IDENTIFICATION OF THE PROTEIN-KINASE-C ISOENZYMES IN HUMAN LUNG AND AIRWAYS SMOOTH-MUSCLE AT THE PROTEIN AND MESSENGER-RNA LEVEL

The protein kinase C (PKC) isoenzymes expressed by human peripheral lu ng and tracheal smooth muscle resected from individuals undergoing hea rt-lung transplantation were identified at the protein and mRNA level. Western immunoblot analyses of human lung identified multiple PKC iso enzymes that were differentially distributed between the soluble and p articulate fraction. Thus, PKC alpha, PKC beta(II), PKC epsilon, and P KC zeta were recovered predominantly in the soluble fraction whereas t he eta isoform was membrane-associated together with trace amounts of PKC alpha and PKC epsilon. PKC beta(I)-like immunoreactivity was occas ionally seen although the intensity of the band was uniformly weak. Im munoreactive bands corresponding to PKCs gamma, delta, or a 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 fr agments that corresponded to the predicted sizes of PKC alpha, PKC bet a(I), PKC beta(II), PKC epsilon, PKC zeta, and PKC eta (consistent wit h the expression of PKC isoenzyme protein) and, in addition, mRNA for PKC delta; PCR fragments of the expected size for the supposedly muscl e-specific isoform, PKC theta, or the atypical isoenzyme, PKC lambda, were never obtained. The complement and distribution of PKC isoforms i n human trachealis were similar, but not identical, to human lung. Thu s, immunoreactive bands corresponding to the alpha, beta(I), beta(II), epsilon, and zeta isoenzymes of PKC were routinely labelled in the cy tosolic fraction. In the particulate material PKC alpha, PKC epsilon, PKC alpha, PKC eta, and PKC mu were detected by immunoblotting. With t he exception of PKC zeta, RT-PCR analyses confirmed the expression of the PKC isoforms detected at the protein level and, in addition, ident ified 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 r oles in these tissues. (C) 1997 Elsevier Science Inc.