Syndromic features and mild cognitive impairment in mice with genetic reduction on p300 activity: Differential contribution of p300 and CBP to Rubinstein–Taybi syndrome etiology
Introduction
Rubinstein–Taybi syndrome (RSTS, OMIM #180849) is a complex autosomal-dominant disease characterized by mental impairment, growth retardation and characteristic skeletal abnormalities, such as apparent hypertelorism, prominent nose, malpositioned ears and broad thumbs and toes (Rubinstein and Taybi, 1963). Petrij and colleagues (1995) discovered 14 years ago that about 40% of the individuals diagnosed with RSTS carried heterozygous mutations in CREBBP, the gene encoding for the CREB binding protein (CBP). Later studies have increased this number up to 60% of the individual diagnosed with this condition (Bentivegna et al., 2006), and revealed that a small percentage (∼ 3%) of cases diagnosed as RSTS were caused by mutations of the gene EP300, which encodes for the co-activator p300 that is highly homologous to CBP (Bartholdi, 2007, Roelfsema, 2005, Zimmermann, 2007).
CBP and p300 proteins function as co-factors for many transcription factors (Chan and La Thangue, 2001) including several transcription factors known to be regulated or induced by neuronal activity, such as CREB, c-Fos, c-Jun, or NF-κβ. In addition, both proteins have histone acetyltransferase (HAT) activity that targets the N-terminal unstructured tails of histones and may contribute to transcriptional activation by relaxing the structure of the nucleosomes (Ogryzko et al., 1996). Their role in activity-dependent chromatin modification and gene expression may account for at least part of the cognitive and physiological alterations associated to mental retardation in RSTS patients.
Although the primary sequence of p300 and CBP are more than 70% similar and they have many common interaction partners, these two proteins have distinct cellular functions and cannot always replace one another (Kalkhoven, 2004). The recent characterization of a number of mouse strains deficient in CBP activity has provided important new insight into the role of CBP in RSTS molecular etiology and opened new therapeutic avenues to treat this condition (Alarcon, 2004, Bourtchouladze, 2003, Korzus, 2004, Wood, 2006, Wood, 2005; reviewed by Barco, 2007). In contrast, the role of p300 in cognition and behavior remains poorly explored, although it has been recently shown that conditional transgenic mice expressing an inhibitory truncated form of p300 had mild memory deficits (Oliveira et al., 2007).
Here, we carried out a comprehensive analysis, including anatomical, behavioral, biochemical and gene profiling assays, of mice bearing a deleted p300 allele. Our results demonstrate skeletal abnormalities reminiscent of RSTS, mild motor and cognitive deficits and enhanced anxiety in the elevated plus maze, but unaltered levels of acetylated histones and negligible changes in gene expression except for p300 itself. In agreement with recent conclusions of studies in human patients, the comparison of the behavioral phenotypes of cbp and p300 mutants indicates that CBP, as compared to p300, plays a major role in cognitive abilities.
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Animals
The generation of cbp+/− (Tanaka et al., 1997) and p300+/− (Yao et al., 1998) mice has been described before. The neo cassette replaced the sequences stretching exons 4–6 of EP300 and exon 2 of crebbp, respectively. CBP deficient mice were maintained in a mixed genetic background (F2 of C57BL/6J and DBA) because they cannot be maintained into a pure C57BL/6J background (Alarcon et al., 2004). p300 deficient mice were maintained in a C57BL/6J pure background at least otherwise is indicated. Mice
Facial dysmorphia and other syndromic features in p300 deficient mice
Although preliminary studies indicated that mice bearing an inactive p300 allele did not show any gross skeletal malformation during development (Yao et al., 1998), we observed that a large percentage of adult p300+/− mice (∼ 75%) presented facial dysmorphia characterized by a prominent forehead and blunt nose (Fig. 1A). This observation was confirmed using computerized tomography (CT) analysis, which revealed normal cranial length and width, but shortened and depressed nasal bridge in p300+/−
Discussion
We carried out a comprehensive neurological characterization of adult p300+/− mice. We did not observe gross neuroanatomical changes or alterations in histone acetylation or gene expression. Our analysis, however, revealed impaired growth and facial dysmorphia, two traits associated to RSTS in humans. We also observed abnormal gait, reduced swimming speed, enhanced anxiety in the elevated plus maze, and mild cognitive impairment during the transfer task in the water maze. Behavioral studies of
Financial disclosures
No conflict of interest is declared.
Acknowledgments
The authors thank Manuel Desco and Francisca Mulero for their help in the acquisition and analysis of CT data and Maria Jimenez-Minchan for excellence technical assistance in histological preparations. We also thank Luis M. Valor and Lidia Larizza for critical reading of the manuscript and for helpful comments. This work was supported by the European Commission grant MEXT-CT-2003-509550, the Spanish Ministry of Science and Innovation Grants CSD2007-00023 and SAF2008-00611, and a grant from
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