Unveiling novel interactions of histone chaperone Asf1 linked to TREX-2 factors Sus1 and Thp1.
Anti-silencing function 1 (Asf1) is a conserved key eukaryotic histone H3/H4 chaperone that participates in a variety of DNA and chromatin-related processes. These include the assembly and disassembly of histones H3 and H4 from chromatin during replication, transcription, and DNA repair. In addition, Asf1 is required for H3K56 acetylation activity dependent on histone acetyltransferase Rtt109. Thus, Asf1 impacts on many aspects of DNA metabolism. To gain insights into the functional links of Asf1 with other cellular machineries, we employed mass spectrometry coupled to tandem affinity purification (TAP) to investigate novel physical interactions of Asf1. Under different TAP-MS analysis conditions, we describe a new repertoire of Asf1 physical interactions and novel Asf1 post-translational modifications as ubiquitination, methylation and acetylation that open up new ways to regulate Asf1 functions. Asf1 co-purifies with several subunits of the TREX-2, SAGA complexes, and with nucleoporins Nup2, Nup60, and Nup57, which are all involved in transcription coupled to mRNA export in eukaryotes. Reciprocally, Thp1 and Sus1 interact with Asf1. Albeit mRNA export and GAL1 transcription are not affected in asf1Δ a strong genetic interaction exists between ASF1 and SUS1. Notably, supporting a functional link between Asf1 and TREX-2, both Sus1 and Thp1 affect the levels of Asf1-dependent histone H3K56 acetylation and histone H3 and H4 incorporation onto chromatin. Additionally, we provide evidence for a role of Asf1 in histone H2B ubiquitination. This work proposes a functional link between Asf1 and TREX-2 components in histone metabolism at the vicinity of the nuclear pore complex.
Departament de Bioquímica i Biologia Molecular; Universitat de València; Burjassot, Spain.
Mercè
M
Pamblanco
Gene Expression and RNA Metabolism Laboratory; Centro de Investigación Príncipe Felipe (CIPF); València, Spain.
Paula
P
Oliete-Calvo
Gene Expression and RNA Metabolism Laboratory; Centro de Investigación Príncipe Felipe (CIPF); València, Spain.
Encar
E
García-Oliver
Secció de Proteòmica; Servei Central de Suport a la Investigació Experimental (SCSIE); Universitat de València; Burjassot, Spain.
M
M
Luz Valero
Departament de Bioquímica i Biologia Molecular; Universitat de València; Burjassot, Spain.
Manuel M
MM
Sanchez del Pino
Gene Expression and RNA Metabolism Laboratory; Centro de Investigación Príncipe Felipe (CIPF); València, Spain.
Susana
S
Rodríguez-Navarro
D000107:Acetylation; D018797:Cell Cycle Proteins / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal / Q000235:genetics; D056488:Histone Chaperones / Q000378:metabolism*; D006657:Histones / Q000378:metabolism; D008745:Methylation; D018832:Molecular Chaperones / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D034443:RNA Transport / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D016601:RNA-Binding Proteins / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D014158:Transcription, Genetic / Q000235:genetics
C109731:ASF1 protein, S cerevisiae; D018797:Cell Cycle Proteins; D056488:Histone Chaperones; D006657:Histones; D018832:Molecular Chaperones; D009687:Nuclear Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; D029701:Saccharomyces cerevisiae Proteins; C481300:Sus1 protein, S cerevisiae; C476348:Thp1 protein, S cerevisiae
(5-10) yAsf1; SAGA; Sus1; TAP-MS strategy; TREX-2; histones; yeast