The silent information regulator 2 (Sir2) family of NAD-dependent N-acetyl-protein deacetylases participates in the regulation of gene silencing, chromatin structure, and longevity. In the Sir2-catalyzed reaction, the acetyl moiety of N-acetyl-histone is transferred to the ADP-ribose of NAD, yielding O-acetyl-ADP-ribose and nicotinamide. We hypothesized that, if O-acetyl-ADP-ribose were an important signaling molecule, a specific hydrolase would cleave the (O-acetyl)-(ADP-ribose) linkage. We report here that the poly-(ADP-ribose) glycohydrolase ARH3 hydrolyzed O-acetyl-ADPribose to produce ADP-ribose in a time-and Mg 2؉ -dependent reaction and thus could participate in two signaling pathways. This O-acetyl-ADP-ribose hydrolase belongs to a family of three structurally related 39-kDa ADP-ribose-binding proteins (ARH1-ARH3). ARH1 was reported to hydrolyze ADP-ribosylarginine, whereas ARH3 degraded poly(ADP-ribose). ARH3-catalyzed generation of ADP-ribose from O-acetyl-ADP-ribose was significantly faster than from poly(ADP-ribose). Like the degradation of poly(ADP-ribose) by ARH3, hydrolysis of O-acetyl-ADP-ribose was abolished by replacement of the vicinal aspartates at positions 77 and 78 of ARH3 with asparagine. The rate of O-acetyl-ADP-ribose hydrolysis by recombinant ARH3 was 250-fold that observed with ARH1; ARH2 and poly(ADP-ribose) glycohydrolase were inactive. All data support the conclusion that the Sir2 reaction product O-acetyl-ADP-ribose is degraded by ARH3.ADP-ribosylhydrolase ͉ ADP-ribosyltransferase ͉ sirtuin ͉ ADP-ribose S ilent information regulator 2 (Sir2) family proteins are involved in gene silencing, chromosomal stability, and lifespan extension (1, 2). In the presence of NAD, Sir2 couples protein deacetylation with formation of O-acetyl-ADP-ribose and release of nicotinamide (3-5). NAD-dependent histone deacetylation appears to be crucial for the biological effects of Sir2. The second product of the reaction, O-acetyl-ADP-ribose, may be involved in the stabilization of chromatin and formation of Sir complexes (6, 7), although its contribution to the biological effects of Sir2 is unclear.In many biological systems, specific enzymes are responsible for the degradation of small molecules that are generated in signaling cascades, thus terminating their action. Examples include the adenylyl cyclase and guanylyl cyclase pathways, where cyclic nucleotide phosphodiesterases degrade cAMP and cGMP, respectively, thus extinguishing the signal (8). Thus far, enzymatic destruction of O-acetyl-ADP-ribose has been shown only with the Nudix family of ADP-ribose pyrophosphatases (9) (nucleoside diphosphate linked to another moiety, hence the acronym Nudix) (10) and perhaps other less selective pyrophosphatases.The extent of ADP-ribosylation of proteins is determined by the rate of opposing actions of ADP-ribosyltransferases, which catalyze the posttranslational modification, and ADP-riboseprotein hydrolases, which cleave the ADP-ribose-protein linkage, releasing ADP-ribose and regenerating unmodified protein...

Background: Nuclear and cytosolic poly(ADP-ribose) metabolism is established but debated in mitochondria. Results: Novel mitochondrial and cytosolic poly(ADP-ribose) glycohydrolase splice variants are inactive for poly(ADP-ribose) degradation. Conclusion: Degradation of mitochondrial matrix-accumulated poly(ADP-ribose) can be catalyzed only by ADP-ribosylhydrolase 3, whereas small poly(ADP-ribose) glycohydrolase isoforms may have functions different from poly(ADP-ribose) degradation. Significance: Important insights into the regulation of subcellular poly(ADP-ribose) metabolism are provided.

Stathmin is an intracellular phosphoprotein that is overexpressed in a number of human malignancies. Our previous study using proteomic profiling showed that significant upregulation of stathmin occurs in oral squamous-cell carcinoma (OSCC)-derived cell lines. In the current study, to determine the potential involvement of stathmin in OSCC, we evaluated the state of stathmin protein and mRNA expression in OSCC-derived cell lines and human primary OSCCs. A significant increase in stathmin expression was observed in all OSCC-derived cell lines examined compared to human normal oral keratinocytes. In immunohistochemistry, 65% of the OSCCs were positive for stathmin, and no immunoreaction was observed in corresponding normal tissues. Real-time quantitative reverse transcriptase -polymerase chain reaction data were consistent with the protein expression status. Moreover, stathmin expression status was correlated with the TNM stage grading. Furthermore, we found a statistical correlation between the protein expression status and disease-free survival (P ¼ 0.029). These results suggest that expression of stathmin could contribute to cancer progression/prognosis, and that stathmin may have potential as a biomarker and a therapeutic target for OSCC.