Hyperglycemia promotes p53-Mdm2 interaction but reduces p53 ubiquitination in RINm5F cells.
The apoptosis of β cells induced by hyperglycemia has been associated with p53 mobilization to mitochondria and p53 phosphorylation. Murine double minute 2 (Mdm2) induces the degradation of p53 and thereby protects cells from apoptosis. We studied the effect of glucose at high concentration on the ability of Mdm2 to ubiquitinate p53 and promote its degradation. RINm5F cells were grown in RPMI-1640 medium with 5 or 30 mM glucose for varying periods of time. After this treatment, the expression of Mdm2 was measured using real-time PCR. The phosphorylation of Mdm2 at Ser166, p53 at Ser15, and the kinases Akt and ATM were measured by Western blotting. The formation of the p53-Mdm2 complex and p53 ubiquitination was assessed by p53 immunoprecipitation and immunofluorescence. Our results showed that high glucose reduced Mdm2 mRNA expression and protein concentration and increased Mdm2 and Akt phosphorylation, albeit with slower kinetics for Akt. It also promoted p53-Mdm2 complex formation, whereas p53 ubiquitination was suppressed. Furthermore, phosphorylation of both p53 Ser15 and ATM was increased in the presence of 30 mM glucose. These data indicate that high concentration glucose decrease the mRNA expression and cytosolic concentration of Mdm2. However, although the increase in glucose promoted the phosphorylation of Mdm2, it also decreased p53 ubiquitination, thus avoiding p53 degradation. In hyperglycemic conditions, such as diabetes mellitus, the reduction of pancreatic β cells mass is favored by stabilization of p53 in association with low p53 ubiquitination and reduced expression of Mdm2.
Unidad de Investigación Médica en Bioquímica, HE, Centro Médico Nacional Siglo XXI, IMSS, Av Cuauhtémoc 330, Col Doctores, Del. Cuauhtémoc, C.P. 06720, Mexico, DF, Mexico.
R
R
Barzalobre-Gerónimo
Barzalobre-Gerónimo
BG
Raúl
L A
LA
Flores-López
Flores-López Luis
FL
Antonio
L A
LA
Baiza-Gutman
Baiza-Gutman Luis
BG
Arturo
M
M
Cruz
Cruz
C
Miguel
R
R
García-Macedo
García-Macedo
GM
Rebeca
A
A
Ávalos-Rodríguez
Ávalos-Rodríguez
ÁR
Alejandro
A
A
Contreras-Ramos
Contreras-Ramos
CR
Alejandra
A
A
Díaz-Flores
Díaz-Flores
DF
Margarita
C
C
Ortega-Camarillo
Ortega-Camarillo
OC
Clara
D000818:Animals; D017209:Apoptosis / Q000502:physiology; D045744:Cell Line, Tumor; D005947:Glucose / Q000378:metabolism*; D006943:Hyperglycemia / Q000378:metabolism*; D050417:Insulin-Secreting Cells / Q000378:metabolism / Q000502:physiology; D008928:Mitochondria / Q000378:metabolism / Q000502:physiology; D010766:Phosphorylation / Q000502:physiology; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D051736:Proto-Oncogene Proteins c-mdm2 / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D054875:Ubiquitination / Q000502:physiology*
D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; C497208:Mdm2 protein, rat; D051736:Proto-Oncogene Proteins c-mdm2; D051057:Proto-Oncogene Proteins c-akt; D005947:Glucose