Intracellular precursor interleukin (IL)-1alpha, but not mature IL-1alpha, is able to regulate human endothelial cell...

The human umbilical vein endothelial cell (HUVEC) has a finite lifespan in vitro, and senescent HUVEC contain elevated levels of the negative growth regulator interleukin (IL)-1alpha. IL-1alpha is translated as a signal peptide sequence-less cytosolic 31-kDa precursor (IL-1alpha p), which undergoes proteolytic activation to release the mature carboxyl terminus 17-kDa protein (IL-1alpha m). Both the IL-1alpha p and IL-1alpha m proteins are biologically active as exogenous cytokines. Interestingly, only IL-1alpha p contains a nuclear localization sequence between residues 79 and 85. To further study the role of intracellular IL-1alpha in the regulation of human endothelial cell function, a spontaneous HUVEC transformant was stably transfected with IL-1alpha p, IL-1alpha m, and the IL-1alpha p K82N mutant, which attenuates the nuclear traffic of IL-1alpha p. Interestingly, the IL-1alpha p transfectants were found to have a lower migratory potential than either IL-1alpha m or IL-1alpha p K82N transfectants, and the addition of the IL-1 receptor antagonist did not alter the migration of these cells. Immunofluorescence microscopy demonstrated that only the IL-1alpha p transfectants exhibited prominent staining for beta-catenin-associated cell-to-cell contacts, as well as pronounced vimentin intermediate filaments and actin cytoskeleton staining. These data suggest that IL-1alpha p, and not IL-1alpha m, may function as an intracellular regulator of the migratory capacity of the human endothelial cell and that the nuclear localization sequence present within IL-1alpha p may be involved in regulating this function.