Post-transcriptional control of CCAAT/enhancer-binding protein beta (C/EBPbeta) expression: formation of a nuclear HuR-C/EBPbeta mRNA complex determines the amount of message reaching the cytosol.
In 3T3-L1 cells, HuR is constitutively expressed and prior to induction of differentiation localized predominantly to the nucleus. Within minutes of induction of differentiation, nuclear HuR binds to its target ligand mRNAs, and the complexes appear to move to the cytosol. One ligand mRNA is the CCAAT/enhancer-binding protein beta (C/EBPbeta) message. To examine the function and importance of the HuR-C/EBPbeta interaction, retroviral expression constructs were created in which the HuR binding site was altered by deletion (betadel) or deletion and substitution (betad/s). Expression of these constructs in murine embryonic fibroblasts resulted in significant adipose conversion relative to those cells expressing wild type C/EBPbeta. C/EBPbeta protein content was increased markedly in both betadel and betad/s, which correlated with the acquisition of the adipocyte phenotype. Analysis of the betad/s cell line demonstrated a robust expression of C/EBPalpha coincident with peroxisome proliferator-activated receptor gamma expression. Total C/EBPbeta mRNA accumulation indicated no difference between cells harboring either the wild type C/EBPbeta cDNA or betad/s construct. However, cytosolic C/EBPbeta mRNA in the cells expressing the betad/s construct was maintained at levels between 2- and 7-fold greater than in the cells expressing the wild type construct. Alteration in mRNA half-life was not responsible for the increased accumulation. Mechanistically, these data suggest that HuR binding results in nuclear retention of the C/EBPbeta mRNA and is consistent with HuR control, at least in part, of mRNA processing.
Cherry
Joy
J
Department of Biochemistry and Molecular Biology, The Brody School of Medicine at East Carolina University, Greenville, North Carolina 27858, USA.
Jones
Heath
H
Karschner
Vesna A
VA
Pekala
Phillip H
PH
D041721:3T3-L1 Cells; D000595:Amino Acid Sequence; D000818:Animals; D000954:Antigens, Surface / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites / Q000502:physiology; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000235:genetics / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism*; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D047495:PPAR gamma / Q000235:genetics / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D012190:Retroviridae; D017384:Sequence Deletion
D000954:Antigens, Surface; D022782:CCAAT-Enhancer-Binding Protein-beta; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; C098770:ELAVL1 protein, human; D047495:PPAR gamma; D012333:RNA, Messenger; D016601:RNA-Binding Proteins