title: 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.
authors.affiliation: a Department of Genetics and Bioengineering , Fatih University , Istanbul , Turkey.|b Bionanotechnology Center , Fatih University , Istanbul , Turkey.
authors.forename: Umidahan
authors.initials: U
authors.lastname: Djakbarova
authors.affiliation: c Department of Biochemistry and Biophysics , University of North Carolina at Chapel Hill , Chapel Hill , NC , USA.|d Program in Molecular Biology and Biotechnology , University of North Carolina at Chapel Hill , Chapel Hill , NC , USA.
authors.forename: William F
authors.initials: WF
authors.lastname: Marzluff
authors.affiliation: a Department of Genetics and Bioengineering , Fatih University , Istanbul , Turkey.|b Bionanotechnology Center , Fatih University , Istanbul , Turkey.
authors.forename: M Murat
authors.initials: MM
authors.lastname: Köseoğlu
mesh_terms: D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000378:metabolism*; D016923:Cell Death; D044842:Cullin Proteins / Q000378:metabolism; D016195:G2 Phase; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D050505:Mutant Proteins / Q000378:metabolism; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding; D059748:Proteolysis; D034622:RNA Interference; D016196:S Phase; D039221:mRNA Cleavage and Polyadenylation Factors / Q000737:chemistry / Q000378:metabolism*
chemical_list: C114234:CUL4A protein, human; D002352:Carrier Proteins; D044842:Cullin Proteins; D050505:Mutant Proteins; D009687:Nuclear Proteins; C092663:SLBP protein, human; D039221:mRNA Cleavage and Polyadenylation Factors; D046988:Proteasome Endopeptidase Complex
keywords: CRL4; Cell cycle; Cul4A; DCAF11; S phase; SLBP; Wdr23; histone mRNA