The integrated stress response induces R-loops and hinders replication fork progression

Area of Science:

• Cellular Biology
• Molecular Biology
• Genetics

Background:

• The integrated stress response (ISR) is a cellular defense mechanism against various stresses like protein misfolding or viral infections.
• ISR activation leads to the phosphorylation of eIF2alpha, a key factor in translation initiation, causing a shutdown of protein synthesis.

Purpose of the Study:

• To investigate the immediate effects of ISR activation on DNA replication.
• To elucidate the molecular mechanisms by which ISR impacts DNA synthesis.

Main Methods:

• Induction of ISR in cellular models.
• Analysis of DNA replication fork progression using established techniques.
• Assessment of histone synthesis and R-loop formation.
• Pharmacological manipulation of eIF2alpha kinases and R-loop resolving enzymes.

Main Results:

• ISR activation rapidly inhibits DNA replication fork progression within one hour.
• This inhibition is mechanistically linked to a block in histone synthesis, promoting R-loop accumulation.
• Restoration of DNA replication was observed upon inhibition of eIF2alpha kinases, eIF2alpha reactivation, histone overexpression, or RNaseH1-mediated R-loop resolution.

Conclusions:

• The ISR rapidly stalls DNA synthesis through histone deficiency and subsequent R-loop formation.
• This shutdown mechanism is proposed to prevent potentially detrimental DNA replication during cellular stress.
• Targeting ISR-mediated pathways may offer therapeutic strategies for conditions involving cellular stress and DNA replication defects.