integral membrane protein or a soluble, intragranular constituent loosely associated with the specific granule membrane. Purified neutrophils were disrupted by nitrogen cavitation and separated on Percoll density gradients into four fractions enriched for azurophilic granules, specific granules, plasma membrane, and cytosol, respectively. The glycoproteins in these fractions were labeled with 3H-borohydride reduction, extracted with Triton X-114, and immunoprecipitated with 60.3, an anti-Mo1 monoclonal antibody Mo1 was detected only in the specific granule and plasma membrane fractions and partitioned exclusively into the detergent-rich fraction consistent with Mo1 being an integral membrane protein. In addition, treatment of specific granule membranes with a high salt, high urea buffer to remove absorbed or peripheral proteins failed to dissociate Mo1. These data support the hypothesis that Mo1 is an integral membrane protein of plasma and specific granule membranes in human neutrophils.
Insights
The glycoprotein Mo1 is an integral membrane protein in human neutrophils. It is found in specific granules and plasma membranes, not loosely associated, confirming its structural role.
Area of Science:
- Immunology
- Cell Biology
- Biochemistry
Background:
- The glycoprotein Mo1 is present on neutrophil cell surfaces and within specific granules.
- Its precise localization (integral membrane vs. soluble) and association with granule membranes remain unclear.
Purpose of the Study:
- To determine if Mo1 is an integral membrane protein or a soluble component of neutrophil specific granules.
Main Methods:
- Neutrophils were fractionated using nitrogen cavitation and Percoll density gradients.
- Glycoproteins were labeled, extracted with Triton X-114, and immunoprecipitated using an anti-Mo1 antibody.
- Specific granule membranes were treated with high salt/urea buffers to assess protein association.
Main Results:
- Mo1 was detected exclusively in specific granule and plasma membrane fractions.
- Mo1 partitioned into the detergent-rich phase after Triton X-114 extraction, indicating membrane integration.
- Mo1 remained associated with specific granule membranes after treatments designed to remove peripheral proteins.
Conclusions:
- Mo1 functions as an integral membrane protein within both plasma and specific granule membranes of human neutrophils.
- These findings support Mo1's structural role in neutrophil membrane integrity and function.