Regulation and processing of a precursor form of eosinophil granule major basic protein (ProMBP) in differentiating eosinophils.
The cDNA for eosinophil granule major basic protein (MBP) encodes a prepromolecule with a total length of 222 amino acids (preproMBP). PreproMBP includes a secretory leader of 15 amino acids, an acidic propiece of 90 amino acids, and a basic MBP portion of 117 amino acids. The function of the propiece, which has a predicted pI of 3.9, is unknown, but it gives proMBP an overall acidic charge. Because proMBP is not found in mature eosinophils, we analyzed eosinophil differentiation in interleukin-5 (IL-5)-stimulated umbilical cord stem cells cultured for 24 days. By immunofluorescence, proMBP appeared by day 6 and peaked on day 18, whereas MBP was prominent at days 12 to 24. By day 6, Western blots detected heterogeneous glycosylated 33-kD proMBP; its peak expression occurred on day 12. Western blots showed sequential processing of 33-kD proMBP to an 18-kD intermediate form and finally to 14-kD MBP. By dual label immunoelectron microscopy, proMBP was localized primarily to large uncondensed eosinophil granules, whereas MBP was localized to granules containing a condensed central area. Thus, proMBP is likely expressed and processed as the granule condenses in a multistep process to 14-kD MBP in differentiating eosinophils.
Popken-Harris
P
P
Department of Immunology, Mayo Clinic and Mayo Foundation, Rochester, MN 55905, USA.
Checkel
J
J
Loegering
D
D
Madden
B
B
Springett
M
M
Kephart
G
G
Gleich
G J
GJ
D000595:Amino Acid Sequence; D001798:Blood Proteins / Q000096:biosynthesis* / Q000235:genetics*; D002454:Cell Differentiation; D018076:DNA, Complementary / Q000032:analysis / Q000235:genetics; D047091:Eosinophil Granule Proteins; D004804:Eosinophils / Q000166:cytology / Q000378:metabolism*; D005312:Fetal Blood; D018507:Gene Expression Regulation, Developmental; D006801:Humans; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000096:biosynthesis / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D012260:Ribonucleases
D001798:Blood Proteins; D018076:DNA, Complementary; D047091:Eosinophil Granule Proteins; D011498:Protein Precursors; D011994:Recombinant Proteins; D012260:Ribonucleases