HYPOXIA INDUCES THE SYNTHESIS OF TROPOMYOSIN IN CULTURED PORCINE PULMONARY-ARTERY ENDOTHELIAL-CELLS

The present study examined the effect of hypoxia on protein synthesis by porcine pulmonary artery endothelial cells (PAEC). Hypoxia decrease d protein synthesis in PAEC, but two-dimensional gel electrophoresis o f [S-35] methionine-labeled PAEC proteins demonstrated the increased s ynthesis of a set of proteins having molecular masses (M(r)) of 35, 36 .5, 45, 116, and 205 kDa. The synthesis of the 35-, 36.5-, and 45-kDa proteins was increased in preconfluent and postconfluent cells. The 35 - and 45-kDa proteins were not induced by hyperthermia, whereas the 36 .5-kDa protein was induced slightly by hyperthermia. Induction of the 36.5- and 45-kDa proteins required a minimum of 8 h of hypoxia, wherea s induction of the 35-kDa protein required only 4 h of exposure to hyp oxia. The upregulated synthesis of the 35-, 36.5-, and 45-kDa proteins was reversible with return to normoxia. Actinomycin D, an inhibitor o f transcription, did not block the hypoxic induction of the 35- and 36 .5-kDa proteins but did block induction of the 45-kDa protein. The par tial amino acid sequence of the 35-kDa protein obtained from cyanogen bromide cleavage of the molecule was e-Lys-Lys-Lys-Met-Gln-Met-Leu-Lys -Leu-Asp-Lys-Glu. This partial sequence of the 35-kDa protein identica lly matches the sequence of tropomyosin. Amino acid composition data a nd the isoelectric point (4.8) were also typical of tropomyosin. Final ly, specific cross-reactivity was detected between the 35-kDa protein and a monoclonal antibody to chicken gizzard tropomyosin on immunoblot . Thus hypoxia induces the synthesis of tropomyosin, a major microfila ment-associated protein, in porcine PAEC in monolayer culture.