Fig. 2
Mov10 binds repeat element RNA and mRNA in P2 brain and blocks retrotransposition. a Results of RIP followed by sequencing. Left pie chart shows the distribution of all immunoprecipitated RNAs. Right pie chart shows the classification of the repeat elements. b RT-PCR of Mov10 or irrelevant (IRR) IP from P2 brains for Mov10 individual nucleotide cross-linking immunoprecipitation (iCLIP) target mRNA (Prrc2b) and an active mouse L1 RNA (mL1T F ) and for the mRNA Grin2A, which does not bind Mov10 [2]. c, d q-PCR of DNAse I- and RNAse H-treated genomic DNA isolated from P3 heterozygote (HET) or WT littermate brain (n = 3) amplified with ORF2 primers and 5S rDNA for normalization. Values plotted relative to adult genomic DNA content. Error bars represent standard error of the mean (SEM), *p<0.05 (Student’s t test, two-tailed). e Representative gel images of the reverse transcriptase assay set up as shown in the table; SuperScript III Reverse Transcriptase (SSRTIII) was preincubated with the indicated concentrations of purified Mov10 or human purified recombinant Fmrp as a control, followed by RT-PCR of RNAs bound by Mov10 (Prrc2b or mL1T F ) or not (Gapdh). f RT-qPCR of Prrc2b, mL1TF, and Gapdh with indicated ratios of Mov10 and SSRTIII. Biological replicates are shown, and the fold change was compared to the RT-only reaction of each gene. Error bars represent standard deviation (SD), *p < 0.05 (Student’s t test, two-tailed). g Representative gel images for the RT assay using equimolar amounts of the Mov10 helicase-deficient mutant and SSRTIII. h Capture assay with WT, C-terminal and N-terminal of Mov10, and SSRTIII or bovine serum albumin (BSA) covalently coupled to beads. i Mov10 or IRR IP from P2 brains immunoblotted for L1-ORF2 (representative image of n = 3). The averages from biological replicates of the ratio between ORF2 and Mov10 for each lane are indicated below, the p values of which are not significant