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Gain of function mutations of RTK conserved residues display differential effects on NTRK1 kinase activity
Claudia Miranda1, Giuseppe Zanotti, Sonia Pagliardini
1Department of Experimental Oncology, Istituto Nazionale Tumori, via G. Venezian 1, 20133, Milan, Italy.
Abstract:
Activation of tyrosine kinase receptors is associated with human tumors. Tumorigenic versions of several RTKs, such as Ret, Kit and Met carry activating mutations at highly conserved residues of the tyrosine kinase domain. We have investigated the effect of some of these mutations on the NTRK1/NGF receptor, for which no naturally occurring activating point mutations have been so far detected. We introduced the following mutations in NTRK1 tyrosine kinase domain: (i) D668N equivalent to Met D1246N associated to HPRC; (ii) D668V modelled on Kit D816V found in mastocytosis; (iii) M688T corresponding to Ret M918T associated to the cancer syndrome MEN2B. The Met-like mutation rendered the NTRK1 receptor more responsive to ligand, as observed for the corresponding mutation in Met. On the contrary the Kit-like D668V resulted as neutral mutation. Surprisingly, the MEN2B-like M688T completely abrogated NTRK1 receptor activity, resulting as a loss of function mutation. Our results show that the mutations tested, although involving conserved amino acids in highly homologous regions, exert distinct effects in different receptors, and suggest a very peculiar auto-inhibitory mechanism for NTRK1.
Insights
Investigating mutations in the NTRK1 receptor revealed distinct functional outcomes. While one mutation enhanced ligand response, another unexpectedly caused a loss of function, suggesting a unique auto-inhibitory mechanism.
Area of Science:
- Molecular Biology
- Oncology
- Biochemistry
Background:
- Tyrosine kinase receptor activation is linked to human cancers.
- Activating mutations in conserved residues of tyrosine kinase domains are common in RTKs like Ret, Kit, and Met.
- Naturally occurring activating point mutations in NTRK1 have not been identified.
Purpose of the Study:
- To investigate the functional impact of specific mutations on the NTRK1/NGF receptor.
- To compare the effects of mutations equivalent to those found in Met, Kit, and Ret receptors on NTRK1.
- To explore the auto-inhibitory mechanisms of the NTRK1 receptor.
Main Methods:
- Introduction of specific point mutations (D668N, D668V, M688T) into the NTRK1 tyrosine kinase domain.
- Assessment of receptor activity and ligand responsiveness following mutation.
- Comparison of mutation effects across different tyrosine kinase receptors.
Main Results:
- The Met-like D668N mutation increased NTRK1 responsiveness to its ligand.
- The Kit-like D668V mutation had a neutral effect on NTRK1 activity.
- The Ret-like M688T mutation resulted in a complete loss of NTRK1 receptor activity.
Conclusions:
- Mutations in conserved regions of tyrosine kinase domains can have distinct functional consequences depending on the specific receptor.
- The M688T mutation in NTRK1 acts as a loss-of-function mutation, unlike its activating role in Ret.
- NTRK1 appears to possess a unique auto-inhibitory mechanism that influences the outcome of these mutations.