APE1/Ref-1 interacts with NPM1 within nucleoli and plays a role in the rRNA quality control process.
APE1/Ref-1 (hereafter, APE1), a DNA repair enzyme and a transcriptional coactivator, is a vital protein in mammals. Its role in controlling cell growth and the molecular mechanisms that fine-tune its different cellular functions are still not known. By an unbiased proteomic approach, we have identified and characterized several novel APE1 partners which, unexpectedly, include a number of proteins involved in ribosome biogenesis and RNA processing. In particular, a novel interaction between nucleophosmin (NPM1) and APE1 was characterized. We observed that the 33 N-terminal residues of APE1 are required for stable interaction with the NPM1 oligomerization domain. As a consequence of the interaction with NPM1 and RNA, APE1 is localized within the nucleolus and this localization depends on cell cycle and active rRNA transcription. NPM1 stimulates APE1 endonuclease activity on abasic double-stranded DNA (dsDNA) but decreases APE1 endonuclease activity on abasic single-stranded RNA (ssRNA) by masking the N-terminal region of APE1 required for stable RNA binding. In APE1-knocked-down cells, pre-rRNA synthesis and rRNA processing were not affected but inability to remove 8-hydroxyguanine-containing rRNA upon oxidative stress, impaired translation, lower intracellular protein content, and decreased cell growth rate were found. Our data demonstrate that APE1 affects cell growth by directly acting on RNA quality control mechanisms, thus affecting gene expression through posttranscriptional mechanisms.
Vascotto
Carlo
C
Department of Biomedical Sciences and Technologies, University of Udine, P.le Kolbe 4, 33100 Udine, Italy.
Fantini
Damiano
D
Romanello
Milena
M
Cesaratto
Laura
L
Deganuto
Marta
M
Leonardi
Antonio
A
Radicella
J Pablo
JP
Kelley
Mark R
MR
D'Ambrosio
Chiara
C
Scaloni
Andrea
A
Quadrifoglio
Franco
F
Tell
Gianluca
G
D001667:Binding, Competitive; D002453:Cell Cycle; D002466:Cell Nucleolus / Q000378:metabolism*; D049109:Cell Proliferation; D004247:DNA / Q000378:metabolism; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase / Q000737:chemistry / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D006367:HeLa Cells; D006801:Humans; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D000090243:Nucleophosmin; D010084:Oxidation-Reduction; D010449:Peptide Mapping; D011485:Protein Binding; D014176:Protein Biosynthesis; D025941:Protein Interaction Mapping; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D012335:RNA, Ribosomal / Q000235:genetics / Q000378:metabolism*; D012337:RNA, Ribosomal, 18S / Q000378:metabolism; D012339:RNA, Ribosomal, 28S / Q000378:metabolism; D014158:Transcription, Genetic
C000716967:NPM1 protein, human; D009687:Nuclear Proteins; D012335:RNA, Ribosomal; D012337:RNA, Ribosomal, 18S; D012339:RNA, Ribosomal, 28S; D000090243:Nucleophosmin; D004247:DNA; C085108:APEX1 protein, human; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase