Exonic splicing code and protein binding sites for calcium

Area of Science:

• Molecular Biology
• Evolutionary Biology
• Genetics

Background:

• Exonic splicing enhancers (ESEs) and silencers (ESSs) are crucial for RNA splicing, influenced by protein binding.
• Previous studies linked ESE/ESS evolution to metal binding, specifically the Irving-Williams (I-W) series.
• The role of specific metal ions, like Ca2+, in shaping splicing codes is not well understood.

Purpose of the Study:

• To investigate the splicing activities of exonic sequences encoding Ca2+-binding sites.
• To determine if Ca2+ coordination residues influence exon inclusion levels.
• To explore the evolutionary implications of Ca2+-binding sites in RNA splicing.

Main Methods:

• Analysis of predicted exon inclusion levels for EF-hand motifs and other Ca2+-binding residues.
• Comparison of splicing patterns between canonical and noncanonical EF-hands.
• Utilizing splicing assays and crosslinking/immunoprecipitation data.
• Identification of trans-acting factors binding to specific motifs.

Main Results:

• Exonic sequences with Ca2+-binding sites, particularly EF-hand motifs, exhibit higher predicted exon inclusion levels.
• A specific hierarchy (1>12>3∼5>9) was observed for Ca2+-coordinating residues in canonical EF-hands.
• Exon splits were preferentially located in the N-terminal halves and around codon 12 of EF-hand loops.
• Candidate trans-acting factors binding GA-rich motifs encoding negatively charged amino acids were identified.

Conclusions:

• Codons for Ca2+-coordinating residues have a high capacity for retention in mature transcripts.
• This retention facilitates exon-level expansion during eukaryotic evolution.
• Ca2+ binding sites play a significant role in the evolution of RNA splicing codes.