Sequence and structural analyses of nuclear export signals in the NESdb database.

Area of Science:

• Molecular Biology
• Structural Biology
• Bioinformatics

Background:

• Nuclear export signals (NESs) mediate the transport of CRM1 cargoes from the nucleus to the cytoplasm.
• Accurate identification of NESs is critical for understanding cellular transport mechanisms.
• Existing methods for NES identification may lead to false positives, complicating research.

Purpose of the Study:

• To compile a comprehensive database of NES-containing CRM1 cargoes (NESdb).
• To analyze sequence and structural properties differentiating experimentally validated NESs from false positives.
• To develop a more precise NES consensus and improve prediction strategies.

Main Methods:

• Compilation of >200 NES-containing CRM1 cargoes into the NESdb database.
• Analysis of amino acid frequencies, sequence logos, and 3D structures of identified NESs and false positives.
• Experimental testing of CRM1-binding for selected NES peptides.

Main Results:

• Experimentally identified NESs exhibit distinct sequence patterns (e.g., Φ1-X(3)-Φ2-X(2)-Φ3-X-Φ4) and preferences for charged/hydrophobic amino acids compared to false positives.
• Real NESs are more likely to adopt specific α-helical conformations and be surface-accessible or disordered in 3D structures.
• 16 out of 40 tested NES peptides failed to bind CRM1, highlighting the issue of misidentification.

Conclusions:

• Sequence and structural analyses provide distinguishing features for experimentally validated NESs.
• A refined NES consensus sequence and understanding of structural preferences can enhance prediction accuracy.
• The findings underscore the potential for misidentifying NESs and emphasize the need for rigorous validation.