Die p300 Protein Acetyltransferaseaktivität supprimiert eine dem humanen Systemischen Lupus Erythematosus ähnliche Autoimmunerkrankung in Mäusen
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Philipps-Universität Marburg
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Abstract
p300, one of 15 mammalian acetyltransferases known, interacts with a huge number of proteins which directly function as substrates for its acetyltransferase activity. Among them are several proteins involved in hematopoiesis. A conditional knock-in mouse with heterozygous expression of an acetyltransferase-deficient p300 (p300AS) in the germline is embryonic lethal, whereas heterozygous p300 knock-out mice and embryonic stem cells homozygous for the p300 mutation are viable. Moreover, several mutations of p300 acetyltransferase activity were found in different leukemia patients. All together, these data suggest a function of p300 acetyltransferase activity in hematopoiesis. Therefore, the task of my thesis work was to define the role of p300 acetyltransferase activity during B cell development and differentiation.
During my studies, I showed that mice expressing an acetyltransferase-deficient p300 only in B cells develop an autoimmune disease similar to the Systemic Lupus Erythematosus (SLE) in human and undergo premature death. The pathological manifestations of these mice include splenomegaly, glomerulonephritis, vasculitis, immune complex depositions in different organs, and production of autoantibodies against dsDNA and other nuclear antigens. A premature death of females in comparison to males was observed, reminiscent to another hallmark of human SLE which affects over 90% of women. In addition, the deficient acetyltransferase activity of p300 in B cells leads to a reduced number of transitional and marginal zone B cells in the spleen besides a partial loss of B cell maturation in these mice. Similar results are obtained by using mice having an inducible and ubiquitously expressed form of mutant p300. These results suggest that the p300 acetyltransferase activity is necessary for the regulation of self tolerance in B cells as well as for their differentiation. Moreover, the acetyltransferase activity of p300 suppresses the development of an SLE-like autoimmune disease.
In addition to the observed B cell phenotype, I prove that B cells expressing acetyltransferase-deficient p300 are still able to produce in vivo immunoglobulins with a specific increase for the isotypes IgG2b and IgM. FACS analysis show that the increased levels of total immunoglobulins are correlating with higher amounts of activated B cells in the spleen of those mice. An analysis of the humoral immune response against a T-cell-dependent antigen, in that case ovalbumin, reveals increased Ovalbumin-specific antibodies before the immunization. Whereas, after 14 days of immunization the primary immune response against ovalbumin of mice deficient for the p300 acetyltransferase activity in B cells is similar to control mice. In contrast, mice show an impaired memory response. Notably, in addition to an increased amount of activated B cells, p300 acetyltransferase-deficient mice show a significant augmented number of activated T cells independently of ovalbumin.
Since the mice expressing the B-cell-specific acetyltransferase-deficient p300 are displaying B cell activation after administration of the antigen, the proliferation of the B cells is analyzed after activation of different signaling pathways ex vivo. Only the activation of the BCR signaling pathway via addition of α-IgM antibodies leads to a reduced proliferation of the B cells. Notably, the reduced amount of proliferating cells is not due to an increased apoptosis rate suggesting a blockage of the proliferation by the acetyltransferase activity of p300. In addition, an observed deregulation of the BCR signaling pathway in microarray analysis in those B cells is consistent with that suggestion.
In order to check whether the presence of an acetyltransferase activity deficient form of p300 had an incidence on the overall acetylation status, biochemical analyses of the histone acetylation levels didn’t show any difference in the global or promoter-specific histone H3K18 acetylation in B cells with reduced p300 acetyltransferase activity. Rather, a reduced acetylation of proteins larger than histones is detectable suggesting that other proteins might be the critical substrates of the p300 acetyltransferase activity in vivo. For further analyses, I established a B cell line via homologous recombination expressing the mutant p300AS. This cell line which shows a reduced acetyltransferase activity in vitro will provide a helpful tool for the identification of the substrates which are dependent on the p300 acetyltransferase activity through proteomics or other analyses.
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Created: 2008Issued: 2008-12-01Updated: 2011-08-10
Faculty
Medizin
Publisher
Philipps-Universität Marburg
Language
ger
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DoctoralThesis
Keywords
MausmodellMouse Modelp300p300SLESLESystemic Lupus ErythematosusAutoimmunityAcetylation
DFG-subjects
AcetylierungSystemischer ErythematodesAutoimmunität
DDC-Numbers
610
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Forster, Nicole: Die p300 Protein Acetyltransferaseaktivität supprimiert eine dem humanen Systemischen Lupus Erythematosus ähnliche Autoimmunerkrankung in Mäusen. : Philipps-Universität Marburg 2008-12-01. DOI: https://doi.org/10.17192/z2008.0921.
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This item has been published with the following license: In Copyright