International Journal of Molecular Sciences (Jul 2022)

The Mechanistic Understanding of RAD51 Defibrillation: A Critical Step in BRCA2-Mediated DNA Repair by Homologous Recombination

  • Fabrizio Schipani,
  • Marcella Manerba,
  • Roberto Marotta,
  • Laura Poppi,
  • Arianna Gennari,
  • Francesco Rinaldi,
  • Andrea Armirotti,
  • Fulvia Farabegoli,
  • Marinella Roberti,
  • Giuseppina Di Stefano,
  • Walter Rocchia,
  • Stefania Girotto,
  • Nicola Tirelli,
  • Andrea Cavalli

Journal volume & issue
Vol. 23, no. 15
p. 8338


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The cytotoxic action of anticancer drugs can be potentiated by inhibiting DNA repair mechanisms. RAD51 is a crucial protein for genomic stability due to its critical role in the homologous recombination (HR) pathway. BRCA2 assists RAD51 fibrillation and defibrillation in the cytoplasm and nucleus and assists its nuclear transport. BRC4 is a peptide derived from the fourth BRC repeat of BRCA2, and it lacks the nuclear localization sequence. Here, we used BRC4 to (i) reverse RAD51 fibrillation; (ii) avoid the nuclear transport of RAD51; and (iii) inhibit HR and enhance the efficacy of chemotherapeutic treatments. Specifically, using static and dynamic light scattering, transmission electron microscopy, and microscale thermophoresis, we show that BRC4 eroded RAD51 fibrils from their termini through a “domino” mechanism and yielded monomeric RAD51 with a cumulative nanomolar affinity. Using cellular assays (BxPC-3, pancreatic cancer), we show that a myristoylated BRC4 (designed for a more efficient cell entry) abolished the formation of nuclear RAD51 foci. The present study provides a molecular description of RAD51 defibrillation, an essential step in BRCA2-mediated homologous recombination and DNA repair.