Translational regulation of C/EBPs
Model of integrated control of osteoclastogenesis by C/EBPβ.Top to Bottom: Long (LAP) and truncated (LIP) C/EBPβ isoforms are generated from a single mRNA. The ratio between LAP and LIP is determined by the activity of translation initiation factors which are downstream of growth factor, hormone, nutrient, and stress signaling pathways. The mammalian Target Of Rapamycine (mTOR) kinase integrates these signals and plays an important role in adjusting the LAP/LIP ratio (blue: low activity; red: high activity). The LAP isoform induces MafB, an inhibitor of osteoclastogenesis that suppresses the activity of several osteogenic regulators (Mitf, Fos, NFATc1). Osteoclast precursors that lack C/EBPβ or that express LIP only, display strongly augmented formation of giant osteoclasts that readily destroy bone substance.
Klaus Wethmar & Julia Schulz
C/EBPα and C/EBPβ transcripts harbor small upstream open reading frames (uORF) in their mRNAs that sense the activity of translation initiation factors and relay initiation to alternative in-frame start sites. As a result, truncated proteins that lack part of the N-terminal sequences are generated. Besides C/EBPs, many transcripts of key regulatory genes involved in growth, differentiation, and proliferation harbor uORFs, suggesting an important role of uORF-mediated translational control in mammalian development and physiology. Nevertheless, genetic models in mammals are lacking to explore the physiological relevance of uORF-regulated translation initiation.
Truncated C/EBP isoforms sustain proliferation, whereas full-length forms are inhibitors of cell division.Previously, we showed that anaplastic large cell lymphoma and Hodgkin Lymphoma express predominantly truncated C/EBPβ. Rapamycin, an antibiotic that inhibits mTOR signaling, shuts down the truncated C/EBP isoform and concomitantly inhibits growth in both types of lymphomas. Ectopic expression of truncated C/EBPβ restored proliferation, suggesting that truncated C/EBPβ represents a translationally controlled oncogene.
By targeted recombination we have now generated mouse C/EBPβ mutants that express distinct C/EBPβ isoforms. The murine mutants revealed that C/EBPβmisoform switching is important during liver regeneration, bone homeostasis and tumorigenesis. Mice deficient for the C/EBPβ uORF initiation codon(C/EBPβΔuORF) fail to initiate translation of the autoantagonistic, truncated “LIP” C/EBPβ isoform. Livers of C/EBPβΔuORF mice displayed defective regeneration. After partial hepatectomy, delayed and blunted entry of hepatocytes into S-phase, persistent repression of E2Fregulated genes, and hyperactivation of acute phase response genes became evident. Thus, switching to the truncated isoform is important during liver regeneration and in shutting off acute phase response.
C/EBPβ deficient mice and mice expressing only the truncated “LIP” isoform (LIP knock-in mice, L/L) both display increased bone resorption. This is due to enhanced differentiation of the bone resorbing cell, the osteoclast. Failure to switch back to the long C/EBPβ isoform augmented osteoclastogenesis. Indeed, ectopic expression of the long isoform “LAP” in monocytes inhibited formation of multi-nucleated osteoclasts. Rapamycin, an inhibitor of mTOR signaling that increases the “LAP” over “LIP” ratio, also inhibited osteoclastogenesis in wild type but not in C/EBP deficient or L/L osteoclasts that both can not switch to “LAP” expression. Profiling analysis of the transcriptome showed that rapamycin treatmentor ectopic expression of “LAP” activated expression of MafB, a negative regulator of osteoclastogenesis.This suggested that restriction of osteoclastogenesis by “LAP” or rapamycin is dependent on MafB. Inaccordance, knock-down of MafB induced osteoclastogenesis, regardless of rapamycin treatment or C/EBP isoform expression. Altogether, the data showed that differential regulation of MafB gene expression by C/EBPβ isoforms determines the balance of bone turnover and that the control of C/EBPβisoform translation represents a target for osteoporosis treatment.