Regular ArticleCloning of Genomic Loci and Chromosomal Localization of the Human PCTAIRE-1 and -3 Protein Kinase Genes
Abstract
Recent studies on the molecular mechanisms controlling the mammalian cell cycle have disclosed a large family of cdc2-related serine/threonine kinases. Among this gene family, the PCTAIRE protein kinases comprise a distinct subfamily of unknown cellular function. To analyze the genomic structure and chromosomal location of the PCTAIRE-1 and -3 genes, we isolated human cosmid clones for each gene by screening a human genomic library with murine PCTAIRE cDNA probes. Overlapping clones encompassing approximately 60 kb of genomic DNA were obtained for both PCTAIRE-1 and -3 . These clones were confirmed to encode authentic PCTAIRE genes by the detection of exon-intron structures and the coincidence of the nucleotide sequence of exons to that of the published human cDNAs. Using these cosmid clones as probes for FISH analyses, the chromosomal loci for PCTAIRE-1 and PCTAIRE-3 were assigned to bands Xp11 and 1q31-q32, respectively.
References (0)
Cited by (22)
Antioxidant activity of microwave-assisted extract of longan (Dimocarpus Longan Lour.) peel
2008, Food ChemistryThe longan (Dimocarpus Longan Lour.) peel was extracted with 95% ethanol employing microwave-assisted extraction and Soxhlet extraction method, the total phenolic content of microwave-assisted extract of Langan peel (MEL) and Soxhlet extract of Langan peel (SEL) reached 96.78 mg/g and 90.35 mg/g dry weight, respectively, expressed as pyrocatechol equivalents, which were quantified using Folin–Ciocalteu reagent. Subsequently, antioxidant properties of two extracts were investigated employing various established systems in vitro including 2,2′-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay, hydroxyl radical scavenging assay using a new resonance scattering (RS) method, reducing power and total antioxidant capacity. MEL and SEL showed excellent antioxidant in all test systems compared to synthetic antioxidant 2,6-di-ter-butyl-4-methylphenol (BHT) and the antioxidant activities of MEL were all superior to those of SEL. Furthermore, the suitability of MEL and SEL as substitute of BHT were determined in peanut oil, and the decrease of lipid oxidation were monitored using thiobarbituric acid-reactive substances (TBARS) assay. MEL and SEL treatment significantly (P < 0.05) reduced lipid oxidation in peanut oil compared to the control. No significant differences ( in lipid oxidation were detected between MEL, SEL and BHT samples of peanut oil.
Studies on antioxidant potential of methanol extract/fractions of Acacia auriculiformis A. Cunn
2007, Food ChemistryAntioxidant activities of the methanol extract/fractions of Acacia auriculiformis A. Cunn were evaluated by three in vitro experiments, namely, DPPH, relative reducing power and hydroxyl radical (site specific and non site specific) assays. The differential activities of methanol extract/fractions could be correlated with their respective total phenolic contents and compared with standards (BHT and l-ascorbic acid.). The bark powder of the plant was extracted with different solvents of increasing and decreasing polarity by a maceration extraction method and then the methanol extract was further partitioned with ethyl acetate and water. The scavenging activity of extract was found to be enhanced on fractionating the extract. Moreover, among the two fractions (ethyl acetate and water fraction) and the crude extract, the water fraction exhibited good scavenging responses of 72.0%, (57.2%), 1.76 (1.52), 88.0% (82.6%) and 93.6% (83.47) in DPPH, reducing power, site specific and non-site specific hydroxyl radical scavenging assay in increasing and (decreasing) order of solvent polarity at maximum concentration, respectively. Studies are in progress to evaluate the effect of extracts/fractions in other antioxidant assays and to identify the factors responsible for the activity.
The genomic repertoire for cell cycle control and DNA metabolism in S. purpuratus
2006, Developmental BiologyA search of the Strongylocentrotus purpuratus genome for genes associated with cell cycle control and DNA metabolism shows that the known repertoire of these genes is conserved in the sea urchin, although with fewer family members represented than in vertebrates, and with some cases of echinoderm-specific gene diversifications. For example, while homologues of the known cyclins are mostly encoded by single genes in S. purpuratus (unlike vertebrates, which have multiple isoforms), there are additional genes encoding novel cyclins of the B and K/L types. Almost all known cyclin-dependent kinases (CDKs) or CDK-like proteins have an orthologue in S. purpuratus; CDK3 is one exception, whereas CDK4 and 6 are represented by a single homologue, referred to as CDK4. While the complexity of the two families of mitotic kinases, Polo and Aurora, is close to that found in the nematode, the diversity of the NIMA-related kinases (NEK proteins) approaches that of vertebrates. Among the nine NEK proteins found in S. purpuratus, eight could be assigned orthologues in vertebrates, whereas the ninth is unique to sea urchins. Most known DNA replication, DNA repair and mitotic checkpoint genes are also present, as are homologues of the pRB (two) and p53 (one) tumor suppressors. Interestingly, the p21/p27 family of CDK inhibitors is represented by one homologue, whereas the INK4 and ARF families of tumor suppressors appear to be absent, suggesting that these evolved only in vertebrates. Our results suggest that, while the cell cycle control mechanisms known from other animals are generally conserved in sea urchin, parts of the machinery have diversified within the echinoderm lineage. The set of genes uncovered in this analysis of the S. purpuratus genome should enhance future research on cell cycle control and developmental regulation in this model.
PCTAIRE 3 is a member of the PCTAIRE subfamily of cdc2-related serine/threonine protein kinases. In the present study, cDNAs encoding two isoforms of PCTAIRE 3 have been cloned and the genomic organization of the human PCTAIRE 3 gene is reported. The gene spans 28.15 kb on chromosome 1q31–32 and contains 16 exons. The major transcript of PCTAIRE 3, designated PCTAIRE 3a, has an open reading frame that is 474 amino acids in length. Transcripts for PCTAIRE 3a were evident throughout the brain and in the majority of tissues analyzed. A second transcript containing an insert that adds 90 nucleotides to the third exon of the gene was also identified. This transcript, designated PCTAIRE 3b, encodes a polypeptide of 504 amino acids. Expression of PCTAIRE 3b was limited to several subcortical nuclei of the basal gangli and the spinal cord and substantial levels of this transcript were not evident outside of the central nervous system. Primary sequence comparisons between different cdc2-related serine/threonine protein kinases reveal that these proteins are most heterogeneous in their N-terminal domains and the PCTAIRE subfamily is further diversified by the presence of isoforms within this region.
Identification of an alternatively spliced form of the mouse AML1/RUNX1 gene transcript AML1c and its expression in early hematopoietic development
2001, Biochemical and Biophysical Research CommunicationsAcute myeloid leukemia 1 (AML1: or runt-related transcription factor, RUNX1) encodes the DNA binding subunit of the heterodimering transcription factor complex PEBP2 (CBF), which plays an essential role for definitive hematopoiesis. Transcription of AML1 is controlled by two distinct promoter regions, which results in the generation of the respective AML1b and AML1c isoforms. Here we report the isolation of the mouse homologue of human AML1c, whose unique N-terminus is 100% identical at the amino acid level to its human counterpart and 63 and 37% identical to the respective family members AML2 and AML3. Semiquantitative RT-PCR assay on mouse embryonic stem cell clones during in vitro differentiation and Northern blot analysis of a mouse embryo revealed that AML1b is expressed in undifferentiated ES cells and upregulated in the early developmental stage, in contrast to the gradual upregulation and steady maintenance of AML1c expression during embryogenesis. In addition, maintenance of AML1c expression depended on the presence of active AML1 allele(s) while that of AML1b did not. Thus, these two AML1 isoforms driven by their respective promoters are differentially expressed and are likely to have distinct functions in early hematopoietic development.
Multiple subcellular localizations of PCTAIRE-1 in brain
2000, Molecular and Cellular NeurosciencesWe developed a selective antibody to a synthetic peptide corresponding to an N-terminal sequence of the PCTAIRE-1 protein. In rodent brain extracts it recognized only the protein doublet characteristic of PCTAIRE-1, and this signal is completely abolished by preincubation of the antibody with the immunopeptide. Immunolabeling experiments done with this PCTAIRE-1-specific antibody reveal that the protein is widely distributed in the rodent brain as are the mRNAs visualized using an antisense riboprobe corresponding to the entire PCTAIRE-1 open reading frame. Two types of PCTAIRE-1 protein localizations were observed: first a diffuse labeling of almost all brain regions, particularly intense in the molecular layer of the cerebellum and the mossy fiber region of the hippocampus, and second a spot-like localization in the nuclei of large neurons such as cerebellar Purkinje cells and pyramidal cells of the hippocampus. Colocalization with the B23 protein allows one to identify these compartments as nucleoli. Our results suggest a nucleolar function of PCTAIRE-1 in large neurons and a role in regions containing important granule cell projections.