Mechanisms of iPS cell generation and beyond

Area of Science:

• Cellular reprogramming and regenerative medicine
• Epigenetic landscape and cellular identity
• Gene regulation and cell fate determination

Background:

• Induced pluripotent stem cells (iPSCs) and master transcription factor (TF)-mediated transdifferentiation have transformed cell engineering.
• The molecular mechanisms underlying diverse cellular identity changes during reprogramming are not fully understood.
• Constitutive active Smad3 overexpression enhances iPSC generation and other TF-mediated cell conversions, suggesting common underlying mechanisms.

Purpose of the Study:

• To investigate the common molecular mechanisms of master TF-mediated cell conversions.
• To identify novel genes involved in or hindering the reprogramming process.
• To provide a valuable resource for improving cell conversion efficiency in regenerative medicine.

Main Methods:

• CRISPR/Cas9-mediated genome-wide knockout screening was employed during reprogramming.
• A lentiviral gRNA library containing 90,000 gRNAs was utilized for the screening.
• Reprogramming efficiency and cellular identity changes were analyzed.

Main Results:

• Approximately 15 novel reprogramming roadblock genes were identified.
• Around 20 candidate genes essential for reprogramming but not ES cell self-renewal were discovered.
• The study demonstrated common mechanisms underlying different master TF-mediated cell conversions.

Conclusions:

• Master TF-mediated cell conversions share common underlying mechanisms.
• The identified genes provide insights into cellular identity alteration during reprogramming.
• This research offers a valuable resource for advancing regenerative medicine through enhanced cell conversion strategies.