Biomedical subjects

Samuel Aparicio

Publications and source records attributed to Samuel Aparicio.

6 recordsLinked to original sources

PubMed2005

PEDF and the serpins: phylogeny, sequence conservation, and functional domains.

Pigment epithelium derived factor (PEDF) is non-inhibitory serpin with neurotrophic and antiangiogenic functions. In this study, we have assembled PEDF sequences for 9 additional species by data base mining and performed cross-species alignment for 14 PEDF sequences to identify conserved structural domains. We found evolutionary conservation of a leader sequence, a single C-terminal glycosylation site, collagen-binding residues, and four specific conserved PEDF peptides. The C-terminus, 384--415 and an N-terminal region 78--95, show close homology with many other serpins, and there is strong conservation of 39 of 51 consensus key residues involved in serpin structure and function. Two peptide regions, 40--67 and 277--301, are unique to PEDF but conserved in all species. Conserved residues at the N-terminus, helix d (hD), and helix A (hA) of PEDF form a structure similar to the heparin-binding groove of other serpins. We identified a motif in PEDF that is homologous to the nuclear localization signals of other proteins. A bitopographical localization of PEDF was confirmed by immunocytochemistry and Western blots. Our results suggest that secretion is required for PEDF's activity, that PEDF can migrate to the nucleus, and that PEDF has structural and functional features more common with inhibitory serpins.

Amino Acid Motifs↗

PubMed2004

p300 regulates p53-dependent apoptosis after DNA damage in colorectal cancer cells by modulation of PUMA/p21 levels.

Activation of the tumor suppressor p53 by DNA damage induces either cell cycle arrest or apoptosis, but what determines the choice between cytostasis and death is not clear. In this report, we show that the E1A-binding p300 nucleoprotein is a key determinant of p53-dependent cell fate in colorectal cancer cells: absence of p300 increases apoptosis in response to DNA damage. In addition, p300-deficient (p300(-)) cells fail to undergo G(1)/S arrest after UV irradiation. These abnormalities are associated with prolongation of p53 stability, reduced p53-acetylation, blunting of MDM2 activation, failure to transactivate p21, and a disproportionate increase in PUMA levels. When xenografted, p300(-) cells are more sensitive to chemotherapy with doxorubicin. These results show that p300 is a key regulator of the p53 response and suggest that p300 inhibition could be used to modulate chemotherapy.

Apoptosis↗

PubMed2004

Fugu genome analysis provides evidence for a whole-genome duplication early during the evolution of ray-finned fishes.

With about 24,000 extant species, teleosts are the largest group of vertebrates. They constitute more than 99% of the ray-finned fishes (Actinopterygii) that diverged from the lobe-finned fish lineage (Sarcopterygii) about 450 MYA. Although the role of genome duplication in the evolution of vertebrates is now established, its role in structuring the teleost genomes has been controversial. At least two hypotheses have been proposed: a whole-genome duplication in an ancient ray-finned fish and independent gene duplications in different lineages. These hypotheses are, however, based on small data sets and lack adequate statistical and phylogenetic support. In this study, we have made a systematic comparison of the draft genome sequences of Fugu and humans to identify paralogous chromosomal regions ("paralogons") in the Fugu that arose in the ray-finned fish lineage ("fish-specific"). We identified duplicate genes in the Fugu by phylogenetic analyses of the Fugu, human, and invertebrate sequences. Our analyses provide evidence for 425 fish-specific duplicate genes in the Fugu and show that at least 6.6% of the genome is represented by fish-specific paralogons. We estimated the ages of Fugu duplicate genes and paralogons using the molecular clock. Remarkably, the ages of duplicate genes and paralogons are clustered, with a peak around 350 MYA. These data strongly suggest a whole-genome duplication event early during the evolution of ray-finned fishes, probably before the origin of teleosts.

Animals↗

PubMed2004

Expression microarray reproducibility is improved by optimising purification steps in RNA amplification and labelling.

BACKGROUND: Expression microarrays have evolved into a powerful tool with great potential for clinical application and therefore reliability of data is essential. RNA amplification is used when the amount of starting material is scarce, as is frequently the case with clinical samples. Purification steps are critical in RNA amplification and labelling protocols, and there is a lack of sufficient data to validate and optimise the process. RESULTS: Here the purification steps involved in the protocol for indirect labelling of amplified RNA are evaluated and the experimentally determined best method for each step with respect to yield, purity, size distribution of the transcripts, and dye coupling is used to generate targets tested in replicate hybridisations. DNase treatment of diluted total RNA samples followed by phenol extraction is the optimal way to remove genomic DNA contamination. Purification of double-stranded cDNA is best achieved by phenol extraction followed by isopropanol precipitation at room temperature. Extraction with guanidinium-phenol and Lithium Chloride precipitation are the optimal methods for purification of amplified RNA and labelled aRNA respectively. CONCLUSION: This protocol provides targets that generate highly reproducible microarray data with good representation of transcripts across the size spectrum and a coefficient of repeatability significantly better than that reported previously.

Carbocyanines↗

PubMed2003

EMSY links the BRCA2 pathway to sporadic breast and ovarian cancer.

The BRCA2 gene is mutated in familial breast and ovarian cancer, and its product is implicated in DNA repair and transcriptional regulation. Here we identify a protein, EMSY, which binds BRCA2 within a region (exon 3) deleted in cancer. EMSY is capable of silencing the activation potential of BRCA2 exon 3, associates with chromatin regulators HP1beta and BS69, and localizes to sites of repair following DNA damage. EMSY maps to chromosome 11q13.5, a region known to be involved in breast and ovarian cancer. We show that the EMSY gene is amplified almost exclusively in sporadic breast cancer (13%) and higher-grade ovarian cancer (17%). In addition, EMSY amplification is associated with worse survival, particularly in node-negative breast cancer, suggesting that it may be of prognostic value. The remarkable clinical overlap between sporadic EMSY amplification and familial BRCA2 deletion implicates a BRCA2 pathway in sporadic breast and ovarian cancer.

BRCA2 Protein↗

PubMed2002

Whole-genome shotgun assembly and analysis of the genome of Fugu rubripes.

The compact genome of Fugu rubripes has been sequenced to over 95% coverage, and more than 80% of the assembly is in multigene-sized scaffolds. In this 365-megabase vertebrate genome, repetitive DNA accounts for less than one-sixth of the sequence, and gene loci occupy about one-third of the genome. As with the human genome, gene loci are not evenly distributed, but are clustered into sparse and dense regions. Some "giant" genes were observed that had average coding sequence sizes but were spread over genomic lengths significantly larger than those of their human orthologs. Although three-quarters of predicted human proteins have a strong match to Fugu, approximately a quarter of the human proteins had highly diverged from or had no pufferfish homologs, highlighting the extent of protein evolution in the 450 million years since teleosts and mammals diverged. Conserved linkages between Fugu and human genes indicate the preservation of chromosomal segments from the common vertebrate ancestor, but with considerable scrambling of gene order.

Animals↗