Cell specific transformation by c-fms activating loop mutations is attributable to constitutive receptor degradation

Abstract

Expression of a receptor for human macrophage-colony stimulating factor (M-CSF or CSF-1), containing a point mutation which changes an aspartate to a valine at position 802 of the activating loop of the kinase domain, potently transforms the haemopoietic cell line FDC-P1 yet prevents Rat-2 ®broblast transformation. In order to understand this apparent paradox, aspartate 802 was changed by cassette mutagenesis to each of the other 19 amino acids. All hydrophobic amino acid substitutions were transforming when tested in FDC-P1 cells yet inactivating when tested in Rat-2 ®broblasts. These same amino acid substitutions also activated receptor degradation, strongly suggesting a causal relationship between receptor degradation and inactivation in ®broblasts. Point mutations or small deletions of Y708 within the kinase insert region of the mutant D802V receptor partly inhibited receptor degradation. The more stable D802V receptor derivatives were able to transform both FDC-P1 cells and Rat-2 ®broblasts, so establishing that the cell speci®c e ect of the c-fmsD802V activating loop mutation is attributable to receptor degradation which accompanies kinase activation and prevents the transformation of Rat-2 but not of FDC-P1 cells.

Polyomavirus Middle T-Antigen Is a Transmembrane Protein That Binds Signaling Proteins in Discrete Subcellular Membrane Sites

“…Our results add support to the evidence that MT and certain growth factor receptors can transform fibroblasts only if correctly trafficked to the plasma membrane (2,28,38). Further investigation of this process promises to uncover novel targets for cancer drug development.…”

Activating mutation of D835 within the activation loop of FLT3 in human hematologic malignancies

“…24 In murine c-FMS, D802Q, D802R, and D802G were inactivating mutations, whereas all other mutants were active. 19 According to these results, all types of c-KIT or c-FMS mutations, which corresponded to FLT3 D835 mutations found in this study, caused constitutive activation of the receptor, suggesting that D835 mutations have adverse effects on leukemia cells. This is further supported by the report that Tyr and Val substitutions for Asp, which occupy most of the mutations found in this study (27 of 32, 84%), had the strongest kinase activity in c-KIT.…”

“…17,18 This mutation is thought to cause constitutive activation by triggering the A-loop into an active conformation. Since this Asp codon is highly conserved in RTKs, and substitutions to Tyr or Val in murine c-FMS and murine FLT3 result in constitutive activation of the receptors, 19,20 the Asp within the A-loop is likely to be a key regulatory residue of the RTKs.…”

Pediatric mastocytosis–associated KIT extracellular domain mutations exhibit different functional and signaling properties compared with KIT-phosphotransferase domain mutations

“…This suggestion is supported by the fact that ECD mutants in Rat2 cells are able to form colonies in soft-agar, whereas PTD mutants demonstrate a clear differentiation capacity program associated with limited oncogenic activity. This incapacity of PTD mutants to transform cells was also reported for other PTD mutants of class III receptors FMS 40 and FLT3. 41 PTD mutants are either not or only rarely observed in dog MCT and human GIST, confirming the lack of agressivity of this class of mutants.…”