Inhibition of protein kinase C promotes differentiation of neuroblastoma × glioma NG108-15 hybrid cells

Abstract

Differentiation of neuroblastoma × glioma NG108-15 hybrid cells can be induced by different means, but the mechanisms involved are unclear. Our aim was to characterize the role of protein kinase C (PKC) in this process. The PKCs present in NG108-15 cells, i.e. PKCα, PKCδ, PKCε and PKCζ, were inhibited using a cocktail of Go6983 and Ro318220 or were downregulated by treatment with phorbol 12-myristate 13-acetate (PMA). In high-glucose Dulbecco's modified Eagle medium, neuritogenesis was induced by 24 h treatment with a cocktail of Go6983 and Ro318220 or by 48 h treatment with PMA, the latter process thus requiring a longer treatment. However, when cells treated with PMA for only 24 h were placed in extracellular standard salts solution, e.g. Locke's buffer, for 3 h, morphological and functional differentiation occurred, with rounding of the cell body, actin polymerization subjacent to the plasma membrane and an increase in voltage-sensitive Ca(2+) channel activity in the absence of cell death. This rapid differentiation was not due to autophagy, growth arrest or increased cyclic AMP response element binding protein phosphorylation, but coincided with combined activation of p38 mitogen-activated protein kinase (MAPK) and inhibition of extracellular signal-regulated kinase (ERK) and Akt, as confirmed by the effects of selective inhibitors. Furthermore, PKC activation blocked thapsigargin-induced neuritogenesis, whereas PKC downregulation did not. These results show that PKC downregulation promotes differentiation and this effect is accelerated by exposure to Locke's buffer. Although this experimental paradigm cannot be related to the in vivo situation and disease, it implies that combined inhibition of Akt and p44/p42 ERK and activation of p38 MAPK promotes differentiation.

Impairment of adenylyl cyclase 2 function and expression in hypoxanthine phosphoribosyltransferase-deficient rat B103 neuroblastoma cells as model for Lesch–Nyhan disease: BODIPY–forskolin as pharmacological tool

“…AC6 was also up-regulated in HPRT − cells, although to a lesser extent than AC5. For the other AC isoforms, minor changes in expression were observed, and these changes were not consistently found in Effects on FS, BODIPY-FS, and BODIPY control dye on morphology of B103 control and HPRT − cells It is well known that cAMP-increasing substances induce differentiation in neuronal cells (Braumann et al 1986;Monaghan et al 2008;Chang et al 2011). Differentiation is associated with cell rounding and clustering and neurite extension (Schubert et al 1974;Ishii et al 1990;Jalink and Moolenaar 1992;Hirose et al 1998;Ishii and Chun 2002).…”

PI3K/Akt signaling pathway triggers P2X7 receptor expression as a pro-survival factor of neuroblastoma cells under limiting growth conditions

“…S3A online), suggesting that P2rx7 gene expression is controlled by aPKCs. Atypical isotypes (PKCι/λ and PKCζ) have been reported to be critically involved in cell proliferation and survival, although showing different distribution patterns: while PKCι/λ is restricted to testis, insulin secreting cells and epidermis; PKCζ is widely expressed in all tissues, including neuroblastoma 32 33 . In N2a cells, serum deprivation resulted in rapid and transient phosphorylation of PKCζ activation loop (Thr 410 ), reaching top levels 15 min after serum starvation (see supplementary Fig.…”

Global signaling effects of a schizophrenia-associated missense mutation in neuregulin 1: an exploratory study using whole genome and novel kinome approaches

“…PKA has been implicated in neurite growth and has been shown to interact with the MAPK pathway in regulating neuronal differentiation (Vogt Weisenhorn et al 2001; Yao et al 1998). ERK is a crucial regulator of neurite growth (Sarina et al 2013; Auer et al 2012) and is involved in maintaining balance between gliogenesis and neurogenesis during development (Chang et al 2011). Protein Kinase C mediates activation of ERK-regulated dendritic spine density (Goldin, Segal 2003).…”