HDAC1 regulates the stability of glutamate carboxypeptidase II protein by modulating acetylation status of lysine 479 residue

다국어 초록 (Multilingual Abstract)

<P><B>Abstract</B></P> <P>Our previous study showed that the level of glutamate carboxypeptidase II (GCPII) protein is regulated by valproic acid, a histone deacetylase (HDAC) inhibitor, through acetylation of lysine residue in the GCPII protein in human astrocytes, U-87MG. The present study further investigated which HDAC subtype is involved in the acetylation of GCPII. The results revealed that GCPII interacted with HDAC1 but not with HDAC2, HDAC3, HDAC4, HDAC5, and HDAC6. Overexpression of catalytic domain (1–56 aa)-deleted HDAC1, which poorly binds to GCPII, enhanced lysine acetylation in GCPII and increased the level of GCPII protein when compared with that of the wild-type HDAC1. Further experiments showed that HDAC1 regulated the stability of GCPII protein. These data suggest that acetylation of GCPII is facilitated by HDAC1, and the acetylated GCPII is more stable than the non-acetylated GCPII. Additional experiments using siRNA HDAC1 and by <I>HDAC1</I> overexpression confirmed the role of HDAC1 in regulating the stability of GCPII protein. Further, database search of acetylation and ubiquitination sites showed four candidate lysine sites in human GCPII protein that can be both acetylated and ubiquitinylated (K207, K479, K491, and K699). Mutation (lysine residues to arginine (R)) analysis showed that in the presence of cycloheximide K479R- and K491R-hGCPII mutants were less ubiquitinylated and degraded, and decrease in the level of GCPII protein by HDAC1 was significantly blocked by K479R mutants. These data suggest that K479 is a possible site of acetylation or ubiquitination. Furthermore, the results also demonstrate that the stability of GCPII protein is regulated by HDAC1 through acetylation at the lysine 479 residue.</P> <P><B>Highlights</B></P> <P> <UL> <LI> GCPII interacts specifically with HDAC1 which is present in the non-nuclear fraction. </LI> <LI> HDAC1 regulates the stability of GCPII protein. </LI> <LI> Mutation at lysine sites in human GCPII (K479R and K491R) reduces the ubiquitination. </LI> <LI> Mutation at 479 lysine residue of human GCPII blocks HDAC1-mediated decrease in the level of GCPII protein. </LI> </UL> </P>