Background: Human Sp140 protein is a leukocyte-specific member of the speckled protein (Sp) family (Sp100, Sp110, Sp140, Sp140L), a class of multi-domain nuclear proteins involved in intrinsic immunity and transcriptional regulation. Sp140 regulates macrophage transcriptional program and is implicated in several haematologic malignancies. Little is known about Sp140 structural domains and its post-translational modifications. Methods: We used mass spectrometry and biochemical experiments to investigate endogenous Sp140 SUMOylation in Burkitt's Lymphoma cells and Sp140 SUMOylation sites in HEK293T cells, FLAG-Sp140 transfected and His6-SUMO-1T95K infected. NMR spectroscopy and in vitro SUMOylation reactions were applied to investigate the role of Sp140 PHD finger in the SUMOylation of the adjacent BRD. Results: Endogenous Sp140 is a SUMO-1 target, whereby FLAG-Sp140 harbors at least 13 SUMOylation sites distributed along the protein sequence, including the BRD. NMR experiments prove direct binding of the SUMO E2 ligase Ubc9 and SUMO-1 to PHD-BRDSp140. In vitro SUMOylation reactions show that the PHDSp140 behaves as SUMO E3 ligase, assisting intramolecular SUMOylation of the adjacent BRD. Conclusions: Sp140 is multi-SUMOylated and its PHD finger works as versatile protein-protein interaction platform promoting intramolecular SUMOylation of the adjacent BRD. Thus, combinatorial association of Sp140 chromatin binding domains generates a multifaceted interaction scaffold, whose function goes beyond the canonical histone recognition. General significance: The addition of Sp140 to the increasing lists of multi-SUMOylated proteins opens new perspectives for molecular studies on Sp140 transcriptional activity, where SUMOylation could represent a regulatory route and a docking surface for the recruitment and assembly of leukocyte-specific transcription regulators.
Sp140 is a multi-SUMO-1 target and its PHD finger promotes SUMOylation of the adjacent Bromodomain / C. Zucchelli, S. Tamburri, G. Filosa, M. Ghitti, G. Quilici, A. Bachi, G. Musco. - In: BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS. - ISSN 0304-4165. - 1863:2(2019 Feb), pp. 456-465.
|Titolo:||Sp140 is a multi-SUMO-1 target and its PHD finger promotes SUMOylation of the adjacent Bromodomain|
|Parole Chiave:||Bromodomain; NMR; PHD; Proteomics; Sp140; SUMOylation; Antigens, Nuclear; HEK293 Cells; Humans; Protein Binding; Transcription Factors; PHD Zinc Fingers; Protein Domains; Sumoylation|
|Settore Scientifico Disciplinare:||Settore BIO/11 - Biologia Molecolare|
|Data di pubblicazione:||feb-2019|
|Digital Object Identifier (DOI):||http://dx.doi.org/10.1016/j.bbagen.2018.11.011|
|Appare nelle tipologie:||01 - Articolo su periodico|