ArchAlign: coordinate-free chromatin alignment reveals novel architectures

To facilitate identification and characterization of genomic functional elements, we have developed a chromatin architecture alignment algorithm (ArchAlign). ArchAlign identifies shared chromatin structural patterns from high-resolution chromatin structural datasets derived from next-generation sequencing or tiled microarray approaches for user defined regions of interest. We validated ArchAlign using well characterized functional elements, and used it to explore the chromatin structural architecture at CTCF binding sites in the human genome. ArchAlign is freely available at http://www.acsu.buffalo.edu/~mjbuck/ArchAlign.html.     

Virulence determinants, drug resistance and mobile genetic elements of Laribacter hongkongensis: a genome-wide analysis

Background

Laribacter hongkongensis is associated with community-acquired gastroenteritis and traveler's diarrhea. In this study, we performed an in-depth annotation of the genes in its genome related to the various steps in the infective process, drug resistance and mobile genetic elements.

Results

For acid and bile resistance, L. hongkongensis possessed a urease gene cassette, two arc gene clusters and bile salt efflux systems. For intestinal colonization, it possessed a putative adhesin of the autotransporter family homologous to those of diffusely adherent Escherichia coli (E. coli) and enterotoxigenic E. coli. To evade from host defense, it possessed superoxide dismutase and catalases. For lipopolysaccharide biosynthesis, it possessed the same set of genes that encode enzymes for synthesizing lipid A, two Kdo units and heptose units as E. coli, but different genes for its symmetrical acylation pattern, and nine genes for polysaccharide side chains biosynthesis. It contained a number of CDSs that encode putative cell surface acting (RTX toxin and hemolysins) and intracellular cytotoxins (patatin-like proteins) and enzymes for invasion (outer membrane phospholipase A). It contained a broad variety of antibiotic resistance-related genes, including genes related to β-lactam (n = 10) and multidrug efflux (n = 54). It also contained eight prophages, 17 other phage-related CDSs and 26 CDSs for transposases.

Conclusions

The L. hongkongensis genome possessed genes for acid and bile resistance, intestinal mucosa colonization, evasion of host defense and cytotoxicity and invasion. A broad variety of antibiotic resistance or multidrug resistance genes, a high number of prophages, other phage-related CDSs and CDSs for transposases, were also identified.     

Differential roles of RIPK1 and RIPK3 in TNF-induced necroptosis and chemotherapeutic agent-induced cell death

Apoptosis is a key mechanism for metazoans to eliminate unwanted cells. Resistance to apoptosis is a hallmark of many cancer cells and a major roadblock to traditional chemotherapy. Recent evidence indicates that inhibition of caspase-dependent apoptosis sensitizes many cancer cells to a form of non-apoptotic cell death termed necroptosis. This has led to widespread interest in exploring necroptosis as an alternative strategy for anti-cancer therapy. Here we show that in human colon cancer tissues, the expression of the essential necroptosis adaptors receptor interacting protein kinase (RIPK)1 and RIPK3 is significantly decreased compared with adjacent normal colon tissues. The expression of RIPK1 and RIPK3 was suppressed by hypoxia, but not by epigenetic DNA modification. To explore the role of necroptosis in chemotherapy-induced cell death, we used inhibitors of RIPK1 or RIPK3 kinase activity, and modulated their expression in colon cancer cell lines using short hairpin RNAs. We found that RIPK1 and RIPK3 were largely dispensable for classical chemotherapy-induced cell death. Caspase inhibitor and/or second mitochondria-derived activator of caspase mimetic, which sensitize cells to RIPK1- and RIPK3-dependent necroptosis downstream of tumor necrosis factor receptor-like death receptors, also did not alter the response of cancer cells to chemotherapeutic agents. In contrast to the RIPKs, we found that cathepsins are partially responsible for doxorubicin or etoposide-induced cell death. Taken together, these results indicate that traditional chemotherapeutic agents are not efficient inducers of necroptosis and that more potent pathway-specific drugs are required to fully harness the power of necroptosis in anti-cancer therapy.Cell death by apoptosis is a natural barrier to cancer development, as it limits uncontrolled proliferation driven by oncogenes.¹ Chemotherapeutic agents that target apoptosis have been successful in anti-cancer therapy. However, cancer cells, especially cancer stem cells, often evolve multiple mechanisms to circumvent growth suppression by apoptosis.² This resistance to apoptosis is a major challenge for many chemotherapeutic agents. Targeting other non-apoptotic cell death pathways is an attractive therapeutic alternative.A growing number of recent studies show that there are distinct genetic programmed cell death modes other than apoptosis.³ Necroptosis is mediated by receptor interacting protein kinase 3 (RIPK3).⁴ In the presence of caspase inhibition and cellular inhibitor of apoptosis proteins (cIAPs) depletion, tumor necrosis factor (TNF) receptor 1 triggers a signaling reaction that culminates in binding of RIPK3 with its upstream activator RIPK1 through the RIP homotypic interaction motif (RHIM).⁴ RIPK1 and RIPK3 phosphorylation stabilizes this complex and promotes its conversion to an amyloid-like filamentous structure termed the necrosome.⁵ Once activated, RIPK3 recruits its substrate mixed lineage kinase domain-like (MLKL).⁶ Phosphorylated MLKL forms oligomers that translocate to intracellular membranes and the plasma membrane, which eventually leads to membrane rupture.^{7, 8, 9, 10}In addition to phosphorylation, RIPK1 and RIPK3 are also tightly regulated by ubiquitination, a process mediated by the E3 ligases cIAP1, cIAP2, and the linear ubiquitin chain assembly complex.¹¹ The ubiquitin chains on RIPK1 act as a scaffold to activate nuclear factor-κB (NF-κB) and mitogen-activated protein kinase pathways and inhibit formation of the necrosome. As such, depletion of cIAP1/2 by second mitochondria-derived activator of caspase (Smac) mimetics or removal of the ubiquitin chains by the de-ubiquitinating enzyme cylindromatosis (CYLD) promotes necroptosis.^{12, 13, 14, 15} In addition, RIPK1 and RIPK3 are cleaved and inactivated by caspase 8.^{16, 17, 18} Mice deficient for caspase 8 or FADD, an essential adaptor protein of caspase 8, suffer from embryonic lethality due to extensive RIPK1- or RIPK3-dependent necroptosis.^{19, 20, 21} Hence, caspase inhibition and IAP depletion are key priming signals for necroptosis.The physiological functions of RIPK1 and RIPK3 have been extensively investigated in infectious and sterile inflammatory diseases.^{4, 22} By contrast, their roles in cancer cells'' response to chemotherapeutics are poorly understood. Here we show that RIPK1 and RIPK3 expression is significantly decreased in human colon cancer tissues, suggesting that suppression of RIPK1 or RIPK3 expression is advantageous for cancer growth. However, the loss of RIPK1 and RIPK3 expression in colon cancer was not due to epigenetic DNA modification. Interestingly, RIPK1 and RIPK3 expression in colon cancer cells is reduced by hypoxia, a hallmark of solid tumor. We found that chemotherapeutic agents did not effectively elicit RIPK1/RIPK3-dependent necroptosis in colon cancer cells. Moreover, caspase inhibition and Smac mimetics, which are potent sensitizers for necroptosis, also did not enhance chemotherapeutic agent-induced cell death. These results show that traditional chemotherapeutic agents are not strong inducers of classical necroptosis in colon cancers and suggest that development of pathway-specific drugs is needed to harness the power of necroptosis in anti-cancer therapy.     

Melatonin suppresses AOM/DSS-induced large bowel oncogenesis in rats

The inhibitory effects of exogenous melatonin (MEL) on colon oncogenesis were investigated using an azoxymethane (AOM)/dextran sodium sulfate (DSS) rat model. Male F344 rats initiated with a single intraperitoneal injection of AOM (20 mg/kg bw) were promoted by 1% (w/v) DSS in drinking water for 7 days. They were then given 0.4, 2 or 10 ppm MEL in drinking water for 17 weeks. At week 20, the development of colonic adenocarcinoma was significantly inhibited by the administration with MEL dose-dependently. MEL exposure modulated the mitotic and apoptotic indices in the colonic adenocarcinomas that developed and lowered the immunohistochemical expression of nuclear factor kappa B, tumor necrosis factor α, interleukin-1β and STAT3 in the epithelial malignancies. These results may indicate the beneficial effects of MEL on colitis-related colon carcinogenesis and a potential application for inhibiting colorectal cancer development in the inflamed colon.     

Zebrafish whole-adult-organism chemogenomics for large-scale predictive and discovery chemical biology

The ability to perform large-scale, expression-based chemogenomics on whole adult organisms, as in invertebrate models (worm and fly), is highly desirable for a vertebrate model but its feasibility and potential has not been demonstrated. We performed expression-based chemogenomics on the whole adult organism of a vertebrate model, the zebrafish, and demonstrated its potential for large-scale predictive and discovery chemical biology. Focusing on two classes of compounds with wide implications to human health, polycyclic (halogenated) aromatic hydrocarbons [P(H)AHs] and estrogenic compounds (ECs), we generated robust prediction models that can discriminate compounds of the same class from those of different classes in two large independent experiments. The robust expression signatures led to the identification of biomarkers for potent aryl hydrocarbon receptor (AHR) and estrogen receptor (ER) agonists, respectively, and were validated in multiple targeted tissues. Knowledge-based data mining of human homologs of zebrafish genes revealed highly conserved chemical-induced biological responses/effects, health risks, and novel biological insights associated with AHR and ER that could be inferred to humans. Thus, our study presents an effective, high-throughput strategy of capturing molecular snapshots of chemical-induced biological states of a whole adult vertebrate that provides information on biomarkers of effects, deregulated signaling pathways, and possible affected biological functions, perturbed physiological systems, and increased health risks. These findings place zebrafish in a strategic position to bridge the wide gap between cell-based and rodent models in chemogenomics research and applications, especially in preclinical drug discovery and toxicology.     

Territorial defense against various food competitors in the Tanganyikan benthophagous cichlid <Emphasis Type="Italic">Neolamprologus tetracanthus</Emphasis>

Territorial defense of nonbreeding female Neolamprologus tetracanthus, a shrimp-eating Tanganyikan cichlid, was investigated. Females defended territories (=home ranges, ca. 1m across) against a variety of intruding fishes. Conspecific females were usually attacked outside the territories, heterospecific benthivores (shrimp eaters) and omnivores near the border of the territories, and piscivores, algae and detritus feeders, and herbivores inside the territories. Females used some parts of the sandy substrate in the territories for foraging (foraging areas). Territorial defense prevented most of the conspecific females and benthivores from intruding into the foraging areas. In omnivores, piscivores, and algae and detritus feeders, about half the intruders were repelled from the foraging areas, although herbivores were infrequently repelled in the areas. Soon after removal of the resident females, many food competitors invaded the foraging areas and eagerly devoured prey, suggesting that the territories are maintained for food resource protection from these competitors. Females are likely to discriminate intruding fishes and change their territorial defense primarily on the basis of the degree of dietary overlap, resulting in monofunctional serial territories.