The activity of human enhancers is modulated by the splicing of their associated lncRNAs

Pervasive enhancer transcription is at the origin of more than half of all long noncoding RNAs in humans. Transcription of enhancer-associated long noncoding RNAs (elncRNA) contribute to their cognate enhancer activity and gene expression regulation in cis. Recently, splicing of elncRNAs was shown to be associated with elevated enhancer activity. However, whether splicing of elncRNA transcripts is a mere consequence of accessibility at highly active enhancers or if elncRNA splicing directly impacts enhancer function, remains unanswered. We analysed genetically driven changes in elncRNA splicing, in humans, to address this outstanding question. We showed that splicing related motifs within multi-exonic elncRNAs evolved under selective constraints during human evolution, suggesting the processing of these transcripts is unlikely to have resulted from transcription across spurious splice sites. Using a genome-wide and unbiased approach, we used nucleotide variants as independent genetic factors to directly assess the causal relationship that underpin elncRNA splicing and their cognate enhancer activity. We found that the splicing of most elncRNAs is associated with changes in chromatin signatures at cognate enhancers and target mRNA expression. We provide evidence that efficient and conserved processing of enhancer-associated elncRNAs contributes to enhancer activity.

Development of an In Vivo Probe to Track SARS-CoV-2 Infection in Rhesus Macaques

Infection with the novel coronavirus, SARS-CoV-2, results in pneumonia and other respiratory symptoms as well as pathologies at diverse anatomical sites. An outstanding question is whether these diverse pathologies are due to replication of the virus in these anatomical compartments and how and when the virus reaches those sites. To answer these outstanding questions and study the spatiotemporal dynamics of SARS-CoV-2 infection a method for tracking viral spread in vivo is needed. We developed a novel, fluorescently labeled, antibody-based in vivo probe system using the anti-spike monoclonal antibody CR3022 and demonstrated that it could successfully identify sites of SARS-CoV-2 infection in a rhesus macaque model of COVID-19. Our results showed that the fluorescent signal from our antibody-based probe could differentiate whole lungs of macaques infected for 9 days from those infected for 2 or 3 days. Additionally, the probe signal corroborated the frequency and density of infected cells in individual tissue blocks from infected macaques. These results provide proof of concept for the use of in vivo antibody-based probes to study SARS-CoV-2 infection dynamics in rhesus macaques.

Social evolution of shared biofilm matrix components

Biofilm formation is an important and ubiquitous mode of growth among bacteria. Central to the evolutionary advantage of biofilm formation is cell-cell and cell-surface adhesion achieved by a variety of factors, some of which are diffusible compounds that may operate as classical public goods - factors that are costly to produce but may benefit other cells. An outstanding question is how diffusible matrix production, in general, can be stable over evolutionary timescales. In this work, using Vibrio cholerae as a model, we show that shared diffusible biofilm matrix proteins are indeed susceptible to cheater exploitation, and that the evolutionary stability of producing these matrix components fundamentally depends on biofilm spatial structure, intrinsic sharing mechanisms of these components, and flow conditions in the environment. We further show that exploitation of diffusible adhesion proteins is localized within a well-defined spatial range around cell clusters that produce them. Based on this exploitation range and the spatial distribution of cell clusters, we construct a model of costly diffusible matrix production and relate these length scales to the relatedness coefficient in social evolution theory. Our results show that production of diffusible biofilm matrix components is evolutionarily stable under conditions consistent with natural biofilm habitats and host environments. We expect the mechanisms revealed in this study to be relevant to other secreted factors that operate as cooperative public goods in bacterial communities, and the concept of exploitation range and the associated analysis tools to be generally applicable.

Synergistic epistasis of the deleterious effects of transposable elements

The replicative nature and generally deleterious effects of transposable elements (TEs) raise an outstanding question about how TE copy number is stably contained in host populations. Classic theoretical analyses predict that, when the decline in fitness due to each additional TE insertion is greater than linear, or when there is synergistic epistasis, selection against TEs can result in a stable equilibrium of TE copy number. While several mechanisms are predicted to yield synergistic deleterious effects of TEs, we lack empirical investigations of the presence of such epistatic interactions. Purifying selection with synergistic epistasis generates repulsion linkage between deleterious alleles. We investigated this population genetic signal in the likely ancestral Drosophila melanogaster population and found evidence supporting the presence of synergistic epistasis among TE insertions, especially TEs expected to exert large fitness impacts. Even though synergistic epistasis of TEs has been predicted to arise through ectopic recombination and TE-mediated epigenetic silencing mechanisms, we only found mixed support for the associated predictions. We observed signals of synergistic epistasis for a large number of TE families, which is consistent with the expectation that such epistatic interaction mainly happens among copies of the same family. Curiously, significant repulsion linkage was also found among TE insertions from different families, suggesting the possibility that synergism of TEs’ deleterious fitness effects could arise above the family level and through mechanisms similar to those of simple mutations. Our findings set the stage for investigating the prevalence and importance of epistatic interactions in the evolutionary dynamics of TEs.

Hierarchical length and sequence preferences establish a single major piRNA 3'-end

PIWI-interacting RNAs (piRNAs) guard germline genomes against the deleterious action of retroviruses and other mobile genetic elements. How piRNAs faithfully discriminate between self and non-self to restrict all mobile elements while sparing essential genes remains a key outstanding question in genome biology. PiRNAs use extensive base-pairing to recognize their targets and variable 3'ends could change the specificity and efficacy of piRNA silencing. Here, we identify conserved rules that ensure the generation of a single major piRNA 3'end in flies and mice. Our data suggest that the PIWI proteins initially define a short interval on pre-piRNAs that grants access to the ZUC-processor complex. Within this Goldilocks zone, the preference of the ZUC-processor to cut in front of a Uridine determines the ultimate processing site. We observe a mouse-specific roadblock that relocates the Goldilocks zone and generates an opportunity for consecutive trimming by PNLDC1. Our data reveal a conserved hierarchy between length and sequence preferences that controls the piRNA sequence space. The unanticipated precision of 3'end formation bolsters the emerging understanding that the functional piRNA sequence space is tightly controlled to ensure effective defense.

Prerenal Transplant Education and Evaluation Positively Impacts Outcomes

Introduction: An outstanding question in kidney transplantation is how to prepare candidates and their social supports for optimal posttransplant outcomes. Project Aims: This program evaluation assessed whether a pretransplant quality improvement clinic improved clinical outcomes in the year posttransplant compared to recipients receiving standard of care. Design: The Countdown to Transplant Clinic was implemented with kidney transplant candidates expected to receive a transplant within the next few months. The clinic included an enhanced education session on posttransplant lifestyle management, confirmation of support (≥2 adults), and evaluations by transplant social work, psychology, and nephrology. Results: Seventy-five patients participated in the clinic and underwent a transplant. A retrospective chart review of posttransplant laboratory values, rehospitalizations (within 3-months posttransplant), biopsy-confirmed graft failure, and mortality (within 1-year posttransplant) were collected from both groups. Univariate and multivariate propensity score-weighted linear or logistic regression models were used to evaluate the association between clinic participation and outcomes. In models adjusting for relevant covariates, participation in The Countdown to Transplant Clinic (vs standard care) was associated with a lower coefficient of variation of serum tacrolimus (all values collected 3-12 months posttransplant), 30-day posttransplant white blood cell counts (but not 90-day), 90-day posttransplant potassium, and 30 and 31 to 90 days rehospitalizations. Clinic participation did not predict serum glucose levels at 30- or 90-days posttransplant. Due to low rates of rejection and mortality, meaningful comparisons were not possible. Conclusion: Participation in a pretransplant, multicomponent clinic may improve certain outcomes of interest posttransplantation. Pilot testing for feasibility for randomized controlled trials is a necessary next step.

Early Form Based Morphological Decomposition in Tagalog: MEG Evidence from Reduplication, Infixation and Circumfixation

Neuro- and psycholinguistic experimentation supports the early decomposition of morphologically complex words within the ventral processing stream, which MEG has localized to the M170 response in the (left) visual word form area (VWFA). Decomposition into an exhaustive parse of visual morpheme forms extends beyond words like “farmer” to those imitating complexity (e.g. “brother”, Lewis et al. 2011), and to “unique” stems occurring in only one word but following the syntax and semantics of their affix (e.g. “vulnerable”, Gwilliams & Marantz 2018). Evidence comes primarily from suffixation; other morphological processes have been under-investigated. This study explores circumfixation, infixation, and reduplication in Tagalog. In addition to investigating whether these are parsed like suffixation, we address an outstanding question concerning semantically empty morphemes. Some words in Tagalog resemble English “winter” as decomposition is not supported (wint-er); these apparently reduplicated pseudoreduplicates lack the syntactic and semantic features of reduplicated forms. However, unlike “winter,” these words exhibit phonological behavior predicted only if they involve a reduplicating morpheme. If these are decomposed, this provides evidence that words are analyzed as complex, like English “vulnerable”, when the grammar demands it. In a lexical decision task with MEG, we find that VWFA activity correlates with stem:word transition probability for circumfixed, infixed and reduplicated words. Furthermore, a Bayesian analysis suggests that pseudoreduplicates with reduplicate-like phonology are also decomposed; other pseudoreduplicates are not. These findings are consistent with an interpretation that decomposition is modulated by phonology in addition to syntax and semantics.

Possibilities and limits for using the gut microbiome to improve captive animal health

Because of its potential to modulate host health, the gut microbiome of captive animals has become an increasingly important area of research. In this paper, we review the current literature comparing the gut microbiomes of wild and captive animals, as well as experiments tracking the microbiome when animals are moved between wild and captive environments. As a whole, these studies report highly idiosyncratic results with significant differences in the effect of captivity on the gut microbiome between host species. While a few studies have analyzed the functional capacity of captive microbiomes, there has been little research directly addressing the health consequences of captive microbiomes. Therefore, the current body of literature cannot broadly answer what costs, if any, arise from having a captive microbiome in captivity. Addressing this outstanding question will be critical to determining whether it is worth pursuing microbial manipulations as a conservation tool. To stimulate the next wave of research which can tie the captive microbiome to functional and health impacts, we outline a wide range of tools that can be used to manipulate the microbiome in captivity and suggest a variety of methods for measuring the impact of such manipulation preceding therapeutic use. Altogether, we caution researchers against generalizing results between host species given the variability in gut community responses to captivity and highlight the need to understand what role the gut microbiome plays in captive animal health before putting microbiome manipulations broadly into practice.

Mechanisms of ATP release in pain: role of pannexin and connexin channels

Pain is a physiological response to bodily damage and serves as a warning of potential threat. Pain can also transform from an acute response to noxious stimuli to a chronic condition with notable emotional and psychological components that requires treatment. Indeed, the management of chronic pain is currently an important unmet societal need. Several reports have implicated the release of the neurotransmitter adenosine triphosphate (ATP) and subsequent activation of purinergic receptors in distinct pain etiologies. Purinergic receptors are broadly expressed in peripheral neurons and the spinal cord; thus, purinergic signaling in sensory neurons or in spinal circuits may be critical for pain processing. Nevertheless, an outstanding question remains: what are the mechanisms of ATP release that initiate nociceptive signaling? Connexin and pannexin channels are established conduits of ATP release and have been suggested to play important roles in a variety of pathologies, including several models of pain. As such, these large-pore channels represent a new and exciting putative pharmacological target for pain treatment. Herein, we will review the current evidence for a role of connexin and pannexin channels in ATP release during nociceptive signaling, such as neuropathic and inflammatory pain. Collectively, these studies provide compelling evidence for an important role of connexins and pannexins in pain processing.

Keeping ahead of floods and droughts: a case study of Sdau Kaong Commune, Cambodia

<p>The Lower Mekong Basin covers four countries, Lao PDR, Thailand, Cambodia and Vietnam. These countries are often affected by floods and sometimes by droughts. These natural hazards silently and adversely affect people’s livelihoods in the region. In the face of future environmental changes, especially climate change and dam construction along the Mekong River, patterns of floods and droughts are more likely to exacerbate the situation. For this case study of a vulnerable commune in this setting, I developed a hybrid model of the development and complexity paradigms to both organise my research data and extend my analysis. This holistic hybrid paradigm enabled me to explore the interrelationships between natural hazards, disasters, and vulnerability, and adopt a multidisciplinary approach in which I attempt to integrate disaster risk management and climate change adaptation models to highlight problems and to propose interventions. The results obtained indicate that in the future floods and droughts are likely to be more frequent and severe and just what impact additional dams currently being planned or built will have over the control of water levels remains an outstanding question. Plans need to be made to enable people to cope with floods and droughts because these can have a hugely detrimental impact on their livelihoods including crops and personal property, people, community infrastructure and environment. Although current coping strategies are in place, disasters still occur. Based on the vulnerability context of the Sustainable Livelihoods Framework and the Pressure and Release (PAR) model, I was able to show how vulnerability is exacerbated by dissonant social, economic, and political structures. This research also proposes an integrated framework, including adaptive management and participatory action research, as a way of monitoring interventions that could possibly resolve some of the challenges.</p>