The authors declare no competing financial interests. Data availability HTGTS V(D)J-seq, Hi-C, 3C-HTGTS, GRO-Seq and ChIP-Seq sequencing data reported in this study has been deposited in the GEO database under the accession number GSE130224. Specifically, HTGTS V(D)J-seq data is deposited in the GEO database under the accession number GSE130216 and is related to Fig. 1e-h; 2b, c; 3a-c, e; 4c; Extended Data Fig. 2c-e, g, h; 3e, f; 4a, c, d; 5a-c; 6c; 7d, e; 9b; and Supplementary Information Table 1&2. Hi-C data is deposited in the GEO database under accession number GSE134543 and is related to Extended Data Fig. 8a. 3C-HTGTS data is deposited in the GEO database under the accession number GSE130214 and is related to Fig. 3f; 4d; and Extended Data Fig. 8a; 9c; 10q, r. GRO-Seq data is deposited in the GEO database under the accession number GSE130215 and is related to Fig. 3d; 4b; and Extended Data Fig. 4e; 6d; 7f; 9b. ChIP-Seq data is deposited in the GEO database under the accession number GSE130213 and is related to Extended Data Fig. 8c, d; 9d. Code availability. HTGTS V(D)J-seq and 3C-HTGTS data was processed through published pipeline available at (http://robinmeyers.github.io/ transloc_pipeline/). Code for Hi-C data process is available at (github.com/aidenlab). GRO-Seq and ChIP-Seq were aligned to mm9 genome with bowtie2 v2.2.8 (http://bowtie-bio.sourceforge.net/bowtie2/index.shtml), processed by samtools v1.8 (https:// sourceforge.net/projects/samtools/files/samtools/1.8/) and generated graph files via RSeQC tool v2.6 (http://rseqc.sourceforge.net/ #bam2wig-py).

The neurotrophin brain-derived neurotrophic factor (BDNF) and its receptor TrkB participate in diverse neuronal functions, including activity-dependent synaptic plasticity that is crucial for learning and memory. On binding to BDNF, TrkB is not only autophosphorylated at tyrosine residues but also undergoes serine phosphorylation at S478 by the serine/threonine kinase cyclin-dependent kinase 5 (Cdk5). However, the in vivo function of this serine phosphorylation remains unknown. We generated knock-in mice lacking this serine phosphorylation (Trkb(S478A/S478A) mice) and found that the TrkB phosphorylation-deficient mice displayed impaired spatial memory and compromised hippocampal long-term potentiation (LTP). S478 phosphorylation of TrkB regulates its interaction with the Rac1-specific guanine nucleotide exchange factor TIAM1, leading to activation of Rac1 and phosphorylation of S6 ribosomal protein during activity-dependent dendritic spine remodeling. These findings reveal the importance of Cdk5-mediated S478 phosphorylation of TrkB in activity-dependent structural plasticity, which is crucial for LTP and spatial memory formation.