Generation of an Avian Myeloblastosis Virus (AMV) Reverse Transcriptase Prime Editor

Yi-Ting Tsai; Bruna Lopes da Costa; Salvatore Marco Caruso; Nicolas D Nolan; Sarah R Levi; Stephen H Tsang; Peter M J Quinn

doi:10.1007/978-3-031-27681-1_17

Generation of an Avian Myeloblastosis Virus (AMV) Reverse Transcriptase Prime Editor

Adv Exp Med Biol. 2023:1415:109-114. doi: 10.1007/978-3-031-27681-1_17.

Authors

Yi-Ting Tsai¹, Bruna Lopes da Costa^{1

2

3}, Salvatore Marco Caruso^{1

2

3

4}, Nicolas D Nolan^{1

2

3

4}, Sarah R Levi^{2

3}, Stephen H Tsang^{1

2

3

4

5

6}, Peter M J Quinn^{7

8}

Affiliations

¹ Department of Biomedical Engineering, Columbia University, New York, NY, USA.
² Edward S. Harkness Eye Institute, Department of Ophthalmology, Columbia University Irving Medical Center/New York-Presbyterian Hospital, New York, NY, USA.
³ Jonas Children's Vision Care, and Bernard & Shirlee Brown Glaucoma Laboratory, Department of Ophthalmology, Columbia University, New York, NY, USA.
⁴ Columbia Stem Cell Initiative, Columbia University, New York, NY, USA.
⁵ Department of Pathology & Cell Biology, Columbia University, New York, NY, USA.
⁶ Institute of Human Nutrition, Columbia University, New York, NY, USA.
⁷ Edward S. Harkness Eye Institute, Department of Ophthalmology, Columbia University Irving Medical Center/New York-Presbyterian Hospital, New York, NY, USA. pq2138@cumc.columbia.edu.
⁸ Jonas Children's Vision Care, and Bernard & Shirlee Brown Glaucoma Laboratory, Department of Ophthalmology, Columbia University, New York, NY, USA. pq2138@cumc.columbia.edu.

PMID: 37440022
DOI: 10.1007/978-3-031-27681-1_17

Abstract

Prime editing (PE) is a novel, double-strand break (DSB)-independent gene editing technology that represents an exciting avenue for the treatment of inherited retinal diseases (IRDs). Given the extensive and heterogenous nature of the 280 genes associated with IRDs, genome editing has presented countless complications. However, recent advances in genome editing technologies have identified PE to have tremendous potential, with the capability to ameliorate small deletions and insertions in addition to all twelve possible transition and transversion mutations. The current PE system is based on the fusion of the Streptococcus pyogenes Cas9 (SpCas9) nickase H840A mutant and an optimized Moloney murine leukemia virus (MMLV) reverse-transcriptase (RT) in conjunction with a PE guide RNA (pegRNA). In this study, we developed a prime editor based on the avian myeloblastosis virus (AMV)-RT and showed its applicability for the installation of the PRPH2 c.828+1G>A mutation in HEK293 cells.

Keywords: AMV-RT prime editors; Avian myeloblastosis virus (AMV); PRPH2; Prime editing; Retinitis pigmentosa (RP).

MeSH terms

Animals
Avian Myeloblastosis Virus* / genetics
Avian Myeloblastosis Virus* / metabolism
CRISPR-Cas Systems
Gene Editing
HEK293 Cells
Humans
Mice
Moloney murine leukemia virus / genetics
Moloney murine leukemia virus / metabolism
RNA-Directed DNA Polymerase* / genetics
RNA-Directed DNA Polymerase* / metabolism

Substances

RNA-Directed DNA Polymerase