On the Lysosomal Degradation of Neurofibromin and Its Phosphorylation in Cultured Melanocytes

Abstract

Neurofibromatosis type 1 (NF1) is one of the most common inherited disorders in humans. Most of the NF1 gene mutations result in a reduction of the amount of neurofibromin to about 50%. Recently, we found that the level of neurofibromin can be regulated post-translationally through the alteration of its half-life. Here, we investigated whether lysosomes are involved in this post-translational regulation in cultured melanocytes of NF1 patients and controls. When the lysosomal degradation was inhibited by chloroquine, an increase of neurofibromin by a factor of 2 to 3, correlating with an increased half-life, was measured. Incubation with phosphoprotein-phosphatase inhibitors also increased the neurofibromin content in melanocytes. Investigations on phosphorylation of neurofibromin revealed a basal phosphorylation in melanocytes cultured with growth factor-deprived medium that increased upon incubation with the growth stimulators PMA or bFGF. Because both factors are also able to increase the half-life of neurofibromin, we suggest its phosphorylation to be an important step in protecting neurofibromin against specific lysosomal degradation.

Differential Regulation of RasGAPs in Cancer

“…77 It is unknown if these findings are linked to earlier studies reporting that NF1 phosphorylation inhibits its lysosomal degradation. 78 Interestingly, ETEA/UBXD8 does not interact with p120GAP, 77 the other ubiquitously expressed RasGAP, indicative of this interaction contributing to differential and spatiotemporal Ras signaling.…”

Pigment cell‐related manifestations in neurofibromatosis type 1: an overview

“…Here, stimulation with PMA resulted in less increase in neurofibromin half‐life compared with NF1 and healthy donor normal skin melanocytes (Kaufmann et al., 1999a). Neurofibromin, which is susceptible to lysosomal degradation through the presence of a special amino acid motif in its GRD, undergoes phosphorylation under PMA or bFGF stimulated culture conditions, thereby protecting it against lysosomal degradation (Kaufmann et al., 1999b). The possible role of neurofibromin in melanocyte differentiation was further elaborated by the transient co‐expression in a melanoma cell line of rat neurofibromin cDNA and a fusion gene, consisting of the tyrosinase promotor region cloned upstream of the firefly luciferase reporter gene.…”

Interplay Between Protein Kinase C Isoforms Alpha and Epsilon, Neurofibromin, and the Ras/MAPK Pathway in Neuroblastoma Differentiation

“…Whether neurofibromin phosphorylation on Ser2808 (or other sites) is directly involved is, however, unknown. In sharp contrast, phosphorylation of neurofibromin in-non tumor cellular contexts has been correlated with increased stability of the protein, at least in melanocytes (Kaufmann et al, 1999) and neurons (Mangoura et al, 2006a). In conclusion, PKCs have the potential to directly impact on the activation state of Ras by modulating the activity of both GEFs and GAPs in a variety of ways.…”