Transgenic mice expressing an inhibitory truncated form of p300 exhibit long-term memory deficits

Abstract

The formation of many forms of long-term memory requires several molecular mechanisms including regulation of gene expression. The mechanisms directing transcription require not only activation of individual transcription factors but also recruitment of transcriptional coactivators. CBP and p300 are transcriptional coactivators that interact with a large number of transcription factors and regulate transcription through multiple mechanisms, including an intrinsic histone acetyltransferase (HAT) activity. HAT activity mediates acetylation of lysine residues on the amino-terminal tails of histone proteins, thereby increasing DNA accessibility for transcription factors to activate gene expression. CBP has been shown to play an important role in long-term memory formation. We have investigated whether p300 is also required for certain forms of memory. p300 shares a high degree of homology with CBP and has been shown to interact with transcription factors known to be critical for long-term memory formation. Here we demonstrate that conditional transgenic mice expressing an inhibitory truncated form of p300 (p300Delta1), which lacks the carboxy-terminal HAT and activation domains, have impaired long-term recognition memory and contextual fear memory. Thus, our study demonstrates that p300 is required for certain forms of memory and that the HAT and carboxy-terminal domains play a critical role.

Figure 1.

p300 expression in the brain. Fluorescent immunohistochemistry of p300 in brain coronal sections from naïve C57Bl/6J mice. p300 is ubiquitously expressed in the mouse forebrain including the CA1, CA3, and DG regions of the hippocampus (Hp) (A,B), amygdala (Amy) (C), and cortex (Cx) (D).

Figure 2.

Generation and characterization of p300Δ1 transgenic mice. (A) Schematic representation of the truncated form of p300 (p300Δ1). p300Δ1 lacks the bromo and histone acetyltransferase (HAT) domains contained within the carboxy-terminal portion of this protein. (KIX) Kinase inducible interaction domain. (B) Transient transfection of p300Δ1 and a CRE-luciferase reporter in HEK293 cells showed that p300Δ1 significantly reduces CRE-dependent transcription induced by forskolin and IBMX. (C) Schematic representation of the tetracycline system used to regionally and temporally control the expression of p300Δ1 transgene in mice. p300Δ1 transgene is under the control of a tetracycline operator sequence (tetO) and a minimal promoter and mice carrying this transgene were generated and crossed to mice that contain the tTA transactivator driven by the CaMKIIα promoter (line B, Mayford et al. 1996) giving rise to bitransgenic mice (referred to in this paper as p300Δ1 transgenic mice). In the absence of doxycycline (dox), p300Δ1 is expressed in postnatal forebrain neurons and in the presence of doxycycline p300Δ1 expression should be turned off. (D) RT-PCR analyses of cDNA synthesized from RNA isolated from amygdala (Amy), hippocampus (Hp), cortex (Cx), and cerebellum (Cb) of wild-type and p300Δ1 transgenic mice on and off dox. (E) Western blot analysis of levels of acetylated lysines K9 and K14 in histone H3 (AcH3) in forebrain of wild-type (wt) and p300Δ1 transgenic mice on and off dox (n = 6 per group) normalized to the levels of actin in each sample.

Figure 3.

p300Δ1 transgenic mice show normal spatial memory in the Morris water maze. (A) Wild-type (n = 8) and p300Δ1 transgenic (n = 8) mice do not show a significant difference in the latency to find the hidden platform during the acquisition phase of the task. (B) During probe trial (performed after session 8), p300Δ1 transgenic mice and wild-type littermates showed a similar percentage of time spent swimming in the target quadrant. (C) During the probe trial (performed after session 8) the number of crossings over the area where the platform was located during acquisition was not different between wild-type and p300Δ1 transgenic mice.

Figure 4.

CBPΔ1 and p300Δ1 transgenic mice show impaired long-term recognition memory. (A) CBPΔ1 transgenic mice (n = 5) and wild-type littermates (n = 6) show similar preference for the novel object when tested 30 min after training. However, when tested 24 h after training, CBPΔ1 transgenic mice (n = 9) show significant lower preference for the novel object than their wild-type littermates (n = 8). (B) p300Δ1 transgenic mice (n = 16) and wild-type littermates (n = 16) show similar short-term recognition memory tested 30 min after training. However, p300Δ1 transgenic mice (n = 15) show impaired novel object recognition tested 24 h after training compared with their wild-type littermates (n = 15). (C) p300Δ1 transgenic mice (n = 10) and wild-type littermates (n = 12) fed with doxycycline show similar recognition memory tested 24 h after object familiarization.

Figure 5.

p300Δ1 transgenic mice show impaired long-term contextual fear memory. (A) p300Δ1 transgenic mice (n = 18) show significant lower percentage of freezing compared with their wild-type littermates (n = 19) when tested 24 h after training in the contextual fear conditioning. (B) p300Δ1 transgenic mice fed with doxycycline (n = 20) show a reduced (but not statistically significant) percentage of freezing compared with their wild-type littermates (n = 20). (C) p300Δ1 transgenic mice (n = 10) and wild-type littermates (n = 10) show comparable contextual fear memory tested 1 h after training. (D) p300Δ1 transgenic mice (n = 7) and wild-type littermates (n = 7) show comparable long-term cued fear memory tested 24 h after training.