A novel histone deacetylase, HDAC10, was isolated from a mixed tissue human cDNA library. HDAC10 was classified as a class II subfamily member based upon similarity to HDAC6. The genomic structure of HDAC10 was found to consist of 20 exons. HDAC10 has two sequence variants, HDAC10v1 and HDAC10v2, and two transcripts were detectable by Northern blot analysis. HDAC10v1 and HDAC10v2 were found to be identical through exon 17 but diverged after this exon. HDAC10v2 has an 82-bp alternate exon that generates a frameshift and shortens the sequence by 11 amino acids. In this study, the characterization of HDAC10v1 was performed. HDAC10v1 has an N-terminal catalytic domain, two putative C-terminal retinoblastoma protein binding domains, and a nuclear hormone receptor binding motif. The HDAC10v1 enzyme was found to be catalytically active based upon its ability to deacetylate a 3 H-acetylated histone H4 N-terminal peptide. Immunofluorescence detection of transfected HDAC10v1-FLAG indicated that the enzyme is a nuclear protein. Furthermore, coimmunoprecipitation experiments indicated that HDAC10v1 associated with HDAC2 and SMRT (silencing mediator for retinoid and thyroid hormone receptors). In addition, based upon the public data base, a single nucleotide polymorphism was found in the C terminus of HDAC10 which changes a Gly residue to Cys, suggesting that HDAC10 molecules containing these single nucleotide polymorphisms may be folded improperly. HDAC10 extends the HDAC superfamily and adds to a growing number of HDACs that have been found to have splice variants, suggesting that RNA processing may play a role in mediating the activity of HDACs.Chromatin remodeling plays a major regulatory role in transcription and DNA replication (1, 2). One model for how chromatin remodeling occurs involves ATP-dependent displacement of histones by nucleosome remodeling complexes (3, 4) and changes in the acetylation status of histones and transcription factors catalyzed by histone acetylases and histone deacetylases (HDACs) 1 (5-7).There are currently 16 reported human HDAC isoforms (8 -11) that are divided into three classes based upon sequence homology, intracellular localization, and association with proteins that form DNA-binding complexes. HDAC1, HDAC2, HDAC3, and HDAC8 were categorized as class I based upon their similarity to the yeast gene Rpd3 (8). HDAC 4/HDACA, HDAC5/HDACB, HDAC6, HDAC7, and HDAC9 were designated as class II, based upon their similarity to yeast gene Hda1 (9, 10). The third class of HDACs consists of seven human genes that are similar to yeast silent information regulator gene (Sir2) (13,14). A unique characteristic of class III HDACs is their NAD ϩ -dependent protein deacetylase and ADP-ribosylase activity (15-17).HDACs have been found in multiprotein complexes, implicating HDACs in transcription regulation, hormone signaling, cell cycle, differentiation, and DNA repair. Class I and class II HDACs were found to be components of different complexes (8 -10). HDAC1 and 2 formed a core complex with retinoblastom...

Using a bioinformatics approach, we have isolated a novel Gprotein-coupled receptor (GPCR), R527, and have demonstrated that this receptor shows no significant homology to previously deorphanized GPCRs. Quantitative reverse transcription-polymerase chain reaction analysis of the expression of GPCR R527 indicated a very high level of mRNA expression in eosinophils, with high expression also detected in neutrophils and lung macrophages. Stable cell lines were generated expressing this receptor together with the G-protein ␣-subunit G␣ 16 . These cells were used to screen an agonist collection in a calcium mobilization assay and 5-oxo-6E,8Z,11Z,14Z-eicosatetraenoic acid (5-oxo-ETE) was identified as a putative ligand. 5(S)-Hydroxyperoxy-6E,8Z,11Z,14Z-eicosatetraenoic acid was also shown to activate the receptor, whereas the leukotrienes LTB 4 , LTC 4 , LTD 4 , and LTE 4 failed to elicit a response. In cAMP assays, pertussis toxin reversed the inhibitory effects of 5-oxo-ETE on cAMP production, indicating that the receptor is G␣ i -coupled. The GPCR R527 shows pharmacological properties similar to those of the previously described 5-oxo-ETE receptor expressed on eosinophils, neutrophils, and monocytes. These cell types show chemotactic responses to 5-oxo-ETE, and this eicosanoid has been proposed to play a key role in the inflammatory response. The molecular identification of a receptor binding 5-oxo-ETE will expand our understanding of the physiological role of this mediator and may provide new therapeutic opportunities.

N-terminal methionine removal is an important cellular process required for proper biological activity, subcellular localization, and eventual degradation of many proteins. The enzymes that catalyze this reaction are called Methionine Aminopeptidases (MAPs). To date, only two MAP family members, MAP1A and MAP2, have been well characterized and studied in mammals. In our studies, we have cloned a full length MAP1D gene. Expression and purification of full length recombinant protein shows that the sequence encodes an enzyme with MAP activity. MAP1D is overexpressed in colon cancer cell lines and in colon tumors as compared to matched normal tissue samples. Downregulation of MAP1D expression by shRNA in HCT-116 colon carcinoma cells reduces anchorage-independant growth in soft agar. These data suggest that MAP1D is a potentially oncogenic, novel member of the MAP gene family that may play an important role in colon tumorigenesis.