Functional characterization of MODY2 mutations in the nuclear export signal of glucokinase

BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR BASIS OF DISEASE

卷:1864

Article

Gutierrez-Nogues, Angel¹  Garcia-Herrero, Carmen-Maria¹  Oriola, Josep²  Vincent, Olivier³  Navas, Maria-Angeles^1,4,5 

[1] Univ Complutense Madrid, Fac Med, Dept Bioquim & Biol Mol, Madrid, Spain

[2] Univ Barcelona, Hosp Clin, Dept Ciencias Fisiol 1, Serv Bioquim & Genet Mol,Fac Med, Barcelona, Spain

[3] Univ Autonoma Madrid, Consejo Super Invest Cient, Inst Invest Biomed Alberto Sols, Madrid, Spain

[4] Inst Salud Carlos III, Ctr Invest Biomed Red Diabet & Enfermedades Metab, Madrid, Spain

[5] Hosp Clin San Carlos IdISSC, Inst Invest Sanitaria, Madrid, Spain

关键词: MODY;    Glucokinase;    Glucokinase mutation;    Glucokinase regulatory protein;    Nuclear export signal;   

DOI  :  10.1016/j.bbadis.2018.04.020

来源: Elsevier

【 摘 要 】

Glucokinase (GCK) plays a key role in glucose homeostasis. Heterozygous inactivating mutations in the GCK gene cause the familial, mild fasting hyperglycaemia named MODY2. Besides its particular kinetic characteristics, glucokinase is regulated by subcellular compartmentation in hepatocytes. Glucokinase regulatory protein (GKRP) binds to GCK, leading to enzyme inhibition and import into the nucleus at fasting. When glucose concentration increases, GCK-GKRP dissociates and GCK is exported to the cytosol due to a nuclear export signal (NES). With the aim to characterize the GCK-NES, we have functionally analysed nine MODY2 mutations located within the NES sequence. Recombinant GCK mutants showed reduced catalytic activity and, in most cases, protein instability. Most of the mutants interact normally with GKRP, although mutations L306R and L309P impair GCK nuclear import in cotransfected cells. We demonstrated that GCK-NES function depends on exportin 1. We further showed that none of the mutations fully inactivate the NES, with the exception of mutation L304P, which likely destabilizes its a-helicoidal structure. Finally, we found that residue G1u300 negatively modulates the NES activity, whereas other residues have the opposite effect, thus suggesting that some of the NES spacer residues contribute to the low affinity of the NES for exportin 1, which is required for its proper functioning. In conclusion, our results have provided functional and structural insights regarding the GCK-NES and contributed to a better knowledge of the molecular mechanisms involved in the nucleo-cytoplasmic shuttling of glucokinase. Impairment of this regulatory mechanism by some MODY2 mutations might contribute to the hyperglycaemia in the patients.

【 授权许可】

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