Single-cell multi-omic atlas and morphogen screening informs midbrain and hindbrain organoid engineering.

Area of Science:

• Neuroscience
• Developmental Biology
• Genomics

Background:

• Neural tube patterning is crucial for developing midbrain and hindbrain structures essential for motor control, sensory processing, and cognition.
• Dysfunction in these brain regions is linked to various neurological disorders.
• In vitro organoid models offer a promising avenue for studying neural development and modeling diseases.

Purpose of the Study:

• To comprehensively map cell composition and regulatory mechanisms in midbrain and hindbrain organoid models.
• To identify key transcription factors and morphogen concentrations that guide neuronal differentiation and regional patterning.
• To advance the utility of brain organoids for understanding posterior brain development and disease.

Main Methods:

• Paired single-cell transcriptome and accessible chromatin sequencing (scATAC-seq) were employed to analyze organoid cell types and regulatory elements.
• Gene regulatory network inference and transcription factor perturbation experiments were conducted to understand differentiation mechanisms.
• A single-cell multiplexed patterning screen was performed to identify optimal morphogen concentrations for specific cell type generation.

Main Results:

• Existing midbrain organoid protocols successfully generate diverse ventral and dorsal cell types, including those from the floor plate, midbrain, and adjacent hindbrain regions.
• The study resolved key mechanisms governing neuronal differentiation through gene regulatory network analysis.
• The patterning screen identified specific morphogen conditions capable of generating medulla glycinergic neurons and cerebellum glutamatergic subtypes, expanding the organoid model's potential.

Conclusions:

• The multi-omic atlas provides a detailed resource for understanding cellular and regulatory landscapes in posterior brain organoids.
• Morphogen-regulon relationships were elucidated, demonstrating their role in guiding region-specific progenitor differentiation.
• These findings enhance the capacity of brain organoids to model posterior brain development and neurological conditions.