The Hexameric Structures of Human Heat Shock Protein 90

The human 90-kDa heat shock protein (HSP90) functions as a dimeric molecular chaperone. HSP90 identified on the cell surface has been found to play a crucial role in cancer invasion and metastasis, and has become a validated anti-cancer target for drug development. It has been shown to self-assemble into oligomers upon heat shock or divalent cations treatment, but the functional role of the oligomeric states in the chaperone cycle is not fully understood.Here we report the crystal structure of a truncated HSP90 that contains the middle segment and the carboxy-terminal domain, termed MC-HSP90. The structure reveals an architecture with triangular bipyramid geometry, in which the building block of the hexameric assembly is a dimer. In solution, MC-HSP90 exists in three major oligomer states, namely dimer, tetramer and hexamer, which were elucidated by size exclusion chromatography and analytical ultracentrifugation. The newly discovered HSP90 isoform HSP90N that lacks the N-terminal ATPase domain also exhibited similar oligomerization states as did MC-HSP90.While lacking the ATPase domain, both MC-HSP90 and HSP90N can self-assemble into a hexameric structure, spontaneously. The crystal structure of MC-HSP90 reveals that, in addition to the C-terminal dimerization domain, the residue W320 in the M domain plays a critical role in its oligomerization. This study not only demonstrates how the human MC-HSP90 forms a hexamer, but also justifies the similar formation of HSP90N by using 3D modeling analysis

Presence of activating KRAS mutations correlates significantly with expression of tumour suppressor genes DCN and TPM1 in colorectal cancer

<p>Abstract</p> <p>Background</p> <p>Despite identification of the major genes and pathways involved in the development of colorectal cancer (CRC), it has become obvious that several steps in these pathways might be bypassed by other as yet unknown genetic events that lead towards CRC. Therefore we wanted to improve our understanding of the genetic mechanisms of CRC development.</p> <p>Methods</p> <p>We used microarrays to identify novel genes involved in the development of CRC. Real time PCR was used for mRNA expression as well as to search for chromosomal abnormalities within candidate genes. The correlation between the expression obtained by real time PCR and the presence of the <it>KRAS </it>mutation was investigated.</p> <p>Results</p> <p>We detected significant previously undescribed underexpression in CRC for genes <it>SLC26A3</it>, <it>TPM1 </it>and <it>DCN</it>, with a suggested tumour suppressor role. We also describe the correlation between <it>TPM1 </it>and <it>DCN </it>expression and the presence of <it>KRAS </it>mutations in CRC. When searching for chromosomal abnormalities, we found deletion of the <it>TPM1 </it>gene in one case of CRC, but no deletions of <it>DCN </it>and <it>SLC26A3 </it>were found.</p> <p>Conclusion</p> <p>Our study provides further evidence of decreased mRNA expression of three important tumour suppressor genes in cases of CRC, thus implicating them in the development of this type of cancer. Moreover, we found underexpression of the <it>TPM1 </it>gene in a case of CRCs without <it>KRAS </it>mutations, showing that <it>TPM1 </it>might serve as an alternative path of development of CRC. This downregulation could in some cases be mediated by deletion of the <it>TPM1 </it>gene. On the other hand, the correlation of <it>DCN </it>underexpression with the presence of <it>KRAS </it>mutations suggests that <it>DCN </it>expression is affected by the presence of activating <it>KRAS </it>mutations, lowering the amount of the important tumour suppressor protein decorin.</p

Recombinant adenovirus IL-24-Bax promotes apoptosis of hepatocellular carcinoma cells in vitro and in vivo

Gene therapy promises to become an alternative choice for the treatment of hepatic cancer. In many cancers, the delivery of chimeric proteins by adenovirus vector has been reported to induce apoptosis. This study was performed to evaluate whether the recombinant adenovirus interleukin (IL)-24-Bax can induce apoptosis in hepatocellular carcinoma cells in vitro and in vivo. Several recombinant adenoviruses were constructed, and the expression of their encoded proteins was measured. The effects of the recombinant adenovirus on hepatocellular carcinoma cells and the normal hepatocyte cell line were investigated through cell viability and apoptosis assays after the cells were treated with Ad.Luc, Ad.IL-24, Ad.Bax or Ad.IL-24-Bax. The mechanism involved was also explored. A tumor-bearing mouse model was used to evaluate the effects of the adenovirus on tumor volume and cell apoptosis in vivo. Ad.IL-24-Bax selectively suppressed growth of hepatocellular carcinoma cells and induced apoptosis, but it had little influence on the normal hepatocytes. The mechanism of this response may include the effect of the 10HRE/VEGF385 promoter and the synergistic effect of IL-24 and Bax. Ad.IL-24-Bax also suppressed tumor growth in nude mice and induced apoptosis. Ad.IL-24-Bax may be a useful tool for gene therapy of hepatic cancer

Gene expression profile and genomic alterations in colonic tumours induced by 1,2-dimethylhydrazine (DMH) in rats

<p>Abstract</p> <p>Background</p> <p>Azoxymethane (AOM) or 1,2-dimethylhydrazine (DMH)-induced colon carcinogenesis in rats shares many phenotypical similarities with human sporadic colon cancer and is a reliable model for identifying chemopreventive agents. Genetic mutations relevant to human colon cancer have been described in this model, but comprehensive gene expression and genomic analysis have not been reported so far. Therefore, we applied genome-wide technologies to study variations in gene expression and genomic alterations in DMH-induced colon cancer in F344 rats.</p> <p>Methods</p> <p>For gene expression analysis, 9 tumours (TUM) and their paired normal mucosa (NM) were hybridized on 4 × 44K Whole rat arrays (Agilent) and selected genes were validated by semi-quantitative RT-PCR. Functional analysis on microarray data was performed by GenMAPP/MappFinder analysis. Array-comparative genomic hybridization (a-CGH) was performed on 10 paired TUM-NM samples hybridized on Rat genome arrays 2 × 105K (Agilent) and the results were analyzed by CGH Analytics (Agilent).</p> <p>Results</p> <p>Microarray gene expression analysis showed that <it>Defcr4</it>, <it>Igfbp5</it>, <it>Mmp7, Nos2, S100A8 </it>and <it>S100A9 </it>were among the most up-regulated genes in tumours (Fold Change (FC) compared with NM: 183, 48, 39, 38, 36 and 32, respectively), while <it>Slc26a3</it>, <it>Mptx</it>, <it>Retlna </it>and <it>Muc2 </it>were strongly down-regulated (FC: -500; -376, -167, -79, respectively). Functional analysis showed that pathways controlling cell cycle, protein synthesis, matrix metalloproteinases, TNFα/NFkB, and inflammatory responses were up-regulated in tumours, while Krebs cycle, the electron transport chain, and fatty acid beta oxidation were down-regulated. a-CGH analysis showed that four TUM out of ten had one or two chromosomal aberrations. Importantly, one sample showed a deletion on chromosome 18 including <it>Apc</it>.</p> <p>Conclusion</p> <p>The results showed complex gene expression alterations in adenocarcinomas encompassing many altered pathways. While a-CGH analysis showed a low degree of genomic imbalance, it is interesting to note that one of the alterations concerned <it>Apc</it>, a key gene in colorectal carcinogenesis. The fact that many of the molecular alterations described in this study are documented in human colon tumours confirms the relevance of DMH-induced cancers as a powerful tool for the study of colon carcinogenesis and chemoprevention.</p

Population Genomics of Parallel Adaptation in Threespine Stickleback using Sequenced RAD Tags

Next-generation sequencing technology provides novel opportunities for gathering genome-scale sequence data in natural populations, laying the empirical foundation for the evolving field of population genomics. Here we conducted a genome scan of nucleotide diversity and differentiation in natural populations of threespine stickleback (Gasterosteus aculeatus). We used Illumina-sequenced RAD tags to identify and type over 45,000 single nucleotide polymorphisms (SNPs) in each of 100 individuals from two oceanic and three freshwater populations. Overall estimates of genetic diversity and differentiation among populations confirm the biogeographic hypothesis that large panmictic oceanic populations have repeatedly given rise to phenotypically divergent freshwater populations. Genomic regions exhibiting signatures of both balancing and divergent selection were remarkably consistent across multiple, independently derived populations, indicating that replicate parallel phenotypic evolution in stickleback may be occurring through extensive, parallel genetic evolution at a genome-wide scale. Some of these genomic regions co-localize with previously identified QTL for stickleback phenotypic variation identified using laboratory mapping crosses. In addition, we have identified several novel regions showing parallel differentiation across independent populations. Annotation of these regions revealed numerous genes that are candidates for stickleback phenotypic evolution and will form the basis of future genetic analyses in this and other organisms. This study represents the first high-density SNP–based genome scan of genetic diversity and differentiation for populations of threespine stickleback in the wild. These data illustrate the complementary nature of laboratory crosses and population genomic scans by confirming the adaptive significance of previously identified genomic regions, elucidating the particular evolutionary and demographic history of such regions in natural populations, and identifying new genomic regions and candidate genes of evolutionary significance

Yeast caspase 1 links messenger RNA stability to apoptosis in yeast

During the past years, yeasts have been successfully established as models to study the mechanisms of apoptotic regulation. We recently showed that mutations in the LSM4 gene, which is involved in messenger RNA decapping, lead to increased mRNA stability and apoptosis in yeast. Here, we show that mitochondrial function and YCA1, which encodes a budding yeast metacaspase, are necessary for apoptosis triggered by stabilization of mRNAs. Deletion of YCA1 in yeast cells mutated in the LSM4 gene prevents mitochondrial fragmentation and rapid cell death during chronological ageing of the culture, diminishes reactive oxygen species accumulation and DNA breakage, and increases resistance to H(2)O(2) and acetic acid. mRNA levels in lsm4 mutants deleted for YCA1 are still increased, positioning the Yca1 budding yeast caspase as a downstream executor of cell death induced by mRNA perturbations. In addition, we show that mitochondrial function is necessary for fast death during chronological ageing, as well as in LSM4 mutated and wild-type cells

Guidelines for the nomenclature of the human heat shock proteins

The expanding number of members in the various human heat shock protein (HSP) families and the inconsistencies in their nomenclature have often led to confusion. Here, we propose new guidelines for the nomenclature of the human HSP families, HSPH (HSP110), HSPC (HSP90), HSPA (HSP70), DNAJ (HSP40), and HSPB (small HSP) as well as for the human chaperonin families HSPD/E (HSP60/HSP10) and CCT (TRiC). The nomenclature is based largely on the more consistent nomenclature assigned by the HUGO Gene Nomenclature Committee and used in the National Center of Biotechnology Information Entrez Gene database for the heat shock genes. In addition to this nomenclature, we provide a list of the human Entrez Gene IDs and the corresponding Entrez Gene IDs for the mouse orthologs