The Prolyl Isomerase Pin1 Targets Stem-Loop Binding Protein (SLBP) To Dissociate the SLBP-Histone mRNA Complex Linking Histone mRNA Decay with SLBP Ubiquitination

Abstract

Histone mRNAs are rapidly degraded at the end of S phase, and a 26-nucleotide stem-loop in the 3′ untranslated region is a key determinant of histone mRNA stability. This sequence is the binding site for stem-loop binding protein (SLBP), which helps to recruit components of the RNA degradation machinery to the histone mRNA 3′ end. SLBP is the only protein whose expression is cell cycle regulated during S phase and whose degradation is temporally correlated with histone mRNA degradation. Here we report that chemical inhibition of the prolyl isomerase Pin1 or downregulation of Pin1 by small interfering RNA (siRNA) increases the mRNA stability of all five core histone mRNAs and the stability of SLBP. Pin1 regulates SLBP polyubiquitination via the Ser20/Ser23 phosphodegron in the N terminus. siRNA knockdown of Pin1 results in accumulation of SLBP in the nucleus. We show that Pin1 can act along with protein phosphatase 2A (PP2A) in vitro to dephosphorylate a phosphothreonine in a conserved TPNK sequence in the SLBP RNA binding domain, thereby dissociating SLBP from the histone mRNA hairpin. Our data suggest that Pin1 and PP2A act to coordinate the degradation of SLBP by the ubiquitin proteasome system and the exosome-mediated degradation of the histone mRNA by regulating complex dissociation.

SUPPLEMENTAL MATERIAL

Supplemental material for this article may be found at http://dx.doi.org/10.1128/MCB.00382-12.

ACKNOWLEDGMENTS

These studies were supported by NIH grant 1RO1-GM076660 to William F. Marzluff and Roopa Thapar and HWI faculty start-up funds to Roopa Thapar. Ronald Berezney and Andrew Fritz were supported by NIH grant 1RO1-GM072131.

The flow cytometry, ImageStream flow cytometry, and confocal microscopy were performed at RPCI's Flow and Image Cytometry Facility, the CCD microscopy and deconvolution image analysis were performed at the University at Buffalo, and the RT-PCR studies were performed at Hauptman-Woodward Institute and the UB Center of Excellence in Bioinformatics and Life Sciences (COE). The NMR studies were performed at the Biomolecular NMR Facility at the University of North Carolina at Chapel Hill. All other studies were performed at HWI. We thank Anthony Means (Duke University) for bacterial expression vectors for human and Xenopus Pin1 and Dirk Bohmann (University of Rochester) for mammalian expression vectors of HA and (His)₆-tagged ubiquitin. We also appreciate Mary LoPresti and Jean Kanyo (W. M. Keck Foundation Biotechnology Resource Laboratory, Yale University) for assistance with sample preparation and collecting some of the mass spectrometry data on the Orbitrap Elite (1S10 RR031795-0).

We declare that we have no financial or other conflicts of interest associated with this work.