A non-catalytic N-terminus domain of WRN prevents mitotic telomere deprotection

Abstract

Telomeric ends form a loop structure (T-loop) necessary for the repression of ATM kinase activation throughout the normal cell cycle. However, cells undergoing a prolonged mitotic arrest are prone to lose the T-loop, resulting in Aurora B kinase-dependent mitotic telomere deprotection, which was proposed as an anti-tumor mechanism that eliminates precancerous cells from the population. The mechanism of mitotic telomere deprotection has not been elucidated. Here, we show that WRN, a RECQ helicase family member, can suppress mitotic telomere deprotection independently of its exonuclease and helicase activities. Truncation of WRN revealed that N-terminus amino acids 168–333, a region that contains a coiled-coil motif, is sufficient to suppress mitotic telomere deprotection without affecting both mitotic Aurora B-dependent spindle checkpoint and ATM kinase activity. The suppressive activity of the WRN168–333 fragment is diminished in cells partially depleted of TRF2, while WRN is required for complete suppression of mitotic telomere deprotection by TRF2 overexpression. Finally, we found that phosphomimetic but not alanine mutations of putative Aurora B target sites in the WRN168–333 fragment abolished its suppressive effect. Our findings reveal a non-enzymatic function of WRN, which may be regulated by phosphorylation in cells undergoing mitotic arrest. We propose that WRN enhances the protective function of TRF2 to counteract the hypothetical pathway that resolves the mitotic T-loop.

A CPC-shelterin-BTR axis regulates mitotic telomere deprotection

“…IMR90 fibroblasts were provided by Fuyuki Ishikawa (Kyoto University), HeLa cells by Megan Chircop (CMRI), HT1080 6TG cells by Eric Stanbridge (University of California, Irvine), and TRF2 Floxed/Floxed Rosa26-CreERT2 pBabeSV40LT MEFs by Eros Lazzerini Denchi (National Cancer Institute) 13 . IMR90 E6E7 hTERT fibroblasts produced as described previously 65 . IMR90 E6E7 hTERT fibroblasts were grown in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (S1810-500, biowest), 200 mM L-glutamine, 7.5% NaHCO 3 , 100 U mL − 1 penicillin, streptomycin, and 5 μg mL − 1 Plasmocin (InvivoGen) and maintained at 37 °C in 5% CO 2 and 3% O 2 .…”