Journal: Learning & Memory

Article Title: Context memory formation requires activity-dependent protein degradation in the hippocampus

doi: 10.1101/lm.045443.117

Figure Lengend Snippet: Proteolytic activity in the dHPC increases immediately and quickly returns to baseline while the vHPC exhibits a biphasic pattern of proteolytic following context exposure. Rats were given 5 min of context exposure, and dHPC and vHPC tissue were collected 0, 30, 60, 90, or 120 min later for Western blotting and in vitro proteasome activity assay. ( A ) Phosphorylation of Rpt6 normalized to β-actin and expressed as a percentage of the No Exposure (NE) control group. The inset is a representative section of dissected dHPC (light gray) tissue. Phosphorylated Rpt6 marginally increased at 0 and 30 min following context exposure and returned to baseline levels by 60 min. ( B ) Raw fluorescent units (RFU) for the in vitro 20S proteasome assay for dHPC chymotrypsin, peptidylglutamyl, and trypsin activities expressed as a percentage of the NE control. Chymotrypsin, peptidylglutamyl, and trypsin activities significantly increased immediately following context exposure and returned to baseline by 30 min. ( C ) Phosphorylation of Rpt6 normalized to β-actin and expressed as a percentage of the No Exposure (NE) control group. The inset is a representative section of dissected vHPC (dark gray) tissue. Phosphorylated Rpt6 significantly increased immediately and following context exposure and returned to baseline by 30 min. ( D ) Raw fluorescent units (RFU) for the in vitro 20S proteasome assay for vHPC chymotrypsin, peptidylglutamyl, and trypsin activities expressed as a percentage of the NE control. Chymotrypsin and trypsin activities significantly increased immediately following context exposure and returned to baseline by 30 min. Peptidylglutamyl activity significantly increased 90 min following context exposure and returned to baseline by 120 min. (*) P < 0.05, (**) P < 0.01.

Article Snippet: Primary antibodies included a custom phospho-Rpt6-Serine 120 (pS120) rabbit polyclonal antibody generated commercially (ProSci) (described in ), pERK (1:500, Cell Signaling), and β-actin (1:1000, Cell Signaling).

Techniques: Activity Assay, Western Blot, In Vitro