Modifying splice site usage with ModCon: Maintaining the genetic code while changing the underlying mRNP code.

Area of Science:

• Molecular Biology
• Genetics
• Bioinformatics

Background:

• Codon degeneracy allows an
• mRNP code
• layer influencing RNA processing alongside the genetic code.
• Splice site selection in pre-mRNA splicing depends on intrinsic strength and regulatory protein binding sites within the sequence context.

Purpose of the Study:

• To investigate modifications of splicing regulatory properties near potential splice sites without altering encoded amino acids.
• To quantify splicing regulatory properties using the Splice Site HEXplorer Weight (SSHW) based on the HEXplorer score algorithm.

Main Methods:

• Designed the ModCon algorithm, a stochastic optimization method using a genetic algorithm with crossover, insertion, mutation, and a sliding window approach.
• Applied ModCon to 1000 human splice donor sites to systematically minimize or maximize SSHW.
• Utilized a splice site competition reporter assay with coding sequences from FANCA, FANCB, and BRCA2 to test ModCon's efficacy.

Main Results:

• Achieved substantial and accurate modifications in SSHW for human splice donors without altering amino acid sequences.
• Successfully switched splice donor usage in reporter constructs, demonstrating functional control over splicing.
• Validated the ModCon algorithm's capability to precisely manipulate splicing regulatory properties.

Conclusions:

• The ModCon algorithm and its R package offer a powerful tool for reporter design, enabling the introduction or silencing of splice sites.
• This approach allows for general modification of the mRNP code while preserving the genetic code.
• Facilitates precise control over RNA processing and gene expression through targeted splicing modulation.