Protein transduction by pseudotyped lentivirus-like nanoparticles

Area of Science:

• Biotechnology
• Molecular Biology
• Cell Biology

Background:

• Establishing efficient and reproducible methods for protein transduction into mammalian cells remains a challenge.
• Current methods may require highly purified proteins or can damage cellular DNA.

Purpose of the Study:

• To develop a novel, simple, efficient, and reproducible method for protein transduction into mammalian cells.
• To demonstrate the feasibility of using lentivirus-like nanoparticles (LENAs) for protein delivery.

Main Methods:

• Development of a lentiviral vector system to package foreign proteins into lentivirus-like nanoparticles (LENAs).
• Fusion of a reporter protein, beta-lactamase (BlaM), to Gag for incorporation into LENAs.
• Production of BlaM-containing LENAs via co-transfection with VSV-G pseudotyping.
• Transduction of mammalian cells (293T and MT-4) with BlaM-loaded LENAs and detection of BlaM activity.

Main Results:

• Successfully produced LENAs containing thousands of beta-lactamase (BlaM) enzyme molecules per capsid.
• Demonstrated efficient transduction of BlaM into the cytoplasm of mammalian cells via LENA infection.
• Confirmed BlaM activity through cleavage of a fluorescent substrate (CCF2-AM).
• Showed that LENA-mediated protein transduction does not damage cellular DNA.

Conclusions:

• LENA technology provides a novel and effective method for transient protein transduction in mammalian cells.
• This approach bypasses the need for highly purified proteins and avoids cellular DNA damage.
• LENA-mediated protein delivery holds significant potential for diverse basic research and clinical applications.