HDinHD: A Rich Data Portal for Huntington's Disease Research.

HDinHD (Huntington's Disease in High Definition; HDinHD.org) is an open online portal for the HD research community that presents a synthesized view of HD-related scientific data. Here, we present a broad overview of HDinHD and highlight the newly launched HDinHD Explorer tool that enables researchers to discover and explore a wide range of diverse yet interconnected HD-related data. We demonstrate the utility of HDinHD Explorer through data mining of a single collection of newly released in vivo therapeutic intervention study reports alongside previously published reports.

Identification of new human cadherin genes using a combination of protein motif search and gene finding methods.

We have combined protein motif search and gene finding methods to identify genes encoding proteins containing specific domains. Particularly, we have focused on finding new human genes of the cadherin superfamily proteins, which represent a major group of cell-cell adhesion receptors contributing to embryonic neuronal morphogenesis. Models for three cadherin protein motifs were generated from over 100 already annotated cadherin domains and used to search the complete translated human genome. The genomic sequence regions containing motif "hits" were analyzed by eukaryotic GeneMark.hmm to identify the exon-intron structure of new genes. Three new genes CDH-J, PCDH-J and FAT-J were found. The predicted proteins PCDH-J and FAT-J were classified into protocadherin and FAT-like subfamilies, respectively, based on the number and organization of cadherin domains and presence of subfamily-specific conserved amino acid residues. Expression of FAT-J was shown in almost all tested tissues. The exon-intron organization of CDH-J was experimentally verified by PCR with specifically designed primers and its tissue-specific expression was demonstrated. The described methodology can be applied to discover new genes encoding proteins from families with well-characterized structural and functional domains.

Enhanced homology searching through genome reading frame predetermination.

MOTIVATION: Many bioinformatic approaches exist for finding novel genes within genomic sequence data. Traditionally, homology search-based methods are often the first approach employed in determining whether a novel gene exists that is similar to a known gene. Unfortunately, distantly related genes or motifs often are difficult to find using single query-based homology search algorithms against large sequence datasets such as the human genome. Therefore, the motivation behind this work was to develop an approach to enhance the sensitivity of traditional single query-based homology algorithms against genomic data without losing search selectivity. RESULTS: We demonstrate that by searching against a genome fragmented into all possible reading frames, the sensitivity of homology-based searches is enhanced without degrading its selectivity. Using the ETS-domain, bromodomain and acetyl-CoA acetyltransferase gene as queries, we were able to demonstrate that direct protein-protein searches using BLAST2P or FASTA3 against a human genome segmented among all possible reading frames and translated was substantially more sensitive than traditional protein-DNA searches against a raw genomic sequence using an application such as TBLAST2N. Receiver operating characteristic analysis was employed to demonstrate that the algorithms remained selective, while comparisons of the algorithms showed that the protein-protein searches were more sensitive in identifying hits. Therefore, through the overprediction of reading frames by this method and the increased sensitivity of protein-protein based homology search algorithms, a genome can be deeply mined, potentially finding hits overlooked by protein-DNA searches against raw genomic data.

Farnesoid X receptor activates transcription of the phospholipid pump MDR3.

The human multidrug resistance gene MDR3 encodes a P-glycoprotein that belongs to the ATP-binding cassette transporter family (ABCB4). MDR3 is a critical trans-locator for phospholipids across canalicular membranes of hepatocytes, evidenced by the fact that human MDR3 deficiencies result in progressive familial intrahepatic cholestasis type III. It has been reported previously that MDR3 expression is modulated by hormones, cellular stress, and xenobiotics. Here we show that the MDR3 gene is trans-activated by the farnesoid X receptor (FXR) via a direct binding of FXR/retinoid X receptor alpha heterodimers to a highly conserved inverted repeat element (a FXR response element) at the distal promoter (-1970 to -1958). In FXR trans-activation assays, both the endogenous FXR agonist chenodeoxycholate and the synthetic agonist GW4064 activated the MDR3 promoter. Deletion or mutation of this inverted repeat element abolished FXR-mediated MDR3 promoter activation. Consistent with these data, MDR3 mRNA was significantly induced by both chenodeoxycholate and GW4064 in primary human hepatocytes in time- and dose-dependent fashions. In conclusion, we demonstrate that MDR3 expression is directly up-regulated by FXR. These results, together with the previous report that the bile salt export pump is a direct FXR target, suggest that FXR coordinately controls secretion of bile salts and phospholipids. Results of this study further support the notion that FXR is a master regulator of lipid metabolism.

Human kininogen gene is transactivated by the farnesoid X receptor.

Human kininogen belongs to the plasma kallikreinkinin system. High molecular weight kininogen is the precursor for two-chain kinin-free kininogen and bradykinin. It has been shown that the two-chain kinin-free kininogen has the properties of anti-adhesion, anti-platelet aggregation, and anti-thrombosis, whereas bradykinin is a potent vasodilator and mediator of inflammation. In this study we show that the human kininogen gene is strongly up-regulated by agonists of the farnesoid X receptor (FXR), a nuclear receptor for bile acids. In primary human hepatocytes, both the endogenous FXR agonist chenodeoxycholate and synthetic FXR agonist GW4064 increased kininogen mRNA with a maximum induction of 8-10-fold. A more robust induction of kininogen expression was observed in HepG2 cells, where kininogen mRNA was increased by chenodeoxycholate or GW4064 up to 130-140-fold as shown by real time PCR. Northern blot analysis confirmed the up-regulation of kininogen expression by FXR agonists. To determine whether kininogen is a direct target of FXR, we examined the sequence of the kininogen promoter and identified a highly conserved FXR response element (inverted repeat, IR-1) in the proximity of the kininogen promoter (-66/-54). FXR/RXRalpha heterodimers specifically bind to this IR-1. A construct of a minimal promoter with the luciferase reporter containing this IR-1 was transactivated by FXR. Deletion or mutation of this IR-1 abolished FXR-mediated promoter activation, indicating that this IR-1 element is responsible for the promoter transactivation by FXR. We conclude that kininogen is a novel and direct target of FXR, and bile acids may play a role in the vasodilation and anti-coagulation processes.

An interactive tool for extracting exons and SNP from genomic sequence: isolation of HCN1 and HCN3 ion channel genes.

Genome Analyzer (GenoA) with a relational database back-end, was developed to extract information from mammalian genomic sequences. This data mining and visualization tool-set enables laboratory bench scientists to identify and assemble virtual cDNA from genomic exon sequences, and provides a starting point to identify potential alternative splice variants and polymorphisms in silico. The study described in this paper demonstrates the use of GenoA to study human brain hyperpolarization-activated cation channel genes HCN1 and HCN3.

Molecular characterization of the murine SIGNR1 gene encoding a C-type lectin homologous to human DC-SIGN and DC-SIGNR.

The C-type lectin human dendritic cell (DC)-specific intercellular adhesion molecule (ICAM)-3-grabbing non-integrin (DC-SIGN) plays important roles in pattern recognition by dendritic cells in the immune system. In addition to binding human immunodeficiency virus (HIV), this type II membrane protein binds with high affinity to the adhesion molecules ICAM-3 and -2 to promote important dendritic cell interactions with naive T cells and endothelial cells, respectively. DC-SIGNR, a human DC-SIGN homologue expressed on sinusoidal endothelial cells in liver and lymph node, also binds and transmits HIV virus. We describe the cloning and characterization of a family of murine complementary DNAs (cDNAs) called SIGNR1, expressed in skin and spleen, that encode C-type lectins highly related to human DC-SIGN and DC-SIGNR. We also report the genomic structure of the SIGNR1 gene and compare it to that of human DC-SIGN and DC-SIGNR. The different transcripts (alpha, beta, gamma, delta) are generated by differences in 5' untranslated sequences, alternative splicing and/or the use of different polyadenylation sites. The predicted open reading frames encoded by the cDNAs are most closely related to human DC-SIGN and DC-SIGNR in the cytoplasmic domain, the transmembrane region and the carbohydrate recognition domain. Moreover, the alternatively spliced transcripts encode proteins that lack the transmembrane region or have modified carbohydrate recognition domains. Northern hybridization experiments with several different SIGNR1 cDNA probes reveal transcripts of 1.3 and 2.1 kb that are expressed in a tissue-restricted fashion in murine skin, spleen and lung. In situ hybridization and immunocytochemistry experiments demonstrate that, like human DC-SIGN, the murine messenger RNAs are expressed in subsets of dendritic cells in the spleen and skin.