Comprehensive Characterization of Alternative Splicing Dynamics During Mammalian Brain Development Reveals Extensive Isoform Diversity and Its Profound Effects on Protein Conformational Changes

Core Concepts

Alternative splicing has a much greater impact in remodeling the transcriptome profile from neural stem cells to neurons than previously thought, independently from changes in gene expression. This includes a progressive increase in exon inclusion resulting in radical protein conformational changes.

Abstract

The study aimed to comprehensively assess alternative splicing (AS) dynamics and its potential impact on protein structure during mammalian brain development.
Key highlights:

Combining short-read and long-read sequencing, the study reconstructed a novel transcriptome assembly of the developing mouse cortex, identifying nearly 50,000 new isoforms.
Analysis of AS events revealed a strong bias towards exon inclusion during the transition from neural stem cells (NSC) to neurogenic progenitors (NP) and neurons (N). Exons that were minor isoforms in NSC became the predominant isoforms in N.
Computational modeling of protein structures using AlphaFold2 showed that a remarkably high proportion (37%) of isoform pairs from the same gene resulted in substantially different 3D conformations, even when differing by as little as two amino acids.
The study also identified numerous instances of local secondary structure element switches, where identical protein sequences adopted profoundly different conformations depending on distant AS events.
These findings highlight the extensive potential of AS to redefine protein sequence and structure, and thus modulate protein function, during cell fate commitment in brain development.

Stats

Approximately 50,000 new isoforms were identified, accounting for nearly half of the total transcriptome diversity.
37% of isoform pairs from the same gene exhibited large global conformational changes, including fold switches.
Nearly 40% of isoforms showed regions with identical sequence yet adopting profoundly different secondary structures.

Quotes

"Alternative splicing alone was revealed to have a much greater impact in remodeling the transcriptome profile from NSC to N than previously thought and independently from changes in gene expression."
"Remarkably, this highlighted that nearly 40% of isoform pairs originating from the same gene exhibited large global conformational changes including fold switches."
"This example highlights how identical sequences can adopt different structural conformations as a result of an AS event occurring 10 amino acids away."

Key Insights Distilled From

Characterization of Alternative Splicing During Mammalian Brain Development Reveals the Magnitude of Isoform Diversity and its Effects on Protein Conformational Changes

by Haj Abdullah... at www.biorxiv.org 10-14-2023

https://www.biorxiv.org/content/10.1101/2023.10.11.561865v1

Deeper Inquiries

How do the observed protein conformational changes resulting from alternative splicing events impact the functional properties and interactions of the affected proteins during brain development?

The observed protein conformational changes resulting from alternative splicing events can have a significant impact on the functional properties and interactions of the affected proteins during brain development. These conformational changes can alter the structure of the protein, leading to changes in its stability, localization, and interactions with other molecules. For example, a small change in the amino acid sequence due to alternative splicing can result in a completely different protein structure, affecting its function. This can lead to changes in protein-protein interactions, enzymatic activity, and signaling pathways crucial for neuronal development and function.
Additionally, alternative splicing can introduce or remove specific protein domains or motifs that are essential for the protein's function. These changes can modulate the protein's activity, subcellular localization, and ability to interact with other proteins or molecules in the cell. Therefore, the protein conformational changes induced by alternative splicing events play a critical role in shaping the functional properties of proteins during brain development, ultimately influencing neuronal differentiation, maturation, and connectivity.

How might the potential limitations of the computational protein structure modeling approach used in this study be addressed in future work to further validate the findings?

The computational protein structure modeling approach used in this study, specifically AlphaFold2, has shown remarkable accuracy in predicting protein structures. However, there are potential limitations that need to be considered to ensure the validity of the findings.
One limitation is the accuracy of predicting protein structures with high disordered regions. To address this limitation, future work could focus on improving the algorithms and training data to better predict the structures of proteins with significant disordered regions. Additionally, experimental validation techniques such as X-ray crystallography, nuclear magnetic resonance (NMR) spectroscopy, or cryo-electron microscopy can be used to confirm the predicted structures and assess the impact of alternative splicing on protein conformation.
Another limitation is the need for experimental validation of the predicted structural changes induced by alternative splicing events. Future studies could incorporate biochemical assays, such as protein-protein interaction studies or functional assays, to validate the structural changes and their impact on protein function. Integrating multiple experimental approaches with computational modeling can enhance the robustness and reliability of the findings.

How might the extensive isoform diversity uncovered in this study be leveraged to develop new therapeutic strategies for neurodevelopmental disorders associated with aberrant alternative splicing?

The extensive isoform diversity uncovered in this study provides valuable insights into the complexity of gene expression regulation through alternative splicing in the context of neurodevelopment. This knowledge can be leveraged to develop new therapeutic strategies for neurodevelopmental disorders associated with aberrant alternative splicing.
One potential therapeutic approach is to target specific alternative splicing events that are dysregulated in neurodevelopmental disorders. By understanding the isoform diversity and its impact on protein function, researchers can design targeted therapies, such as small molecules or gene editing techniques, to modulate the splicing patterns and restore normal protein expression and function.
Furthermore, the identification of novel isoforms and splicing events associated with neurodevelopment provides potential targets for drug discovery and development. By targeting specific isoforms or splicing factors involved in neuronal differentiation, maturation, or synaptic function, novel therapeutic interventions can be designed to correct aberrant splicing patterns and restore normal brain development.
Overall, leveraging the extensive isoform diversity uncovered in this study can lead to the development of precision medicine approaches for neurodevelopmental disorders, offering new opportunities for personalized therapies tailored to the specific splicing profiles of affected individuals.