We have studied effects of ferric transferrin (FeTF), ferric lactoferrin (FeLF), ferric complexes of pyridoxal- or salicylaldehyde-isonicotinoyl hydrazone, (Fe-PIH, Fe-SIH), and ferric ammonium citrate (FAC) on expression of protein kinase C (PKC) mRNA transcripts in a variety of cultured cell lines. FeTF supported an increase of PKC-β mRNA transcripts in T- lymphoblastoid (CCRF-CEM; Jurkat), B-lymphoblastoid (Daudi; Raji), promyelocyte (HL-60), erythroleukemia (K562), and monocyte (U937) cell lines. By contrast, FeLF, Fe-PIH, and Fe-SIH did not support an increase of PKC-β mRNA transcripts in any of these cell lines. Furthermore, FAC supported an increase of PKC-β mRNA transcripts in HL-60, K562, and U937 cells only. Preincubation of cells with desferrioxamine (DF), a cell-permeable iron chelator, abolished the increments of PKC-β mRNA observed in response to FeTF or FAC. In contrast to results with PKC-β, neither FeTF nor FAC caused an increase of PKC-α transcripts in any cell line. To locate iron- responsive DNA regulatory elements of the PKC-β gene, we prepared genetic constructs containing various portions of the human PKC-β 5'-flanking DNA linked to the firefly luciferase gene. Constructs were cotransfected with the neomycin resistance plasmid, Pwl-neo, into HRE H9 cells, and stable transfectants were selected in G418. Treatment with FeTF of transfectants bearing chimeric gene constructs with 2,200 bp of the PKC-β 5'-flanking region increased luciferase activity and mRNA transcripts 2.5-fold. This increase was blocked by DF. Neither luciferase activity nor mRNA increased with FeTF in stable transfectants bearing constructs with 342 bp or 587 bp of the PKC-β 5'-flanking region. These data provide direct confirmation that iron is involved in regulation of PKC-β but not PKC-α gene expression in many cell lines. The form in which iron is presented to these cell lines appears to affect its availability for this function, and cells vary in their capabilities to use nontransferrin iron to support PKC-β gene expression. Finally, transcriptional upregulation of PKC-β by FeTF is mediated by DNA sequences located between -2200 bp and -587 bp in the 5'-flanking region of the human PKC-β gene.
|Original language||English (US)|
|Number of pages||8|
|State||Published - Jan 1 1994|
ASJC Scopus subject areas
- Cell Biology