The Alarmone Diadenosine Tetraphosphate as a Cosubstrate for Protein AMPylation

Area of Science:

• Biochemistry
• Molecular Biology
• Cellular Stress Response

Background:

• Diadenosine polyphosphates (Ap_n As) are alarmones that signal cellular stress.
• The precise functions of Ap_n As, particularly in protein modification, remain largely unknown.
• Protein AMPylation, mediated by FICD using ATP, is a known response to ER stress.

Purpose of the Study:

• To investigate the role of Ap_n As as cosubstrates for protein AMPylation.
• To identify new protein targets of AMPylation mediated by FICD.
• To explore how different nucleotide cosubstrates influence FICD-mediated AMPylation.

Main Methods:

• Utilized chemical proteomics with a novel chemical probe.
• Assessed the consumption of Ap₄ A by the AMPylator FICD.
• Identified protein targets of AMPylation.

Main Results:

• Ap₄ A was found to be efficiently utilized as a cosubstrate for AMPylation by FICD.
• New potential protein targets for AMPylation were identified.
• FICD-mediated AMPylation targets varied depending on the nucleotide cosubstrate used.

Conclusions:

• Ap₄ A can serve as a cosubstrate for FICD-mediated protein AMPylation.
• The identity of AMPylation targets can be modulated by the specific nucleotide cosubstrate.
• Distinct signaling pathways may operate at high versus low Ap₄ A concentrations, enabling stress-specific responses.