Miracle-dz/UniFAR-FTUs huggingface dataset:
The PqsE Active Site as a Target for Small Molecule Antimicrobial Agents against Pseudomonas aeruginosa

The opportunistic pathogen Pseudomonas aeruginosa causes antibiotic resistant, nosocomial infections in immuno-compromised individuals, and is a high priority for antimicrobial development. Key to pathogenicity in P. aeruginosa are biofilm formation and virulence factor production. Both traits are controlled by the cell-to-cell communication process called quorum sensing (QS). QS involves the synthesis, release, and population-wide detection of signal molecules called autoinducers. We previously reported that activity of the RhlR QS transcription factor depends on a protein-protein interaction with the hydrolase, PqsE, and PqsE catalytic activity is dispensable for this interaction. Nonetheless, the PqsE-RhlR interaction could be disrupted by substitution of an active site glutamate residue with tryptophan (PqsE(E182W)). Here, we show that disruption of the PqsE-RhlR interaction via either the E182W change or alteration of PqsE surface residues that are essential for the interaction with RhlR, attenuates P. aeruginosa infection in a murine host. We use crystallography to characterize the conformational changes induced by the PqsE(E182W) substitution to define the mechanism underlying disruption of the PqsE-RhlR interaction. A loop rearrangement that repositions the E280 residue in PqsE(E182W) is responsible for the loss of interaction. We verify the implications garnered from the PqsE(E182W) structure using mutagenic, biochemical, and additional structural analyses. We present the next generation of molecules targeting the PqsE active site, including a structure of the tightest binding of these compounds, BB584, in complex with PqsE. The findings presented here provide insight for drug discovery against P. aeruginosa with PqsE as the target. Author Summary The human pathogen Pseudomonas aeruginosa is resistant to many currently used antibiotics, making it a burden of urgent clinical importance. P. aeruginosa pathogenicity is controlled by the bacterial cell-to-cell communication process called quorum sensing (QS). The function of one protein that controls P. aeruginosa QS-directed virulence, RhlR, requires a protein-protein interaction with an enzyme called PqsE. When PqsE is blocked from interacting with RhlR, P. aeruginosa is avirulent and incapable of infecting an animal host. Here, we validate the PqsE-RhlR interaction as a target for antibiotic development, and we present a mechanism for how such antibiotics could disrupt the PqsE-RhlR interaction. Discovery of new antibiotics would fulfill an unmet healthcare need by providing treatments to combat P. aeruginosa infections.
The ciliopathy protein CCDC66 controls mitotic progression and cytokinesis by promoting microtubule nucleation and organization

Precise spatiotemporal control of microtubule nucleation and organization is critical for faithful segregation of cytoplasmic and genetic material during cell division and signaling via the primary cilium in quiescent cells. Microtubule-associated proteins (MAPs) govern assembly, maintenance, and remodeling of diverse microtubule arrays. While a set of conserved MAPs are only active during cell division, an emerging group of MAPs acts as dual regulators in dividing and non-dividing cells. Here, we elucidated the nonciliary functions and molecular mechanism of action of the ciliopathy-linked protein CCDC66, which we previously characterized as a regulator of ciliogenesis in quiescent cells. We showed that CCDC66 dynamically localizes to the spindle poles, the bipolar spindle, the spindle midzone, the central spindle and the midbody in dividing cells and interacts with the core machinery of centrosome maturation and MAPs involved in cell division. Loss-of-function experiments revealed its functions during mitotic progression and cytokinesis. Specifically, CCDC66 depletion resulted in defective spindle assembly and positioning, kinetochore fiber stability, chromosome alignment in metaphase as well as central spindle and midbody assembly and organization in anaphase and cytokinesis. Notably, CCDC66 regulates mitotic microtubule nucleation via noncentrosomal and centrosomal pathways via recruitment of gamma-tubulin to the spindle poles and the spindle. Additionally, CCDC66 bundles microtubules in vitro and in cells by its C-terminal microtubule-binding domain. Phenotypic rescue experiments showed that the microtubule and centrosome-associated pools of CCDC66 individually or cooperatively mediate its mitotic and cytokinetic functions. Collectively, our findings identify CCDC66 as a multifaceted regulator of the nucleation and organization of the diverse mitotic and cytokinetic microtubule arrays and provides new insight into nonciliary defects that underlie ciliopathies.
Identifying the Best Approximating Model in Bayesian Phylogenetics: Bayes Factors, Cross-Validation or wAIC?

There is still no consensus as to how to select models in Bayesian phylogenetics, and more generally in applied Bayesian statistics. Bayes factors are often presented as the method of choice, yet other approaches have been proposed, such as cross-validation or information criteria. Each of these paradigms raises specific computational challenges, but they also differ in their statistical meaning, being motivated by different objectives: either testing hypotheses or finding the best-approximating model. These alternative goals entail different compromises, and as a result, Bayes factors, cross-validation and information criteria may be valid for addressing different questions. Here, the question of Bayesian model selection is revisited, with a focus on the problem of finding the best-approximating model. Several model selection approaches were re-implemented, numerically assessed and compared: Bayes factors, cross-validation (CV), in its different forms (k-fold or leave-one-out), and the widely applicable information criterion (wAIC), which is asymptotically equivalent to leave-one-out cross validation (LOO-CV). Using a combination of analytical results and empirical and simulation analyses, it is shown that Bayes factors are unduly conservative. In contrast, cross-validation represents a more adequate formalism for selecting the model returning the best approximation of the data-generating process and the most accurate estimates of the parameters of interest. Among alternative CV schemes, LOO-CV and its asymptotic equivalent represented by the wAIC, stand out as the best choices, conceptually and computationally, given that both can be simultaneously computed based on standard MCMC runs under the posterior distribution.
Modelling eNvironment for Isoforms (MoNvIso): A general platform to predict structural determinants of protein isoforms in genetic diseases

The seamless integration of human disease-related mutation data into protein structures is an essential component of any attempt to correctly assess the impact of the mutation. The key step preliminary to any structural modelling is the identification of the correct isoform onto which mutations should be mapped because there are several functionally different protein isoforms from the same gene. To handle large sets of data coming from omics techniques, this challenging task should be automatized. Here we present our code MoNvIso (Modelling eNvironment for Isoforms), which pinpoints the correct isoform associated with the mutation of interest and builds a structural model of both the wild type isoform and the related variants starting from the name of the gene and the list of mutations of interest.
Birth and Death in Terminal Complement Pathway

The cytolytic activity of the membrane attack complex (MAC) has a crucial role in the complement-mediated elimination of pathogens. Terminal complement pathway (TCP) genes encode the proteins that form the MAC. Although the TCP genes are well conserved within most vertebrate species, the early evolution of the TCP genes is poorly understood. Based on the comparative genomic analysis of the early evolutionary history of the TCP homologs, we evaluated four possible scenarios that could have given rise to the vertebrate TCP. Currently available genomic data support a scheme of complex sequential protein domain gains that may be responsible for the birth of the vertebrate C6 gene. The subsequent duplication and divergence of this vertebrate C6 gene formed the C7, C8α, C8β, and C9 genes. Compared to the widespread conservation of TCP components within vertebrates, we discovered that C9 has disintegrated in the genomes of galliform birds. Publicly available genome and transcriptome sequencing datasets of chicken from Illumina short read, PacBio long read, and Optical mapping technologies support the validity of the genome assembly at the C9 locus. In this study, we have generated a >120X coverage whole-genome Chromium 10x linked-read sequencing dataset for the chicken and used it to verify the loss of the C9 gene in the chicken. We find multiple CR1 (chicken repeat 1) element insertions within and near the remnant exons of C9 in several galliform bird genomes. The reconstructed chronology of events shows that the CR1 insertions occurred after C9 gene loss in an early galliform ancestor. Our study of C6 gene birth in an early vertebrate ancestor and C9 gene death in galliform birds provides insights into the evolution of the TCP.
Pallidal neuromodulation of the explore/exploit trade-off in decision-making

Every decision that we make involves a conflict between exploiting our current knowledge of an action’s value or exploring alternative courses of action that might lead to a better, or worse outcome. The sub-cortical nuclei that make up the basal ganglia have been proposed as a neural circuit that may contribute to resolving this explore-exploit ‘dilemma’. To test this hypothesis, we examined the effects of neuromodulating the basal ganglia’s output nucleus, the globus pallidus interna, in patients who had undergone deep brain stimulation (DBS) for isolated dystonia. Neuromodulation enhanced the number of exploratory choices to the lower value option in a 2-armed bandit probabilistic reversal-learning task. Enhanced exploration was explained by a reduction in the rate of evidence accumulation (drift rate) in a reinforcement learning drift diffusion model. We estimated the functional connectivity profile between the stimulating DBS electrode and the rest of the brain using a normative functional connectome derived from heathy controls. Variation in the extent of neuromodulation induced exploration between patients was associated with functional connectivity from the stimulation electrode site to a distributed brain functional network. We conclude that the basal ganglia’s output nucleus, the globus pallidus interna, can adaptively modify decision choice when faced with the dilemma to explore or exploit.
Archaeal bundling pili of Pyrobaculum calidifontis reveal similarities between archaeal and bacterial biofilms

While biofilms formed by bacteria have received great attention due to their importance in pathogenesis, much less research has been focused on the biofilms formed by archaea. It has been known that extracellular filaments in archaea, such as Type IV pili, hami and cannulae, play a part in the formation of archaeal biofilms. We have used cryo-electron microscopy to determine the atomic structure of a previously uncharacterized class of archaeal surface filaments from hyperthermophilic Pyrobaculum calidifontis. These filaments, which we call archaeal bundling pili (ABP), assemble into highly ordered bipolar bundles. The bipolar nature of these bundles most likely arises from the association of filaments from at least two different cells. The component protein shows homology, both at the sequence and structural level, to the bacterial protein TasA, a major component of the extracellular matrix in bacterial biofilms, contributing to biofilm stability. We show that ABP forms very stable filaments in a manner similar to the donor-strand exchange of bacterial TasA fibers and chaperone-usher pathway pili where a β-strand from one subunit is incorporated into a β-sheet of the next subunit. Our results reveal mechanistic similarities and evolutionary connection between bacterial and archaeal biofilms, and suggest that there could be many other archaeal surface filaments that are as yet uncharacterized. Significance Biofilms are communities of microbes where cells attach to each other as well as to surfaces, and bacterial biofilms have been intensively studied due to their importance in many infections. Much less has been known about archaeal biofilms, where archaea are a third domain of life. Using cryo-electron microscopy, we have determined the atomic structure of a surface filament in archaea that forms bi-polar bundles connecting cells. We show that this protein has common ancestry with a protein known to be an important component of bacterial biofilms. This adds to our understanding of the evolutionary relationship between bacteria and archaea and may provide new insights into bacterial biofilms.
Multiple routes of adaptation to high levels of CIN and aneuploidy in budding yeast

Both an increased frequency of chromosome missegregation (chromosomal instability) and the presence of an abnormal complement of chromosomes (aneuploidy) are hallmarks of cancer. Paradoxically, both chromosomal instability and aneuploidy are also associated with substantial decreases in cellular fitness. To better understand how cells are able to adapt to high levels of chromosomal instability, we previously examined yeast cells that were deleted of the gene BIR1, a member of the chromosomal passenger complex (CPC). The CPC is an essential regulator of chromosome segregation fidelity. We found bir1Δ cells quickly adapted by acquiring specific combinations of beneficial aneuploidies. However, targeted mutations of specific genes were notably absent in the short term. In this study, we monitored these yeast strains for longer periods of time to determine how cells adapt to high levels of both CIN and aneuploidy in the long term. We identify suppressor mutations that mitigate the chromosome missegregation phenotype. The mutated proteins fall into four main categories: outer kinetochore subunits, members of the SCFCdc4 complex, the mitotic kinase Mps1, and a member of the CPC itself. These mutants function in two distinct ways, as mutations in the outer kinetochore suppress Bir1 deletion indirectly by destabilizing connections between the chromosomes and the mitotic spindle, whereas the other three categories of mutations affect the CPC directly. As a consequence of the accumulation of suppressor point mutations, overall levels of aneuploidy decreased. These experiments demonstrate a timeline of adaptation to high rates of CIN wherein cells first acquire specific aneuploidies that suppress the CIN phenotype, next develop point mutations that more specifically target the source of CIN, and finally reduce the level of aneuploidy to relieve the fitness burden placed by aneuploidy on the cell.
cMYC protein interactions and chromatin association in NUT carcinoma

The oncogene cMYC (HGNC:7553) is a critical genomic target of the BRD4-NUT (B4N) protein that defines many of the NUTM1-rearrangement cancer subtypes in NUT carcinoma (NC). Previously, we reported that B4N interacts with the EP300 lysine acetyltransferase (KAT3B) and creates hyperacetylated “megadomains” that activate downstream genes such as cMYC. Here we ask how misregulated cMYC in turn interacts with protein partners and target genes in patient-derived NC797 cells, and whether these interactions change in response to B4N inactivation. We used CRISPR-Cas9 mediated knock-in of a BioTAP affinity tag to analyze cMYC protein expressed from its normal chromosomal context. This allowed us to implement a crosslinking purification method termed BioTAP-XL to preserve cMYC integrity and chromatin association for genomic and mass spectrometry-based proteomic analyses. We found that in the NC797 cell line, cMYC interacts primarily with the NuA4 KAT5 lysine acetyltransferase complex, with interactions that are maintained despite a decrease in cMYC levels after JQ1 treatment. We propose that a cascade of aberrant acetyltransferase activities in NC797 cells, first via EP300 recruitment by B4N to mis-regulate cMYC, and then by KAT5 interaction with the resulting overexpressed cMYC protein, drive NC cell proliferation and blockade to differentiation. Graphical Abstract Simple Summary A longstanding goal in biology is to understand how protein interactions influence and reflect cellular disease states. MYC is a critical regulator of cellular proliferation that is mis-regulated in many cancers, including those with NUTM1-rearrangements featured in this issue. NUT carcinoma cells are dependent on MYC expression, which blocks differentiation. Using a crosslinking approach to identify MYC protein interactions in a NUTM1-rearranged patient cell line, we found that MYC interacts primarily with subunits of the NuA4 lysine acetyltransferase (KAT5), one of two KAT complexes previously discovered as MYC interactors in non-NUTM1 cell lines.
ACSS2 Regulates HIF-2α Degradation through the E3-Ubiquitin Ligase MUL1 in Clear Cell Renal Cell Carcinoma

Clear cell renal cell carcinoma (ccRCC) is an aggressive kidney cancer driven by VHL loss and aberrant HIF-2α signaling. Acetate metabolism may contribute to this axis by ACSS2-dependent acetylation of HIF-2α and may provide opportunities to intervention. Here we tested the effects of pharmacological and genetic manipulation of ACSS2 on HIF-2α, ccRCC cells, and tumors. ACSS2 inhibition led to HIF-2α degradation and suppressed ccRCC growth in vitro, in vivo, and in primary cell cultures of ccRCC patient tumors. This treatment resulted in reduced glucose and cholesterol metabolism, mitochondrial biogenesis and altered cristae deformation, that are consistent with loss of HIF-2α. Mechanistically, HIF-2α protein levels are regulated through proteolytic degradation and we found, in parallel to VHL, HIF-2α stability was dependent on ACSS2 activity to prevent direct interaction with the E3 ligase MUL1. These findings highlight ACSS2 as a critical upstream regulator of HIF-2α that may be exploited to overcome resistance to HIF-2α inhibitor therapies. STATEMENT OF SIGNIFICANCE We have unveiled ACSS2 as a critical upstream regulator of HIF-2α in ccRCC. Targeting ACSS2 potently promotes HIF-2α degradation via MUL1 to effectively deplete mitochondrial activity and block ccRCC primary tumor models and growth models resistant to HIF-2α inhibitor therapy.
Achievable degrees of freedom of MIMO two-way relay interference channel with delayed CSIT

In this paper, assuming each node has delayed channel state information at the transmitter (CSIT), we investigate the achievable degrees of freedom (DOF) of MIMO two-way relay interference channel in frequency division duplex (FDD) systems, where there are K user pairs (i.e., 2K users) and each user in a user pair exchanges messages with the other user in the same user pair simultaneously via an intermediate relay. We propose a two-stage transmission scheme and derive the closed-form expressions for its achievable DOF.
Geriatric Walk Assist Robot - Design, Analysis and Implementation of a Modular Lower Limb Exoskeleton Robot

Among the problems of major concern faced by the geriatric community the fore most is considered to be: “as we grow older it gets difficult to walk”. As they lose their strength to withstand their weight they become weak to walk on their own. Through this project a design is proposed for an assistive modular lower limb exoskeleton robot to enable aged people to walk on their own. The design is mainly based on the amplification of the pressure applied on the thighs and ankles of the legs and these pressures are used to move the legs of the robot which support the legs of the user. Since the user is capable of generating a minimum pressure on their own the working of the gait pattern is mainly based on the movement of knee and ankle, and so only the knee and the ankle are considered in this design. Another reason is that the major weight of the body acts on the knee and the foot while walking. The working of the robot is mainly based on the layered control algorithm embedded on the microcontroller acting through the sensors and actuators. Pressure sensors are used to measure the applied pressure and electrical actuators are used due to their lesser weight. Based on the experimental results obtained with the working robot the design would be fine-tuned for optimized performance.
State of the Art in Combination Immuno/Radiotherapy for Brain Metastases: Systematic Review and Meta-Analysis

Objectives: Common origins for brain metastases (BMs) are melanoma, lung, breast, and renal cell cancers. BMs account for a large share of morbidity and mortality caused by these cancers. The advent of new immunotherapeutic treatments has made a revolution in the treatment of cancer patients and particularly, as a new concept, if it is combined with radiotherapy, may lead to considerably longer survival. This systematic review and meta-analysis aimed to evaluate the survival rate and toxicities of such a combination in brain metastases. Methods: To perform a systematic review of the literature until January 2021 using electronic databases such as PubMed, Cochrane Library, and Embase; the Newcastle–Ottawa Scale was used to evaluate the quality of cohort studies. For data extraction, two reviewers extracted the data blindly and independently. Hazard ratio with 95% confidence interval (CI), fixed-effect model, and inverse-variance method was calculated. The meta-analysis has been evaluated with the statistical software Stata/MP v.16 (The fastest version of Stata). Results: In the first step, 494 studies were selected to review the abstracts, in the second step, the full texts of 86 studies were reviewed. Finally, 28 studies were selected consisting of 1465 patients. The addition of IT to RT in the treatment of brain metastasis from melanoma and non-small-cell lung carcinoma was associated with a 39% reduction in mortality rate and has prolonged overall survival, with an acceptable toxicity profile. The addition of IT to RT compared with RT alone has a hazard ratio of 0.39(95% CI 0.34–0.44). Conclusions: A combination of immuno/radiotherapy (IR) for the treatment of patients with BMs from melanoma and non-small-cell lung carcinoma has prolonged overall survival and reduced mortality rate, with acceptable toxicity. In terms of timing, RT seems to have the best effect on the result when performed before or simultaneously with immunotherapy.
Be fruitful and multiply Fitness and health in evolutionary mismatch and clinical research

Evolutionary mismatch is, roughly, poor ﬁt between an organism and its environment. Researchers in evolutionary medicine have proposed mismatch as a possible cause for morbidity and mortality in contemporary Homo sapiens populations. Mismatch hypotheses are often taken to provide an evolutionary explanation for the health outcome in question, while simultaneously oﬀering possible interventions for researchers and clinicians to pursue. A problem: ﬁtness outcomes and health outcomes are distinct. Natural selection operates on ﬁtness, not on health per se. There are cases where increased health may not contribute to ﬁtness in the modern environment. I propose an approach for using evolutionary mismatch in clinical research which sidesteps this problem. The gist of the proposal: given structural analogies between environmental causes of morbidity and environmental causes of ﬁtness reductions, evolutionary mismatch can be used as a heuristic to shrink the space through which clinical and public health researchers must search for possible interventions in response to contemporary health problems.