Regulation of C/EBPbeta isoforms by MAPK pathways in HL60 cells induced to differentiate by 1,25-dihydroxyvitamin D3.
C/EBPbeta is known to be important for monocytic differentiation and macrophage function. Here, we found that expression of all three C/EBPbeta isoforms induced in HL60 cells by 1,25-dihydroxyvitamin D3 (1,25D) was upregulated in a sustained manner that correlates with the appearance of monocytic phenotype and with the G1 phase cell cycle arrest. In 1,25D-resistant HL60-40AF cells, isoforms beta-1 and beta-3 were expressed at levels comparable to 1,25D-sensitive HL60-G cells, but isoform beta-2 was difficult to detect. Treatment of sensitive HL60 cells with 1,25D resulted in predominantly nuclear localization of C/EBP isoforms beta-2 and beta-3, while a large proportion of C/EBPbeta-1 remained in the cytoplasm. Attenuation of the MEK-ERK MAPK pathway by the inhibitor PD98059 markedly reduced the expression, 1,25D-induced phosphorylation and nuclear localization of C/EBPbeta-2 and C/EBPbeta-3. Interestingly, only the lower molecular mass isoforms of C/EBPbeta phosphorylated on Thr235 were found in the nuclei, while C/EBPbeta-1 was constitutively phosphorylated and was detected principally in the cytoplasmic fraction. Although the role of C/EBPbeta isoforms in 1,25D-induced differentiation is complex, our results taken together strongly suggest that the phosphorylation of C/EBPbeta isoforms on Thr235 takes place mainly via the MEK-ERK pathway and that C/EBPbeta-2 is the principal transcription factor in this cell system.
Marcinkowska
Ewa
E
Institute of Biochemistry and Molecular Biology, University of Wroclaw, Tamka 2, 50-137 Wroclaw, Poland.
Garay
Edward
E
Gocek
Elzbieta
E
Chrobak
Agnieszka
A
Wang
Xuening
X
Studzinski
George P
GP
D022782:CCAAT-Enhancer-Binding Protein-beta / Q000502:physiology*; D002117:Calcitriol / Q000494:pharmacology*; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology*; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D018922:HL-60 Cells; D006801:Humans; D007700:Kinetics; D020935:MAP Kinase Signaling System / Q000502:physiology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D010766:Phosphorylation; D020033:Protein Isoforms / Q000502:physiology; D015854:Up-Regulation
D022782:CCAAT-Enhancer-Binding Protein-beta; D020033:Protein Isoforms; D020928:Mitogen-Activated Protein Kinases; D002117:Calcitriol