Quantitation of tryptase, chymase, Fc epsilon RI alpha, and Fc epsilon RI gamma mRNAs in human mast cells and basophils by competitive reverse transcription-polymerase chain reaction.
Competitive reverse transcription-PCR assays developed for human tryptase, chymase, Fc epsilon RI alpha, and Fc epsilon RI gamma mRNA molecules were applied to the HMC-1 leukemic mast cell line, the KU812 leukemic basophil cell line, mast cells dispersed from lung and skin, and peripheral blood basophils. Relative amounts of alpha-tryptase and beta-tryptase mRNA were determined by analysis of BseAI digests of PCR products. Tryptase expression was highest in tissue-derived mast cells, lowest in basophils and KU812 cells, and intermediate in HMC-1 cells. beta-Tryptase mRNA predominated in HMC-1 and KU812 cells; mixtures of alpha- and beta-tryptase were found in tissue mast cells; and alpha-tryptase predominated in basophils. Chymase mRNA was more abundant in skin-derived (nearly all of the MCTC type) than lung-derived (variable amounts of MCTC and MCT cells) mast cells. Small amounts of chymase mRNA were detected in HMC-1 cells; none was found in basophils, in KU812 cells, or in the one preparation of 100% MCT cells derived from lung. Comparable amounts of Fc epsilon RI alpha and Fc epsilon RI gamma mRNA molecules were measured in basophils and tissue-derived mast cells, lesser amounts were detected in KU812 cells, and almost none was detected in HMC-1 cells. Thus, steady state levels of the granule and membrane resident molecules examined in our study are transcriptionally regulated in mast cells and basophils.
Xia
H Z
HZ
Department of Medicine, Virginia Commonwealth University, Richmond 23298, USA.
Kepley
C L
CL
Sakai
K
K
Chelliah
J
J
Irani
A M
AM
Schwartz
L B
LB
D001483:Base Sequence; D001491:Basophils / Q000737:chemistry* / Q000201:enzymology / Q000276:immunology; D001667:Binding, Competitive; D002478:Cells, Cultured; D053818:Chymases; D018076:DNA, Complementary / Q000032:analysis; D006801:Humans; D008407:Mast Cells / Q000737:chemistry* / Q000201:enzymology / Q000276:immunology; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D018075:RNA, Complementary / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis; D017455:Receptors, IgE / Q000032:analysis*; D012697:Serine Endopeptidases / Q000032:analysis*; D053802:Tryptases
D018076:DNA, Complementary; D018075:RNA, Complementary; D012333:RNA, Messenger; D017455:Receptors, IgE; D012697:Serine Endopeptidases; C083294:chymase 2; D053818:Chymases; D053802:Tryptases