Phosphorylation-mediated conformational change regulates human SLFN11

Area of Science:

• Molecular Biology
• Structural Biology
• Biochemistry

Background:

• Human Schlafen 11 (SLFN11) is a protein that sensitizes cells to DNA damaging agents, acting as a predictive biomarker in chemotherapy.
• SLFN11 also functions as an antiviral restriction factor by recognizing single-stranded DNA (ssDNA) and inhibiting translation.
• The precise regulation of SLFN11's diverse functions and enzymatic activities has remained unclear.

Purpose of the Study:

• To elucidate the structural mechanisms underlying SLFN11 function and regulation.
• To investigate the role of phosphorylation in modulating SLFN11's interaction with tRNAs and ssDNA.
• To understand how SLFN11's nuclease activity is controlled.

Main Methods:

• Cryo-electron microscopy (cryo-EM) was used to determine the structures of SLFN11 bound to specific tRNAs.
• Site-directed mutagenesis was employed to create phosphomimetic mutants (e.g., S753D).
• Biochemical assays were performed to assess ssDNA binding, ATP binding, and ribonuclease activity.

Main Results:

• Cryo-EM structures reveal how SLFN11 binds and cleaves tRNA-Leu and tRNA-Met, with regulation by phosphorylation at S219 and T230.
• The S753D phosphomimetic mutant exhibits a monomeric conformation, binds ATP, but loses ssDNA binding and shows reduced ribonuclease activity.
• Phosphorylation at S753 acts as a conformational switch, governing SLFN11 dimerization and binding of ATP and ssDNA.

Conclusions:

• Phosphorylation at S753 is critical for regulating SLFN11 dimerization and substrate binding (ATP and ssDNA).
• Phosphorylation at S219 and T230 specifically modulates tRNA recognition and nuclease activity.
• These findings provide a structural basis for understanding the multifaceted roles of SLFN11 in DNA damage response and antiviral immunity.