Microcell-mediated chromosome transfer between non-identical human iPSCs

Area of Science:

• Stem cell biology
• Genetics
• Developmental biology

Background:

• Microcell-mediated chromosome transfer (MMCT) is a key technique for generating aneuploidy models.
• Existing methods using mouse/hamster cell lines are prone to chromosomal rearrangements and are labor-intensive.
• Human induced pluripotent stem cells (hiPSCs) offer a promising alternative for chromosome transfer and disease modeling.

Purpose of the Study:

• To develop an efficient MMCT method utilizing hiPSCs as both donor and recipient cells.
• To establish a robust system for generating hiPSC-based models of hyperaneuploidy syndromes.
• To overcome limitations of traditional MMCT methods regarding chromosomal stability and generation time.

Main Methods:

• Developed an MMCT protocol using hiPSCs, employing paclitaxel and reversine for micronucleation.
• Utilized ecotropic viral envelope and mCAT-1 interactions for efficient membrane fusion during MMCT.
• Employed CRISPR-Cas9 for tagging specific chromosomes (Chr21, ChrY) and transferring human/mouse artificial chromosomes (Chr21, ChrX, ChrY).

Main Results:

• Successfully established an MMCT method using hiPSCs as donor and recipient cells.
• Generated isogenic hiPSC disease models with controlled hyperaneuploidy.
• Achieved full-length introduction of artificial chromosomes, a feat not previously reported.

Conclusions:

• The developed CRISPR-Cas9 and MMCT strategy provides a systematic approach to create hiPSC disease models with hyperaneuploidy.
• This method facilitates the study of rare genetic disorders and offers insights into early developmental mechanisms.
• Enables the introduction of comprehensive chromosomal sets into hiPSCs for advanced research.