Identification of the protein kinase C isoenzymes in human lung and airways smooth muscle at the protein and mRNA level.
The protein kinase C (PKC) isoenzymes expressed by human peripheral lung and tracheal smooth muscle resected from individuals undergoing heart-lung transplantation were identified at the protein and mRNA level. Western immunoblot analyses of human lung identified multiple PKC isoenzymes that were differentially distributed between the soluble and particulate fraction. Thus, PKC alpha, PKC betaII, PKC epsilon, and PKC zeta were recovered predominantly in the soluble fraction whereas the eta isoform was membrane-associated together with trace amounts of PKC alpha and PKC epsilon. PKC beta1-like immunoreactivity was occasionally seen although the intensity of the band was uniformly weak. Immunoreactive bands corresponding to PKCs gamma, delta, or theta were never detected. Reverse transcription-polymerase chain reaction (RT-PCR) of RNA extracted from human lung using oligonucleotide primer pairs that recognise unique sequences in each of the PKC genes amplified cDNA fragments that corresponded to the predicted sizes of PKC alpha, PKC betaI, PKC betaII, PKC epsilon, PKC zeta, and PKC eta (consistent with the expression of PKC isoenzyme protein) and, in addition, mRNA for PKC delta; PCR fragments of the expected size for the supposedly muscle-specific isoform, PKC theta, or the atypical isoenzyme, PKC lambda, were never obtained. The complement and distribution of PKC isoforms in human trachealis were similar, but not identical, to human lung. Thus, immunoreactive bands corresponding to the alpha, betaI, betaII, epsilon, and zeta isoenzymes of PKC were routinely labelled in the cytosolic fraction. In the particulate material PKC alpha, PKC epsilon, PKC alpha, PKC eta, and PKC mu were detected by immunoblotting. With the exception of PKC zeta, RT-PCR analyses confirmed the expression of the PKC isoforms detected at the protein level and, in addition, identified mRNA for PKC delta. Collectively, these data clearly demonstrate the expression of multiple PKC isoenzymes in human lung and tracheal smooth muscle, suggesting that they subserve diverse multifunctional roles in these tissues.
Webb
B L
BL
Thoracic Medicine, Imperial College School of Medicine at the National Heart and Lung Institute, London, UK.
Lindsay
M A
MA
Seybold
J
J
Brand
N J
NJ
Yacoub
M H
MH
Haddad
E B
EB
Barnes
P J
PJ
Adcock
I M
IM
Giembycz
M A
MA
D000328:Adult; D015153:Blotting, Western; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D008168:Lung / Q000201:enzymology*; D009130:Muscle, Smooth / Q000201:enzymology*; D016133:Polymerase Chain Reaction; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D014132:Trachea / Q000201:enzymology*
D007527:Isoenzymes; D012333:RNA, Messenger; D011493:Protein Kinase C