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ABSTRACT In response to the host environment, the human pathogen Cryptococcus neoformans must rapidly reprogram its translatome from one which promotes growth to one which is responsive to host stress. In this study, we investigate the two events which comprise translatome reprogramming: the removal of abundant, pro-growth mRNAs from the translating pool, and the regulated entry of stress-responsive mRNAs into the translating pool. Removal of pro-growth mRNAs from the translating pool is controlled primarily by two regulatory mechanisms, repression of translation initiation via Gcn2, and decay mediated by Ccr4. We determined that translatome reprogramming in response to oxidative stress requires both Gcn2 and Ccr4, whereas the response to temperature requires only Ccr4. Additionally, we assessed ribosome collision in response to host-relevant stress and found that collided ribosomes accumulated during temperature stress but not during oxidative stress. The phosphorylation of eIF2α that occurred as a result of translational stress led us to investigate the induction of the integrated stress response (ISR). We found that eIF2α phosphorylation varied in response to the type and magnitude of stress, yet all tested conditions induced translation of the ISR transcription factor Gcn4. However, Gcn4 translation did not necessarily result in canonical Gcn4-dependent transcription. Finally, we define the ISR regulon in response to oxidative stress. In conclusion, this study begins to reveal the translational regulation in response to host-relevant stressors in an environmental fungus which is capable of adapting to the environment inside the human host. IMPORTANCE Cryptococcus neoformans is a human pathogen capable of causing devastating infections. It must rapidly adapt to changing environments as it leaves its niche in the soil and enters the human lung. Previous work has demonstrated a need to reprogram gene expression at the level of translation to promote stress adaptation. In this work, we investigate the contributions and interplay of the major mechanisms that regulate entry of new mRNAs into the pool (translation initiation) and the clearance of unneeded mRNAs from the pool (mRNA decay). One result of this reprogramming is the induction of the integrated stress response (ISR) regulon. Surprisingly, all stresses tested led to the production of the ISR transcription factor Gcn4, but not necessarily to transcription of ISR target genes. Furthermore, stresses result in differential levels of ribosome collisions, but these are not necessarily predictive of initiation repression as has been suggested in the model yeast.

Long non-coding RNAs (lncRNAs) are highly expressed and can modulate multiple cellular processes including transcription, splicing, translation, and many diverse signaling events. LncRNAs can act as sponges for miRNAs, RNA and DNA binding proteins, functioning as competitive endogenous RNAs. The contribution of lncRNAs to microbial pathogenesis is largely neglected in eukaryotic pathogens despite the abundance of RNA sequencing datasets encompassing conditions of stress, gene deletions and conditions that mimic the host environment. The human fungal pathogen Cryptococcus neoformans encodes 6975 (84%) protein-coding and 1359 (16%) non-protein-coding RNAs, of which 1182 (14.2%) are lncRNAs defined by a threshold of greater than 200 nucleotides in length. Here, we discuss the current state of knowledge in C. neoformans lncRNA biology. Utilizing existing RNA seq datasets, we examine trends in lncRNA expression and discuss potential implications for pathogenesis.

Fungal infections are one of the leading causes of death for people who live with HIV/AIDS. Even though these pathogens are independently well studied, it is still enigmatic how coinfection with HIV-1 and C. neoformans

Cryptococcus neoformans

The classical form of α1-antitrypsin deficiency (ATD) is characterized by intracellular accumulation of the misfolded variant α1-antitrypsin Z (ATZ) and severe liver disease in some of the affected individuals. In this study, we investigated the possibility of discovering novel therapeutic agents that would reduce ATZ accumulation by interrogating a C. elegans model of ATD with high-content genome-wide RNAi screening and computational systems pharmacology strategies. The RNAi screening was utilized to identify genes that modify the intracellular accumulation of ATZ and a novel computational pipeline was developed to make high confidence predictions on repurposable drugs. This approach identified glibenclamide (GLB), a sulfonylurea drug that has been used broadly in clinical medicine as an oral hypoglycemic agent. Here we show that GLB promotes autophagic degradation of misfolded ATZ in mammalian cell line models of ATD. Furthermore, an analog of GLB reduces hepatic ATZ accumulation and hepatic fibrosis in a mouse model in vivo without affecting blood glucose or insulin levels. These results provide support for a drug discovery strategy using simple organisms as human disease models combined with genetic and computational screening methods. They also show that GLB and/or at least one of its analogs can be immediately tested to arrest the progression of human ATD liver disease.

The insect-transmitted protozoan parasite Trypanosoma cruzi experiences changes in nutrient availability and rate of flux through different metabolic pathways across its life cycle. The species encompasses much genetic diversity of both the nuclear and mitochondrial genomes among isolated strains. The genetic or expression variation of both genomes are likely to impact metabolic responses to environmental stimuli, and even steady state metabolic function, among strains. To begin formal characterization these differences, we compared aspects of metabolism between genetically similar strains CL Brener and Tulahuen with less similar Esmeraldo and Sylvio X10 strains in a culture environment. Epimastigotes of all strains took up glucose at similar rates. However, the degree of medium acidification that could be observed when glucose was absent from the medium varied by strain, indicating potential differences in excreted metabolic byproducts. Our main focus was differences related to electron transport chain function. We observed differences in ATP-coupled respiration and maximal respiratory capacity, mitochondrial membrane potential, and mitochondrial morphology between strains, despite the fact that abundances of two nuclear-encoded proteins of the electron transport chain are similar between strains. RNA sequencing reveals strain-specific differences in abundances of mRNAs encoding proteins of the respiratory chain but also other metabolic processes. From these differences in metabolism and mitochondrial phenotypes we have generated tentative models for the differential metabolic fluxes or differences in gene expression that may underlie these results.

ABSTRACT Trypanosoma cruzi parasites causing Chagas disease are passed between mammals by the triatomine bug vector. Within the insect, T. cruzi epimastigote-stage cells replicate and progress through the increasingly nutrient-restricted digestive tract, differentiating into infectious, nonreplicative metacyclic trypomastigotes. Thus, we evaluated how nutrient perturbations or metacyclogenesis affects mitochondrial gene expression in different insect life cycle stages. We compared mitochondrial RNA abundances in cultures containing fed, replicating epimastigotes, differentiating cultures containing both starved epimastigotes and metacyclic trypomastigotes and epimastigote starvation cultures. We observed increases in mitochondrial rRNAs and some mRNAs in differentiating cultures. These increases predominated only for the edited CYb mRNA in cultures enriched for metacyclic trypomastigotes. For the other transcripts, abundance increases were linked to starvation and were strongest in culture fractions with a high population of starved epimastigotes. We show that loss of both glucose and amino acids results in rapid increases in RNA abundances that are quickly reduced when these nutrients are returned. Furthermore, the individual RNAs exhibit distinct temporal abundance patterns, suggestive of multiple mechanisms regulating individual transcript abundance. Finally, increases in mitochondrial respiratory complex subunit mRNA abundances were not matched by increases in abundances of nucleus-encoded subunit mRNAs, nor were there statistically significant increases in protein levels of three nucleus-encoded subunits tested. These results show that, similarly to that in T. brucei, the mitochondrial genome in T. cruzi has the potential to alter gene expression in response to environmental or developmental stimuli but for an as-yet-unknown purpose. IMPORTANCE Chagas disease is caused by insect-transmitted Trypanosoma cruzi. Halting T. cruzi’s life cycle in one of its various human and insect life stages would effectively stop the parasite’s infection cycle. T. cruzi is exposed to a variety of environmental conditions in its different life stages, and gene expression must be remodeled to survive these changes. In this work, we look at the impact that one of these changes, nutrient depletion, has on the expression of the 20 gene products encoded in the mitochondrial genome that is neglected by whole-genome studies. We show increases in mitochondrial RNA abundances in starved insect-stage cells, under two conditions in which transition to the infectious stage occurs or does not. This report is the first to show that T. cruzi mitochondrial gene expression is sensitive to environmental perturbations, consistent with mitochondrial gene expression regulatory pathways being potential antiparasitic targets.

This paper analyzes the deterrence implications of different inmate assistance programs (IAPs), carefully distinguishing between deterrence of first offenses and recidivism. All IAPs considered in our model reduce recidivism, and we identify IAPs that also reduce the first offense rate. However, IAPs which increase work opportunities, improve the reintegration of exiting inmates, and moderate individual's self-control issues may lower the deterrence of first offenses, if this possibility is not adequately anticipated when designing these IAPs. [ABSTRACT FROM AUTHOR]

We present conservative estimates for the marginal value of public funds (MVPF) associated with providing Medicaid to inmates exiting prison. The MVPF measures the ratio between a policy's social benefits and its governmental costs. Our MVPF estimates suggest that every additional $1 the government spends on providing inmates exiting prison with Medicaid coverage can result in social benefits ranging between $3.45 and $10.62. A large proportion of the benefits we consider stems from the reduced future criminal involvement among former inmates who receive Medicaid. Employing a difference‐in‐differences approach, we find that Medicaid expansions reduce the average number of times a released inmate is reimprisoned within 1 year by approximately 11.5%. By combining this estimate with key values reported elsewhere (e.g., victimization costs, data on victimization and incarceration), we quantify specific benefits arising from the policy. These encompass diminished criminal harm due to lower reoffense rates, direct benefits to former inmates through Medicaid coverage, increased employment opportunities, and reduced loss of liberty resulting from fewer future reimprisonments. Net‐costs consist of the cost of providing Medicaid net of changes in the governmental cost of imprisonment, changes in the tax revenue due to increased employment, and changes in spending on other public assistance programs. We interpret our estimates as conservative since we deliberately err on the side of under‐estimating benefits and over‐estimating costs when data on specific items are imprecise or incomplete. Our findings align closely with others in the sparse literature investigating the crime‐related welfare impacts of Medicaid access, underscoring the substantial indirect benefits public health insurance programs can offer through crime reduction, in addition to their direct health‐related advantages. [ABSTRACT FROM AUTHOR]

We consider a previously neglected cost of whistle‐blower awards: employers may base their hiring decisions, on the margin, not on the productivity of an employee but rather on the probability that the employee will become a whistle‐blower. We develop a three‐stage model to examine how productivity losses due to distortions at the hiring stage influence optimal whistle‐blower rewards. We characterize optimal rewards for whistle‐blowing, and show that when rewards can be chosen according to either the benefits of the employer from offending or the productivity of the worker being hired, productivity‐based rewards are superior to benefit‐based rewards. [ABSTRACT FROM AUTHOR]

A reform movement is underway in antitrust. Citing prior enforcement failures, deviations from the original intent of the antitrust laws, and overall rising levels of sector concentration, some are seeking to fundamentally alter or altogether replace the current consumer welfare standard, which has guided courts over the past fifty years. This policy push has sparked an intense debate over the best approach to antitrust law enforcement. In this Article, we examine a previously unexplored potential social cost from moving away from the consumer welfare standard: a loss in the information value to the public from a finding of liability. A virtue of the current standard is the knowledge that firms who violate the antitrust laws have harmed consumers. This simple reality is a direct, easy-to-interpret signal to market participants and investors. In contrast, a broader and more nebulous standard--such as a "public interest" approach, which has been proposed by some academics and agency officials--could conceivably water down the information value of a finding of liability. In essence, the greater the license that regulators and courts have to condemn a business practice beyond a finding of harm to consumers, the noisier the signal to the public about what the verdict means. We can call this phenomenon "the stigma dilution effect." To that end, we develop a formal model to gain insight into the role of reputation in the enforcement and deterrence effects of antitrust laws. The model reveals that broadening the welfare standard is likely to weaken the reputational impact of antitrust violations. This dilution can, in turn, have implications that go against what the proponents of abolishing the consumer welfare standard desire. Namely, a new standard could increase, rather than decrease, the frequency of conduct they seek to deter. Thus, our analysis suggests there may be important and underappreciated costs associated with departures from the consumer welfare standard. In fact, the presence of reputational considerations suggests that these departures can produce effects contrary to the stated goals of their proponents. [ABSTRACT FROM AUTHOR]