Ewing Sarcoma Related protein 1 recognizes R-loops by binding DNA forks

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• EWSR1 (Ewing Sarcoma Related protein 1) is a ubiquitous RNA-binding protein involved in RNA processing.
• EWSR1 plays a role in regulating R-loop formation; aberrant R-loops cause genomic and transcriptional stress.
• Loss of EWSR1 function is linked to increased R-loop abundance, particularly in Ewing Sarcoma.

Purpose of the Study:

• To elucidate the specific mechanism by which EWSR1 recognizes and binds to R-loop structures.
• To identify the molecular domains responsible for EWSR1's high-affinity binding and specificity to R-loops.

Main Methods:

• Electrophoretic mobility shift assays (EMSA) were employed to assess EWSR1 binding to R-loop-related substrates.
• Analysis focused on identifying the specific DNA regions within R-loops that EWSR1 interacts with.

Main Results:

• EWSR1 demonstrated high-affinity binding to substrates mimicking R-loop components.
• Specificity was mapped to the DNA fork region, the transition zone between double- and single-stranded DNA.
• The Zinc-finger (ZnF) and RNA recognition motif (RRM) domains, along with flanking RGG domains, were identified as critical for binding affinity and specificity.

Conclusions:

• EWSR1 utilizes its ZnF and RRM domains to specifically recognize and bind the DNA fork structures characteristic of R-loops.
• This specific binding mechanism provides a rationale for EWSR1's role in mitigating R-loop-associated genomic stress across various cellular contexts.