Targeted resolution of G4-DNA at the Chl1 and Gphn locus impairs memory. A, Design of the lentiviral construct CRISPR-dCas9-DHX36. B, Infusions of the constructs had no significant effect on within-session extinction learning. C, Timeline of training and lentivirus infusion into the ILPFC of a CRISPR-dCAS9-DHX36. This construct was directed to either a control or Gphn locus. D, It led to a significant impairment in memory for fear extinction following a 60 CS training protocol in Context B as there was a significant difference between the two EXT groups (n = 6–8/group, two-way ANOVA, F_(3,21) = 26.19, p < 0.0001, Tukey’s post hoc test; RC dCas9-DHX36 control guide vs EXT dCas9-DHX36 control guide, ****p < 0.0001; RC dCas9-DHX36 control guide vs EXT dCas9-DHX36 Gphn guide, ****p < 0.0001; EXT dCas9-DHX36 control guide vs EXT dCas9-DHX36 Gphn guide, *p = 0.0453). E, Furthermore, in Context A, only the control EXT group showed significant reduction (n = 6–8/group, two-way ANOVA, F_(3,21) = 7.225, p = 0.0016, Tukey’s post hoc test; RC dCas9-DHX36 control guide vs EXT dCas9-DHX36 control guide, ***p = 0.0003). F, CRISPR-dCAS9-DHX36 directed to the Gphn locus led to a significant decrease in mRNA expression (n = 7/group, t test t₍₁₁₎ = 2.76, *p < 0.05). G, Lentivirus infusion into the infralimbic PFC of a CRISPR-dCAS9-DHX36 construct directed to either a control or Chl1 locus led to a significant reduction in the expression of the original fear memory in Context B with no impairment of extinction (n = 6–8/group, two-way ANOVA, F_(3,24)= 15.67, p < 0.0001, Tukey’s post hoc test; RC dCas9-DHX36 control guide vs EXT dCas9-DHX36 control guide, ****p < 0.0001; RC dCas9-DHX36 control guide vs EXT dCas9-DHX36 Chl1 guide, ****p < 0.0001; RC dCas9-DHX36 control guide vs RC dCas9-DHX36 Gphn guide, **p = 0.0071). H, In Context A, there was no significant impairment of expression of original fear or the extinction memory by Chl1-DHX36-dCas9 (n = 6–8/group, two-way ANOVA, F_(3,24)= 7.26, p < 0.0001, Tukey’s post hoc test; RC dCas9-DHX36 control guide vs EXT dCas9-DHX36 control guide, ***p = 0.0008; RC dCas9-DHX36 control guide vs EXT dCas9-DHX36 Chl1 guide, ***p = 0.0001). I, CRISPR-dCAS9-DHX36 directed to the Chl1 locus led to a significant increase in mRNA expression (n = 6/group, t test t₍₁₁₎ = 2.48, *p < 0.05).