Identification of Hp1bp3 as top candidate modulator of cognitive aging (a) The BXD panel was derived by inbreeding the F2 progeny of a C57BL/6J (B6) and DBA/2J (D2) intercross. (b) Contextual fear memory (CFM) varies widely across 21 BXD strains (n = 2–8/strain, mean age = 15 ± 0.3 mo). Memory index was quantified by percent time spent freezing during 10 min test. (c) Genetic interval mapping revealed a significant CFM QTL on mouse chromosome 4 (Chr4: 137.5–140.5 Mb). Pink horizontal line: Genome-wide statistical significance (p = 0.05), Green additive effect line: D parental allele increases trait values, red: B allele increases trait values. Yellow tick marks: presence of genome sequence variant. (d) Hp1bp3 expression is highly correlated with CFM across BXD strains (Pearson r = 0.6, p < 0.05), with strains inheriting the D allele having lower levels of hippocampal Hp1bp3 and poorer CFM.

Effect of Hp1bp3 genotype on CFM status is age-dependent. (a) Memory index for adult BXDs (age = 8–9 weeks) was calculated using CFM data generated by and is publically available on GeneNetwork.org (Trait ID 110908, see Methods section 2.3). Parental origin of Hp1bp3 allele did not significantly affect memory status in adulthood (D allele memory index = 81.0 ± 1.3, B =84.3 ± 1.3, t(54) = −1.86, p = 0.068). (b) Middle-aged (age = 15.0 ± 0.3 months) BXD strains inheriting the D parental allele performed significantly worse on CFM tests than strains inheriting the B allele (D = 4.8 ± 1.0, B = 41.5 ± 20.7, t(14) = −4.6, p < 0.001). Two-way ANOVA revealed a significant interaction between genotype and age [F(1,66) = 33.5, p < 0.001], suggesting that Hp1bp3 genotype influences memory status in an age-dependent manner.

Differences in CFM were not attributable to baseline freezing or post-shock reactivity. There were no significant effects of strain on either (a) baseline freezing [F(20,50) = 1.009, p = 0.47] or (b) length of post-shock activity burst [F(20,49) = 1.668, p = 0.07], indicating no strain-specific differences in baseline fear/anxiety or pain sensitivity, respectively, across 21 BXD strains tested.

Hp1bp3 knockout (KO) mice exhibit impairment on hippocampus-dependent long-term and working memory tasks. (a) Comparable baseline (BL) freezing and acquisition of conditioned fear (Acq. fear) by the final trial in wild-type (WT) and Hp1bp3 knock-out (KO) mice (n = 6/group) during contextual fear conditioning. (b) There were no differences in post-shock reactivity during fear conditioning training between WT and Hp1bp3 KO mice, indicating no differences in pain sensitivity. Videos were manually analyzed to determine the average length of activity burst for each mouse. WT = 2.6 ± 0.2 s, KO = 2.6 ± 0.2 s; t(1,10)= −0.11, p = 0.91. (c) KO mice were significantly impaired on CFM [t(10) = 3.10, p < 0.001)]. (d) KO mice also exhibited spatial working memory deficits [t(12) = 5.095, p < 0.001].

HP1BP3 protein is correlated with memory status in aging humans (a) Western blot shows hippocampal HP1BP3 protein is reduced in cognitively impaired elderly humans (n = 3, MMSE = 22.6 ± 3.0, age = 84 ± 4 years) relative to cognitively intact controls (n = 3, MMSE = 29.3 ± 0.3, age = 78 ± 10 years). (b) Quantification of hippocampal HP1BP3 protein (Cog. Intact = 1.02 ± 0.21 adjusted density, Impaired = 0.60 ± 0.04, t(1,4) = −3.344, p = 0.03).

Gene Set Enrichment Analysis (GSEA). All nominally significant hippocampal correlates (uncorrected p ≤ 0.05, n = 2074, see ) of Hp1bp3 were extracted from whole-genome transcriptome data from BXD strains age-matched to those used for contextual fear conditioning. The genes were sorted based on correlation coefficient, with the most highly positively correlated genes at the top of the list. Graphs illustrate calculation of enrichment score, or deviation from random distribution throughout the preranked list. Each vertical black line below the graph represents a gene set member and its position in the preranked list (red=genes positively correlated with Hp1bp3, blue=genes negatively correlated with Hp1bp3). A FDR cut-off of q ≤ 0.25 was used to identify significantly enriched sets according to established methods. (a, top) Positive correlates were significantly enriched for localization to the nucleus. (a, bottom) Genes localized to the nucleus were also present in significantly enriched functional gene sets including RNA splicing, nuclear transport, chromosome organization, cell cycle, and transcription cofactor activity. Normalized enrichment score (NES) represents enrichment score after adjustment to account for size of the given gene set. (b) Positive correlates were significantly enriched for the KEGG pathway corresponding to insulin signaling. (c, top) Negative correlates of Hp1bp3 were significantly enriched for localization to the plasma membrane. (c, bottom) Genes localized to the plasma membrane were also present in significantly enriched functional gene sets including ion channel activity, substrate-specific channel activity, transmembrane transporter activity, voltage-gated channel activity, and G-protein coupled receptor activity. (d) Correlates of Hp1bp3 also show significant overlap with a set of genes determined by to be significantly differentially expressed between Alzheimer’s disease patients and corresponding controls.