CRISPR-Cas9 gene editing creates a double-strand break (DSB) at a target sequence guided by a single-guide RNA (sgRNA). The break is repaired by either NHEJ or HDR. In therapeutic gene correction (e.g., for sickle cell disease), which repair pathway is preferred, and why?
- A Non-homologous end joining (NHEJ) is preferred because it permanently corrects the mutation without risk of off-target integration
- B Homology-directed repair (HDR) using a DNA donor template is preferred because it introduces precise corrections; NHEJ causes insertions/deletions (indels) that disrupt coding sequences ✓
- C NHEJ is preferred in post-mitotic cells such as neurons and muscle fibres where HDR is impossible
- D Microhomology-mediated end joining (MMEJ) is always chosen over HDR in clinical settings
Explanation
For therapeutic gene correction requiring precise single nucleotide changes (as in sickle cell disease), HDR using a donor template (ssODN or AAV-delivered homologous DNA) is required because it copies the correct sequence into the break site with high fidelity. NHEJ is error-prone, generating frameshift or missense indels that would further disrupt the gene. The limitation of HDR is that it is active primarily in S/G2 phases of cycling cells. Current clinical approaches for sickle cell disease instead exploit NHEJ by disrupting the BCL11A repressor of HbF (reactivating fetal hemoglobin), not correcting the HbS mutation directly — precisely because efficient HDR is difficult in haematopoietic stem cells.
Reference: Harper's Illustrated Biochemistry, 32nd ed.
High-yield for: NEET PGINI-CETNExTFMGEUSMLEPLABMRCP
Written and medically reviewed by the StethoPrep medical team.