Detection of a kallikrein in the mouse lactating mammary gland: a possible processing enzyme for the epidermal growth factor precursor.
Kallikreins are a multigene subfamily of serine proteases that may have a role in processing precursors of polypeptide hormones and growth factors. The epidermal growth factor (EGF) immunoreactivity in mouse milk is derived from the membrane-bound EGF precursor located on the lumenal border of the alveolar cells in the mammary gland. Release of EGF into the milk requires the hydrolysis of the EGF precursor at Arg-X cleavage sites. We report the presence of a candidate EGF precursor-processing enzyme in the lactating mouse mammary gland. Kallikrein transcripts in the mouse lactating mammary gland were detected by primer-directed enzyme amplification of complementary DNA (cDNA). Primers to selected conserved regions of the kallikrein cDNA resulted in an amplified product of the predicted size (573 basepairs). Sequence analysis of the product over three nonconserved regions identified mGK-6 (mouse renal kallikrein) as the primary kallikrein in BALB/c mouse lactating mammary gland. Transcription products for the EGF-binding protein (mGK-9), mGK-1, MGK-3, and mGK-4 were not detected by enzyme amplification with specific primers corresponding to these kallikrein cDNAs. Positive immunohistochemical staining of the apical membrane of mammary alveolar cells was detected with a polyclonal antiserum to mouse kallikrein. Incubation of cell membranes isolated from lactating mammary glands released soluble EGF-immunoreactive material. Aprotinin partially inhibited the release of this material, whereas other protease inhibitors, such as leupeptin, benzamidine, and limabean trypsin inhibitor, had no detectable effect. These results support the hypothesis that the release of EGF-immunoreactive material into the milk is in part dependent upon a kallikrein enzyme (mGK-6) in the BALB/c mouse lactating mammary gland.
Jahnke
G D
GD
Laboratory of Biochemical Risk Analysis, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709.
Chao
J
J
Walker
M P
MP
Diaugustine
R P
RP
D000818:Animals; D007611:Aprotinin / Q000494:pharmacology; D001483:Base Sequence; D018076:DNA, Complementary / Q000032:analysis / Q000235:genetics; D004815:Epidermal Growth Factor / Q000032:analysis / Q000378:metabolism*; D005260:Female; D007150:Immunohistochemistry; D007610:Kallikreins / Q000032:analysis*; D007774:Lactation / Q000502:physiology*; D008321:Mammary Glands, Animal / Q000737:chemistry* / Q000201:enzymology / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008892:Milk / Q000737:chemistry; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000032:analysis / Q000378:metabolism*; D011863:Radioimmunoassay; D014158:Transcription, Genetic
D018076:DNA, Complementary; D011498:Protein Precursors; D004815:Epidermal Growth Factor; D007611:Aprotinin; D007610:Kallikreins