Context-dependent role of ATG4B as target for autophagy inhibition in prostate cancer therapy

ATG4B belongs to the autophagin family of cysteine proteases required for autophagy, an emerging target of cancer therapy. Developing pharmacological ATG4B inhibitors is a very active area of research. However, detailed studies on the role of ATG4B during anticancer therapy are lacking. By analyzing PC-3 and C4-2 prostate cancer cells overexpressing dominant negative ATG4B^C74Ain vitro and in vivo, we show that the effects of ATG4B^C74A are cell type, treatment, and context-dependent. ATG4B^C74A expression can either amplify the effects of cytotoxic therapies or contribute to treatment resistance. Thus, the successful clinical application of ATG4B inhibitors will depend on finding predictive markers of response.     

Food Prices and Consumer Demand: Differences across Income Levels and Ethnic Groups

Background

Targeted food pricing policies may improve population diets. To assess their effects on inequalities, it is important to determine responsiveness to price changes across income levels and ethnic groups.

Objective

Our goal was to estimate price elasticity (PE) values for major commonly consumed food groups in New Zealand, by income and ethnicity. PE values represent percentage change in demand associated with 1% change in price of that good (own-PE) or another good (cross-PE).

Design

We used food expenditure data from national household economic surveys in 2007/08 and 2009/10 and Food Price Index data from 2007 and 2010. Adopting an Almost Ideal Demand System approach, own-PE and cross-PE estimates were derived for 24 food categories, household income quintiles, and two ethnic groups (Māori and non-Māori).

Results

Own-PE estimates (with two exceptions) ranged from −0.44 to −1.78. Cross-PE estimates were generally small; only 31% of absolute values were greater than 0.10. Excluding the outlier ‘energy drinks’, nine of 23 food groups had significantly stronger own-PEs for the lowest versus highest income quintiles (average regression-based difference across food groups −0.30 (95% CI −0.62 to 0.02)). Six own-PEs were significantly stronger among Māori; the average difference for Māori: non-Māori across food groups was −0.26 (95% CI −0.52 to 0.00).

Conclusions

Food pricing policies have potential to improve population diets. The greater sensitivity of low-income households and Māori to price changes suggests the beneficial effects of such policies on health would be greatest for these groups.     

Regulation of Interferon-Induced Protein Kinase PKR: Modulation of P58IPK Inhibitory Function by a Novel Protein, P52rIPK

The cellular response to environmental signals is largely dependent upon the induction of responsive protein kinase signaling pathways. Within these pathways, distinct protein-protein interactions play a role in determining the specificity of the response through regulation of kinase function. The interferon-induced serine/threonine protein kinase, PKR, is activated in response to various environmental stimuli. Like many protein kinases, PKR is regulated through direct interactions with activator and inhibitory molecules, including P58^IPK, a cellular PKR inhibitor. P58^IPK functions to represses PKR-mediated phosphorylation of the eukaryotic initiation factor 2α subunit (eIF-2α) through a direct interaction, thereby relieving the PKR-imposed block on mRNA translation and cell growth. To further define the molecular mechanism underlying regulation of PKR, we have utilized an interaction cloning strategy to identify a novel cDNA encoding a P58^IPK-interacting protein. This protein, designated P52^rIPK, possesses limited homology to the charged domain of Hsp90 and is expressed in a wide range of cell lines. P52^rIPK and P58^IPK interacted in a yeast two-hybrid assay and were recovered as a complex from mammalian cell extracts. When coexpressed with PKR in yeast, P58^IPK repressed PKR-mediated eIF-2α phosphorylation, inhibiting the normally toxic and growth-suppressive effects associated with PKR function. Conversely, introduction of P52^rIPK into these strains resulted in restoration of both PKR activity and eIF-2α phosphorylation, concomitant with growth suppression due to inhibition of P58^IPK function. Furthermore, P52^rIPK inhibited P58^IPK function in a reconstituted in vitro PKR-regulatory assay. Our results demonstrate that P58^IPK is inhibited through a direct interaction with P52^rIPK which, in turn, results in upregulation of PKR activity. Taken together, our data describe a novel protein kinase-regulatory system which encompasses an intersection of interferon-, stress-, and growth-regulatory pathways.     

GROWTH AND ORGANIZED DEVELOPMENT OF CULTURED CELLS. V. THE GROWTH OF COLONIES FROM FREE CELLS ON NUTRIENT AGAR

When angiosperm cells are cultured in a liquid medium they may grow in the form of free, single cells and form small to large groups of cells. This has been shown in earlier papers. This paper deals with the growth of strains of cells (Daucus carota and Haplopappus gracilis cells were used), washed and filtered free from the larger groups, on nutrient agar media in Petri dishes, thus simulating familiar microbiological technique. Each discrete member of a suspension is referred to as a “unit.” On the order of 1–10% of the separate units of a suspension may be induced to grow into viable colonies, depending on the strain and the conditions employed. Whereas at least 30% of the free single carrot cells were shown to be capable of division, only up to about 4% continued their growth to form macroscopically visible colonies when they were widely dispersed. Coconut milk promotes the growth of carrot cells into colonies. Both coconut milk and napthaleneacetic acid, which interact synergistically, arc required for the growth of Haplopappus cells. Various techniques which affect viability (the frequency with which units grow into colonies) were investigated. The viability of carrot units was found (1) to increase with their density on the plates; (2) to decrease upon washing the suspensions prior to plating; (3) to increase with increasing initial size; and (4) to decrease to a vanishingly low value in rigorously filtered suspensions which consist principally of single cells, although the single cells were found to grow with appreciable frequency when the larger units were also present; and (5) to increase dramatically (100-fold) when a rigorously filtered suspension was plated on a medium upon which pieces of growing cultured tissue were placed. Thus, the induction of growth in free cells is enhanced, even in an environment nutritionally optimal for the growth of the larger cell masses, by as yet unknown factors which are contributed by the cells themselves, or by adjacent cells or groups of cells. It is suggested that within a group of cells growing in culture, and perhaps also in the organized growing regions of intact plants, the dividing cells are nourished or stimulated by adjacent but less frequently dividing cells. The implications of these results are discussed.     

Multi-drug resistant Bacteroides fragilis recovered from blood and severe leg wounds caused by an improvised explosive device (IED) in Afghanistan

This report summarizes the case of a 23 year-old otherwise healthy male that was injured in an improvised explosive device (IED) blast in support of Operation Enduring Freedom (OEF). He sustained bilateral open tibia and fibula fractures in the setting of being exposed to water contaminated with raw sewage. Despite long-term carbapenem therapy, the patient's wounds were repeatedly noted to have purulent drainage during surgical debridement and cultures from these wounds were persistently positive for Bacteroides fragilis. Apparent clinical failure persisted despite the addition of metronidazole to his regimen and an eventual trial of tigecycline. Susceptibility testing of the B.?fragilis isolate was performed and resistance to penicillin, clindamycin,metronidazole, cefoxitin, meropenem, imipenem, piperacillin/tazobactam, and tigecycline was confirmed. The presence of a nimE gene on a potentially transferrable plasmid was also confirmed by plasmid sequencing. The only antibiotics that displayed in vitro susceptibility were moxifloxacin and linezolid. These antibiotics were initiated in combination with aggressive irrigation and serial surgical debridement. Conversion to left-sided internal fixation became feasible and his left lower extremity was salvaged without residual evidence of infection. The patient completed an eight week course of combination moxifloxacin and linezolid therapy without adverse event. This B.?fragilis isolate displayed simultaneous high-level resistance to multiple antibiotics routinely utilized in anaerobic infections. This was evidenced by clinical failure, in vitro susceptibility testing, and demonstration of genes associated with resistance mechanisms. This case warrants review not only due to the rarity of this event but also the potential implications regarding anaerobic infections in traumatic wounds and the success of a novel treatment regimen utilizing combination therapy with moxifloxacin and linezolid.     

Removal of a frameshift between the hsdM and hsdS genes of the EcoKI Type IA DNA restriction and modification system produces a new type of system and links the different families of Type I systems

The EcoKI DNA methyltransferase is a trimeric protein comprised of two modification subunits (M) and one sequence specificity subunit (S). This enzyme forms the core of the EcoKI restriction/modification (RM) enzyme. The 3′ end of the gene encoding the M subunit overlaps by 1 nt the start of the gene for the S subunit. Translation from the two different open reading frames is translationally coupled. Mutagenesis to remove the frameshift and fuse the two subunits together produces a functional RM enzyme in vivo with the same properties as the natural EcoKI system. The fusion protein can be purified and forms an active restriction enzyme upon addition of restriction subunits and of additional M subunit. The Type I RM systems are grouped into families, IA to IE, defined by complementation, hybridization and sequence similarity. The fusion protein forms an evolutionary intermediate form lying between the Type IA family of RM enzymes and the Type IB family of RM enzymes which have the frameshift located at a different part of the gene sequence.