DDB1 and CUL4 associated factor 11 (DCAF11) mediates degradation of Stem-loop binding protein at the end of S phase

Abstract

In eukaryotes, bulk histone expression occurs in the S phase of the cell cycle. This highly conserved system is crucial for genomic stability and proper gene expression. In metazoans, Stem-loop binding protein (SLBP), which binds to 3' ends of canonical histone mRNAs, is a key factor in histone biosynthesis. SLBP is mainly expressed in S phase and this is a major mechanism to limit bulk histone production to the S phase. At the end of S phase, SLBP is rapidly degraded by proteasome, depending on two phosphorylations on Thr 60 and Thr 61. Previously, we showed that SLBP fragment (aa 51-108) fused to GST, is sufficient to mimic the late S phase (S/G2) degradation of SLBP. Here, using this fusion protein as bait, we performed pull-down experiments and found that DCAF11, which is a substrate receptor of CRL4 complexes, binds to the phosphorylated SLBP fragment. We further confirmed the interaction of full-length SLBP with DCAF11 and Cul4A by co-immunoprecipitation experiments. We also showed that DCAF11 cannot bind to the Thr61/Ala mutant SLBP, which is not degraded at the end of S phase. Using ectopic expression and siRNA experiments, we demonstrated that SLBP expression is inversely correlated with DCAF11 levels, consistent with the model that DCAF11 mediates SLBP degradation. Finally, we found that ectopic expression of the S/G2 stable mutant SLBP (Thr61/Ala) is significantly more toxic to the cells, in comparison to wild type SLBP. Overall, we concluded that CRL4-DCAF11 mediates the degradation of SLBP at the end of S phase and this degradation is essential for the viability of cells.

Cyclin F-Mediated Degradation of SLBP Limits H2A.X Accumulation and Apoptosis upon Genotoxic Stress in G2

“…This modification is not associated with degradation, despite a conflicting report by Djakbarova et al, 2016. Instead, multi-monoubiquitylation of SLBP via CRL4 WDR23 is necessary for SLBP’s role in the processing of canonical histone mRNAs (Brodersen et al, 2016).…”

Birth and Death of Histone mRNAs

“…In vertebrates SLBP is rapidly degraded at the end of S-phase, as a result of cyclin A/Cdk2 phosphorylating two threonines in the SFTTP motif. The ubiquitin ligase responsible for SLBP degradation at the end of S-phase is not clear, and both cyclin F [69] and DCAF11 [70] have been implicated in its degradation.…”

Guard the guardian: A CRL4 ligase stands watch over histone production

“…Notably, a recent report suggested that CRL4 WDR23 may also target SLBP for proteasomal degradation. 11 Our results, however, demonstrate that CRL4 WDR23 directly governs SLBP function rather than protein levels. First, there is no evidence that cells lacking WDR23 stockpile excess SLBP.…”

“…10 Further, the authors demonstrate that SLBP levels increase or decline depending on the siRNAmediated depletion or transient overexpression of WDR23. 11 However, given that SLBP levels fluctuate throughout the cell cycle, cell synchronization experiments in combination with inducible cell lines and siRNA resistant constructs are necessary to clarify whether WDR23 may contribute to SLBP degradation. Considering our own results and the mechanism described in Dankert et al, we currently advocate a model of sequential interaction and ubiquitination of SLBP by CRL4 WDR23 and SCF cyclin F .…”