Immediate Early Gene Expression in PC12 Cells Exposed to Lead: Requirement for Protein Kinase C

Abstract

Abstract:We previously demonstrated induction of c-fos mRNA in PC12 cells exposed to lead that was dependent on new transcription. In the current work, we examined two signal transduction mechanisms that are activated by lead and have been shown to mediate induction of c-fos mRNA. One mechanism involves protein kinase C, and the other requires calmodulin-dependent protein kinase II. Significant increases in the levels of c-fos, c-jun, and egr-1 but not NGFIB mRNA were observed in PC12 cells exposed to lead or phorbol 12-myristate 13-acetate. In contrast, PC12 cells depolarized with 56 mM K ϩ displayed an increase in c-fos, egr-1, and NGFIB but not c-jun mRNA. Similar to other activators of protein kinase C, lead increased AP-1 and Egr-1 DNA binding activity. Additionally, lead increased luciferase activity in cerebellar granule cells transfected with an AP-1 luciferase reporter construct. Lead did not increase c-fos mRNA in PC12 cells that were depleted of protein kinase C by a 24-h treatment with phorbol 12,13-dibutyrate or incubated with the protein kinase C inhibitor H-7. In contrast, an inhibitor of calmodulin-dependent protein kinase, KN-62, and an inhibitor of calmodulin, W-7, did not block the induction of c-fos mRNA by lead. An increase in serumresponse element DNA-binding activity was observed in nuclear extracts from PC12 cells exposed to lead. It is interesting that lead activated protein kinase C isoforms ␦ and ⑀, but not isoforms ␣ and ␤. In conclusion, lead appears to induce the expression of immediate early genes by a mechanism that requires protein kinase C.

Induction of Vascular Endothelial Growth Factor in Human Astrocytes by Lead

“…Similarly, PKC-ε has been found to play a role in the induction of immediate early gene expression in the rat PC12 cells (47). The potential involvement of other PKC isoforms in response of human astrocytes to lead is currently under investigation.…”

“…The PKC family comprises more than ten isoforms categorized into three structurally related groups: conventional PKCs (cPKCs: α,β and γ) are regulated by calcium and diacylglycerol or phorbol ester; novel PKCs (nPKCs: δ,θ,η and ε) are sensitive to diacylglycerol and phorbol ester but are calcium-independent; and atypical PKCs (aPKCs: ζ, µ and ι) are insensitive to calcium, diacylglycerol and phorbol ester (45). Activation of both calciumdependent and calcium-independent PKC isoforms by lead has been described in rodent and bovine cells such as glial, endothelial and PC12 pheochromocytoma cell lines (12,46,47). It has been proposed that lead alters PKC function by interacting with a high affinity site within the Nterminal regulatory domain either at the calcium activation site or cysteine-rich zinc-finger-like binding site (11,15,48).…”

Oral Lead Exposure Induces Dysbacteriosis in Rats

“…Previous studies have suggested that lead toxicity is the result of its effect on calcium metabolism 17) , and channel dysfunction 29) . Nevertheless, some mechanisms underlying lead toxicity at low concentrations are still unclear, maybe because of the absence of a systemic approach to understanding lead toxicity.…”

Early lead exposure increases the leakage of the blood-cerebrospinal fluid barrier, in vitro

“…Possible mechanisms include the activation of protein kinase C (PKC) by Pb, which subsequently increases the transcription of immediate early response genes (c-fos and c-jun), leading to an overexpression of vascular endothelial growth factor (VEGF) and dysfunction of the BBB. 38,39,45,46 Activation of PKC by Pb has also been demonstrated in the choroidal epithelial cells, 47 although the relationship between PKC activation and tightness of the BCB has not been established. Our data clearly demonstrate a selective inhibition by Pb of the expression of claudin-1 in choroidal epithelial cells, as supported by both RT-PCR on claudin-1 mRNA expression and Western blot on the protein level.…”