Thus, the S-phase checkpoint works to restrict origin activation once lagging strand PAF15-PCNA assembly is exhausted, revealing a previously unknown, strand-specific rate limitation that governs overall replication dynamics. Hope you will enjoy reading.

Finally, we show that that PAF15 rate-limitation is governed by the dynamic activity of repressive E2F4 transcription factor, which maintain a low PAF15-PCNA ratio to ensure genome protection.

In contrast to the lagging strand, where PCNA interacts with PAF15, PCNA on the leading strand fully engages with POLE. In fact, TIMELESS and CLASPIN—components of replisome that couple key replisome-based interactions—shield leading strand PCNA from PAF15, thereby promoting cell survival.

What happens if PAF15 exceeds physiological levels? Any misregulation—whether by overexpression or mislocalization to the leading strand—impairs PCNA turnover and rapidly replication fork progression and triggers a lethal replication response, ultimately leading to cell death.

What is the role of PAF15 during unperturbed DNA replication? With its high-affinity PIP motif and unique access within the DNA-encircling channel of PCNA, PAF15 stabilizes PCNA on chromatin, maintaining proper interactions between PCNA, FEN1, and LIGASE1 for effective OK fragment maturation.

Searching for the mechanisms underlying this strand-specific PCNA depletion, we identified PAF15 as a rate-limited component. PAF15 associates specifically with PCNA on the lagging strand and becomes fully exhausted upon normal origin firing, delineating the overall range of replication.

Under these conditions, cells resort to non‑canonical lagging strand maturation pathways, ultimately exhausting the genome-protective single-stranded binding protein RPA and resulting in terminal replication collapse.

Essentially, when origin activation exceeds a certain threshold (for instance by attenuation of the S-phase checkpoint), cells instantly pause de novo PCNA chromatin loading and lagging strand-specific activities.