Members of the C/EBP family of transcription factors play important roles in mediating the acute phase response, an inflammatory process resulting from infection and/or tissue damage. Among the C/EBP family of proteins, C/EBPbeta and delta were thought to be the primary mediators of the acute phase response along with STAT3 and NF-kappaB. However, when LPS or IL-1 were used as inflammatory stimuli in neonatal mice harboring a targeted disruption in the C/EBPalpha gene, markers of the acute phase response were completely lacking. Two important components of the acute phase response appeared to be affected by the C/EBPalpha deletion. First, NF-kappaB complexes were not identical in LPS stimulated knockout and wild type mice and second, the composition and relative quantities of the various C/EBPbeta truncated isoforms differed in the livers of C/EBPalpha knockout and wild type animals. Truncated C/EBPbeta isoforms are thought to be generated by translation initiation at internal AUGs in the C/EBPbeta mRNA. However, the principal investigator has discovered a novel posttranslational method of C/EBPbeta isoform formation involving specific proteolytic cleavage of the full-length protein. The proteolytic activity is a calpain that is regulated by C/EBPalpha and is not present in C/EBPalpha knockout mice. The C/EBPalpha-dependent calpain also cleaves cyclin A. All of these observations will be pursued as part of this continuation application which contains four specific aims.
In aim 1, the identity of the components of the NF-kappaB complexes in C/EBPalpha knockout mice will be determined as will the molecular basis for C/EBPalpha regulation of NF-kappaB complex formation and the activity of the novel NF-kappaB complex.
In aim 2, the C/EBPalpha-dependent clapain will be cloned and biochemical basis for its regulation by C/EBPalpha determined.
In aim 3 the consequences of elevated levels of full length cyclin A will be pursued. The PI has noted that hepatocyte proliferation is increased in the livers of newborn C/EBPalpha knockout mice and the relationship between cyclin A cleavage and cdk2 phosphorylation will be examined.
In aim 4, the biological consequences of the truncated isoforms of C/EBPbeta will be examined in transgenic and cell culture models.
|Singh, Pallavi; Goode, Triona; Dean, Adam et al. (2011) Elevated interferon gamma signaling contributes to impaired regeneration in the aged liver. J Gerontol A Biol Sci Med Sci 66:944-56|
|Singh, Pallavi; Coskun, Zeynep Z; Goode, Catriona et al. (2008) Lymphoid neogenesis and immune infiltration in aged liver. Hepatology 47:1680-90|
|Yoshida, Yuichi; Hughes, Douglas E; Rausa 3rd, Francisco M et al. (2006) C/EBPalpha and HNF6 protein complex formation stimulates HNF6-dependent transcription by CBP coactivator recruitment in HepG2 cells. Hepatology 43:276-86|
|Yang, Jianqi; Croniger, Colleen M; Lekstrom-Himes, Julie et al. (2005) Metabolic response of mice to a postnatal ablation of CCAAT/enhancer-binding protein alpha. J Biol Chem 280:38689-99|
|Mackey, Stephanie L; Darlington, Gretchen J (2004) CCAAT enhancer-binding protein alpha is required for interleukin-6 receptor alpha signaling in newborn hepatocytes. J Biol Chem 279:16206-13|
|Welm, Alana L; Timchenko, Nikolai A; Ono, Yasuko et al. (2002) C/EBPalpha is required for proteolytic cleavage of cyclin A by calpain 3 in myeloid precursor cells. J Biol Chem 277:33848-56|
|Welm, A L; Timchenko, N A; Darlington, G J (1999) C/EBPalpha regulates generation of C/EBPbeta isoforms through activation of specific proteolytic cleavage. Mol Cell Biol 19:1695-704|
|Darlington, G J (1999) Molecular mechanisms of liver development and differentiation. Curr Opin Cell Biol 11:678-82|
|Burgess-Beusse, B L; Timchenko, N A; Darlington, G J (1999) CCAAT/enhancer binding protein alpha (C/EBPalpha) is an important mediator of mouse C/EBPbeta protein isoform production. Hepatology 29:597-601|
|Burgess-Beusse, B L; Darlington, G J (1998) C/EBPalpha is critical for the neonatal acute-phase response to inflammation. Mol Cell Biol 18:7269-77|