Molecular mechanisms for the regulation of histone mRNA stem-loop-binding protein by phosphorylation

Area of Science:

• Molecular Biology
• Biochemistry
• Structural Biology

Background:

• Replication-dependent histone mRNAs feature a conserved stem-loop structure.
• Stem-loop-binding protein (SLBP) recognizes and binds this RNA structure.
• SLBP undergoes phosphorylation, including an internal threonine and multiple serines in the C-terminal tail of Drosophila SLBP (dSLBP).

Purpose of the Study:

• To investigate the impact of SLBP phosphorylation on RNA-binding affinity.
• To elucidate the structural and biophysical mechanisms underlying enhanced RNA binding upon phosphorylation.
• To understand the role of the C-terminal tail phosphorylation in SLBP's RNA association.

Main Methods:

• Structural analysis (including crystal structure of human SLBP).
• Biophysical analyses of Drosophila SLBP (dSLBP).
• RNA-binding assays to determine affinity changes.

Main Results:

• Phosphorylation of dSLBP significantly increases its RNA-binding affinity.
• Structural and biophysical data reveal how phosphorylation enhances binding.
• Phosphorylation of the C-terminal tail, while not directly contacting RNA, promotes RNA-binding competent conformations.
• Increased negative charge in the tail balances positive charges, leading to a more compact structure without RNA.

Conclusions:

• Phosphorylation is a key regulatory mechanism for SLBP's RNA-binding activity.
• The C-terminal tail's phosphorylation allosterically enhances RNA binding by reducing the entropic penalty.
• Understanding these mechanisms provides insights into gene regulation and protein-RNA interactions.