Clonality of HTLV-2 in Natural Infection

Human T-lymphotropic virus type 1 (HTLV-1) and type 2 (HTLV-2) both cause lifelong persistent infections, but differ in their clinical outcomes. HTLV-1 infection causes a chronic or acute T-lymphocytic malignancy in up to 5% of infected individuals whereas HTLV-2 has not been unequivocally linked to a T-cell malignancy. Virus-driven clonal proliferation of infected cells both in vitro and in vivo has been demonstrated in HTLV-1 infection. However, T-cell clonality in HTLV-2 infection has not been rigorously characterized. In this study we used a high-throughput approach in conjunction with flow cytometric sorting to identify and quantify HTLV-2-infected T-cell clones in 28 individuals with natural infection. We show that while genome-wide integration site preferences in vivo were similar to those found in HTLV-1 infection, expansion of HTLV-2-infected clones did not demonstrate the same significant association with the genomic environment of the integrated provirus. The proviral load in HTLV-2 is almost confined to CD8⁺ T-cells and is composed of a small number of often highly expanded clones. The HTLV-2 load correlated significantly with the degree of dispersion of the clone frequency distribution, which was highly stable over ∼8 years. These results suggest that there are significant differences in the selection forces that control the clonal expansion of virus-infected cells in HTLV-1 and HTLV-2 infection. In addition, our data demonstrate that strong virus-driven proliferation per se does not predispose to malignant transformation in oncoretroviral infections.     

Resveratrol depletes mitochondrial DNA and inhibition of autophagy enhances resveratrol-induced caspase activation

We recently demonstrated that resveratrol induces caspase-dependent apoptosis in multiple cancer cell types. Whether apoptosis is also regulated by other cell death mechanisms such as autophagy is not clearly defined. Here we show that inhibition of autophagy enhanced resveratrol-induced caspase activation and apoptosis. Resveratrol inhibited colony formation and cell proliferation in multiple cancer cell types. Resveratrol treatment induced accumulation of LC3-II, which is a key marker for autophagy. Pretreatment with 3-methyladenine (3-MA), an autophagy inhibitor, increased resveratrol-mediated caspase activation and cell death in breast and colon cancer cells. Inhibition of autophagy by silencing key autophagy regulators such as ATG5 and Beclin-1 enhanced resveratrol-induced caspase activation. Mechanistic analysis revealed that Beclin-1 did not interact with proapoptotic proteins Bax and Bak; however, Beclin-1 was found to interact with p53 in the cytosol and mitochondria upon resveratrol treatment. Importantly, resveratrol depleted ATPase 8 gene, and thus, reduced mitochondrial DNA (mtDNA) content, suggesting that resveratrol induces damage to mtDNA causing accumulation of dysfunctional mitochondria triggering autophagy induction. Together, our findings indicate that induction of autophagy during resveratrol-induced apoptosis is an adaptive response.     

Catalytic components of proteasomes and the regulation of proteinase activity

The proteasome (multicatalytic proteinase complex) is a large multimeric complex which is found in the nucleus and cytoplasm of eukaryotic cells. It plays a major role in both ubiquitin-dependent and ubiquitin-independent nonlysosomal pathways of protein degradation. Proteasome subunits are encoded by members of the same gene family and can be divided into two groups based on their similarity to the and subunits of the simpler proteasome isolated fromThermoplasma acidophilum. Proteasomes have a cylindrical structure composed of four rings of seven subunits. The 26S form of the proteasome, which is responsible for ubiquitin-dependent proteolysis, contains additional regulatory complexes. Eukaryotic proteasomes have multiple catalytic activities which are catalysed at distinct sites. Since proteasomes are unrelated to other known proteases, there are no clues as to which are the catalytic components from sequence alignments. It has been assumed from studies with yeast mutants that -type subunits play a catalytic role. Using a radiolabelled peptidyl chloromethane inhibitor of rat liver proteasomes we have directly identified RC7 as a catalytic component. Interestingly, mutants in Prel, the yeast homologue of RC7, have already been reported to have defective chymotrypsin-like activity. These results taken together confirm a direct catalytic role for these -type subunits. Proteasome activities are sensitive to conformational changes and there are several ways in which proteasome function may be modulatedin vivo. Our recent studies have shown that in animal cells at least two proteasome subunits can undergo phosphorylation, the level of which is likely to be important for determining proteasome localization, activity or ability to form larger complexes. In addition, we have isolated two isoforms of the 26S proteinase.     

Auditory anatomy of beaked whales and other odontocetes: Potential for cochlear stimulation via a “vibroacoustic duct mechanism”

Computed tomography (CT) and microcomputed tomography (microCT) were used to examine the structures involved in cochlear stimulation in odontocetes and terrestrial mammals. Cranial CT examined the osseous attachment of the skull to the tympanoperiotic complex (TPC) and the path of the endocranial foramen of the vestibulocochlear nerve (EFVN), which was assumed to contain the perilymphatic duct. Additional CTs of TPC were taken postextraction to examine the gross morphology of this structure. MicroCT was used to examine the acoustic windows of the cochlea, including the round and oval windows and the apertures of the cochlear and vestibular aqueducts. Cranial CT scans demonstrated an osseous connection between the skull and TPC in beaked whales and Physeter macrocephalus. EFVN traveled through a greater length of cranial bone and communicated more closely with the periotic bone in beaked whales than in other species. Ziphius cavirostris was observed to have a reduced medial sulcus of the mallear ridge (MSMR) and tympanic plate and an enlarged aperture of the cochlear aqueduct, respectively. The potential significance of these findings, including the role of the perilymphatic duct as a novel route of cochlear stimulation referred to as the “vibroacoustic duct mechanism,” are discussed.     

The Effects of Juvenile Stress on Anxiety,Cognitive Bias and Decision Making in Adulthood: A Rat Model

Stress experienced in childhood is associated with an increased risk of developing psychiatric disorders in adulthood. These disorders are particularly characterized by disturbances to emotional and cognitive processes, which are not currently fully modeled in animals. Assays of cognitive bias have recently been used with animals to give an indication of their emotional/cognitive state. We used a cognitive bias test, alongside a traditional measure of anxiety (elevated plus maze), to investigate the effects of juvenile stress (JS) on adulthood behaviour using a rodent model. During the cognitive bias test, animals were trained to discriminate between two reward bowls based on a stimulus (rough/smooth sandpaper) encountered before they reached the bowls. One stimulus (e.g. rough) was associated with a lower value reward than the other (e.g. smooth). Once rats were trained, their cognitive bias was explored through the presentation of an ambiguous stimulus (intermediate grade sandpaper): a rat was classed as optimistic if it chose the bowl ordinarily associated with the high value reward. JS animals were lighter than controls, exhibited increased anxiety-like behaviour in the elevated plus maze and were more optimistic in the cognitive bias test. This increased optimism may represent an optimal foraging strategy for these underweight animals. JS animals were also faster than controls to make a decision when presented with an ambiguous stimulus, suggesting altered decision making. These results demonstrate that stress in the juvenile phase can increase anxiety-like behaviour and alter cognitive bias and decision making in adulthood in a rat model.     

Resveratrol induces p53-independent, X-linked inhibitor of apoptosis protein (XIAP)-mediated Bax protein oligomerization on mitochondria to initiate cytochrome c release and caspase activation

Resveratrol, a naturally occurring phytoalexin, is known to induce apoptosis in multiple cancer cell types, but the underlying molecular mechanisms remain unclear. Here, we show that resveratrol induced p53-independent, X-linked inhibitor of apoptosis protein (XIAP)-mediated translocation of Bax to mitochondria where it underwent oligomerization to initiate apoptosis. Resveratrol treatment promoted interaction between Bax and XIAP in the cytosol and on mitochondria, suggesting that XIAP plays a critical role in the activation and translocation of Bax to mitochondria. This process did not involve p53 but required accumulation of Bim and t-Bid on mitochondria. Bax primarily underwent homo-oligomerization on mitochondria and played a major role in release of cytochrome c to the cytosol. Bak, another key protein that regulates the mitochondrial membrane permeabilization, did not interact with p53 but continued to associate with Bcl-xL. Thus, the proapoptotic function of Bak remained suppressed during resveratrol-induced apoptosis. Caspase-9 silencing inhibited resveratrol-induced caspase activation, whereas caspase-8 knockdown did not affect caspase activity, suggesting that resveratrol induces caspase-9-dependent apoptosis. Together, our findings characterize the molecular mechanisms of resveratrol-induced caspase activation and subsequent apoptosis in cancer cells.     

The psychoactive substance of cannabis Δ9-tetrahydrocannabinol (THC) negatively regulates CFTR in airway cells

Background

Marijuana consumption is on the rise in the US but the health benefits of cannabis smoking are controversial and the impact of cannabis components on lung homeostasis is not well-understood. Lung function requires a fine regulation of the ion channel CFTR, which is responsible for fluid homeostasis and mucocilliary clearance. The goal of this study was to assess the effect that exposure to Δ9-tetrahydrocannabinol (THC), the psychoactive substance present in marijuana, has on CFTR expression and function.