Probing Differences in Gene Essentiality Between the Human and Animal Adapted Lineages of the Mycobacterium tuberculosis Complex Using TnSeq

Members of the Mycobacterium tuberculosis complex (MTBC) show distinct host adaptations, preferences and phenotypes despite being >99% identical at the nucleic acid level. Previous studies have explored gene expression changes between the members, however few studies have probed differences in gene essentiality. To better understand the functional impacts of the nucleic acid differences between Mycobacterium bovis and Mycobacterium tuberculosis, we used the Mycomar T7 phagemid delivery system to generate whole genome transposon libraries in laboratory strains of both species and compared the essentiality status of genes during growth under identical in vitro conditions. Libraries contained insertions in 54% of possible TA sites in M. bovis and 40% of those present in M. tuberculosis, achieving similar saturation levels to those previously reported for the MTBC. The distributions of essentiality across the functional categories were similar in both species. 527 genes were found to be essential in M. bovis whereas 477 genes were essential in M. tuberculosis and 370 essential genes were common in both species. CRISPRi was successfully utilised in both species to determine the impacts of silencing genes including wag31, a gene involved in peptidoglycan synthesis and Rv2182c/Mb2204c, a gene involved in glycerophospholipid metabolism. We observed species specific differences in the response to gene silencing, with the inhibition of expression of Mb2204c in M. bovis showing significantly less growth impact than silencing its orthologue (Rv2182c) in M. tuberculosis. Given that glycerophospholipid metabolism is a validated pathway for antimicrobials, our observations suggest that target vulnerability in the animal adapted lineages cannot be assumed to be the same as the human counterpart. This is of relevance for zoonotic tuberculosis as it implies that the development of antimicrobials targeting the human adapted lineage might not necessarily be effective against the animal adapted lineage. The generation of a transposon library and the first reported utilisation of CRISPRi in M. bovis will enable the use of these tools to further probe the genetic basis of survival under disease relevant conditions.

Prehension kinematics in humans and macaques

Non-human primates, especially rhesus macaques, have been a dominant model to study sensorimotor control of the upper limbs. Indeed, human and macaques have similar hands and homologous neural circuits to mediate manual behavior. However, few studies have systematically and quantitatively compared the manual behaviors of the two species. Such comparison is critical for assessing the validity of using the macaque sensorimotor system as a model of its human counterpart. In this study, we systematically compared the prehensile behaviors of humans and rhesus macaques using an identical experimental setup. We found human and macaque prehension kinematics to be generally similar but with a few subtle differences. Humans and macaques have similar major axes of movements and similar kinematics subspaces. Human grasps are more object-specific and the movement of human digits are less correlated with each other. Monkeys demonstrate more stereotypical grasping behaviors that are common across all grasp conditions. Our results bolster the use of the macaque model to understand the neural mechanisms of manual dexterity.

An RNA-Seq-Based Framework for Characterizing Canine Prostate Cancer and Prioritizing Clinically Relevant Biomarker Candidate Genes

Prostate cancer (PCa) in dogs is a highly malignant disease akin to its human counterpart. In contrast to the situation in humans, multi-gene approaches facilitating risk stratification of canine PCa are barely established. The aims of this study were the characterization of the transcriptional landscape of canine PCa and the identification of diagnostic, prognostic and/or therapeutic biomarkers through a multi-step screening approach. RNA-Sequencing of ten malignant tissues and fine-needle aspirations (FNA), and 14 nonmalignant tissues and FNAs was performed to find differentially expressed genes (DEGs) and deregulated pathways. The 4098 observed DEGs were involved in 49 pathways. These 49 pathways could be grouped into five superpathways summarizing the hallmarks of canine PCa: (i) inflammatory response and cytokines; (ii) regulation of the immune system and cell death; (iii) cell surface and PI3K signaling; (iv) cell cycle; and (v) phagosome and autophagy. Among the highly deregulated, moderately to strongly expressed DEGs that were members of one or more superpathways, 169 DEGs were listed in relevant databases and/or the literature and included members of the PCa pathway, oncogenes, prostate-specific genes, and druggable genes. These genes are novel and promising candidate diagnostic, prognostic and/or therapeutic canine PCa biomarkers.

Probing differences in gene essentiality between the human and animal adapted lineages of the Mycobacterium tuberculosis complex using TnSeq

Members of the Mycobacterium tuberculosis complex (MTBC) show distinct host adaptations, preferences and phenotypes despite being >99% identical at the nucleic acid level. Previous studies have explored gene expression changes between the members, however few studies have probed differences in gene essentiality. To better understand the functional impacts of the nucleic acid differences between Mycobacterium bovis and Mycobacterium tuberculosis we used the Mycomar T7 phagemid delivery system to generate whole genome transposon libraries in laboratory strains of both species and compared the essentiality status of genes during growth under identical in vitro conditions. Libraries contained insertions in 54% of possible TA sites in M. bovis and 40% of those present in M. tuberculosis, achieving similar saturation levels to those previously reported for the MTBC. The distributions of essentiality across the functional categories were similar in both species. 527 genes were found to be essential in M. bovis whereas 477 genes were essential in M. tuberculosis and 370 essential genes were common in both species. CRISPRi was successfully utilised in both species to determine the impacts of silencing genes including wag31, a gene involved in peptidoglycan synthesis and Rv2182c/Mb2204c, a gene involved in glycerophospholipid metabolism. We observed species specific differences in the response to gene silencing, with the inhibition of expression of Mb2204c in M. bovis showing significantly less growth impact than silencing its ortholog (Rv2182c) in M. tuberculosis. Given that glycerophospholipid metabolism is a validated pathway for antimicrobials, our observations suggest that target vulnerability in the animal adapted lineages cannot be assumed to be the same as the human counterpart. This is of relevance for zoonotic tuberculosis as it implies that the development of antimicrobials targeting the human adapted lineage might not necessarily be effective against the animal adapted lineage. The generation of a transposon library and the first reported utilisation of CRISPRi in M. bovis will enable the use of these tools to further probe the genetic basis of survival under disease relevant conditions.

Molecular and Functional Characteristics of DNA Polymerase Beta-Like Enzymes From Trypanosomatids

Trypanosomatids are a group of primitive unicellular eukaryotes that can cause diseases in plants, insects, animals, and humans. Kinetoplast genome integrity is key to trypanosomatid cell survival and viability. Kinetoplast DNA (kDNA) is usually under attack by reactive oxygen and nitric species (ROS and RNS), damaging the DNA, and the cells must remove and repair those oxidatively generated lesions in order to survive and proliferate. Base excision repair (BER) is a well-conserved pathway for DNA repair after base damage, single-base loss, and single-strand breaks, which can arise from ROS, RSN, environmental genotoxic agents, and UV irradiation. A powerful BER system has been described in the T. cruzi kinetoplast and it is mainly carried out by DNA polymerase β (pol β) and DNA polymerase β-PAK (pol β-PAK), which are kinetoplast-located in T. cruzi as well as in other trypanosomatids. Both pol β and pol β-PAK belong to the X-family of DNA polymerases (pol X family), perform BER in trypanosomatids, and display intrinsic 5-deoxyribose phosphate (dRP) lyase and DNA polymerase activities. However, only Pol β-PAK is able to carry out trans-lesion synthesis (TLS) across 8oxoG lesions. T. cruzi cells overexpressing pol β are more resistant to ROS and are also more efficient to repair 8oxoG compared to control cells. Pol β seems to play a role in kDNA replication, since it associates with kinetoplast antipodal sites in those development stages in trypanosomatids which are competent for cell replication. ROS treatment of cells induces the overexpression of pol β, indicating that plays a role in kDNA repair. In this review, we will summarize the main features of trypanosomatid minicircle kDNA replication and the biochemical characteristics of pol β-like enzymes and their involvement in BER and kDNA replication. We also summarize key structural features of trypanosomatid pol β compared to their mammalian (human) counterpart.

A Survey on Emotion Recognition for Human Robot Interaction

With the recent developments of technology and the advances in artificial intelligent and machine learning techniques, it becomes possible for the robot to acquire and show the emotions as a part of Human-Robot Interaction (HRI). An emotional robot can recognize the emotional states of humans so that it will be able to interact more naturally with its human counterpart in different environments. In this article, a survey on emotion recognition for HRI systems has been presented. The survey aims to achieve two objectives. Firstly, it aims to discuss the main challenges that face researchers when building emotional HRI systems. Secondly, it seeks to identify sensing channels that can be used to detect emotions and provides a literature review about recent researches published within each channel, along with the used methodologies and achieved results. Finally, some of the existing emotion recognition issues and recommendations for future works have been outlined.

Mind the Eyes: Artificial Agents’ Eye Movements Modulate Attentional Engagement and Anthropomorphic Attribution

Artificial agents are on their way to interact with us daily. Thus, the design of embodied artificial agents that can easily cooperate with humans is crucial for their deployment in social scenarios. Endowing artificial agents with human-like behavior may boost individuals’ engagement during the interaction. We tested this hypothesis in two screen-based experiments. In the first one, we compared attentional engagement displayed by participants while they observed the same set of behaviors displayed by an avatar of a humanoid robot and a human. In the second experiment, we assessed the individuals’ tendency to attribute anthropomorphic traits towards the same agents displaying the same behaviors. The results of both experiments suggest that individuals need less effort to process and interpret an artificial agent’s behavior when it closely resembles one of a human being. Our results support the idea that including subtle hints of human-likeness in artificial agents’ behaviors would ease the communication between them and the human counterpart during interactive scenarios.

Generation of a humanized Aβ expressing mouse demonstrating aspects of Alzheimer’s disease-like pathology

The majority of Alzheimer’s disease (AD) cases are late-onset and occur sporadically, however most mouse models of the disease harbor pathogenic mutations, rendering them better representations of familial autosomal-dominant forms of the disease. Here, we generated knock-in mice that express wildtype human Aβ under control of the mouse App locus. Remarkably, changing 3 amino acids in the mouse Aβ sequence to its wild-type human counterpart leads to age-dependent impairments in cognition and synaptic plasticity, brain volumetric changes, inflammatory alterations, the appearance of Periodic Acid-Schiff (PAS) granules and changes in gene expression. In addition, when exon 14 encoding the Aβ sequence was flanked by loxP sites we show that Cre-mediated excision of exon 14 ablates hAβ expression, rescues cognition and reduces the formation of PAS granules.

The Traf2DNxBCL2-tg Mouse Model of Chronic Lymphocytic Leukemia/Small Lymphocytic Lymphoma Recapitulates the Biased IGHV Gene Usage, Stereotypy, and Antigen-Specific HCDR3 Selection of Its Human Counterpart

Chronic lymphocytic leukemia (CLL)/Small lymphocytic lymphoma (SLL) is a heterogeneous disease consisting of at least two separate subtypes, based on the mutation status of the immunoglobulin heavy chain variable gene (IGHV) sequence. Exposure to antigens seems to play a role in malignant transformation and in the selection and expansion of more aggressive CLL clones. Furthermore, a biased usage of particular IGHV gene subgroups and the existence of stereotyped B-cell receptors (BCRs) are distinctive characteristics of human CLL. We have previously described that Traf2DN/BCL2 double-transgenic (tg, +/+) mice develop CLL/SLL with high incidence with aging. In this model, TNF-Receptor Associated Factor (TRAF)-2 deficiency cooperates with B cell lymphoma (BCL)-2 in promoting CLL/SLL in mice by specifically enforcing marginal zone (MZ) B cell differentiation and rendering B cells independent of BAFF for survival. In this report, we have performed the sequencing of the IGHV-D-J rearrangements of B cell clones from the Traf2DN/BCL2-tg+/+ mice with CLL/SLL. The results indicate that these mice develop oligoclonal and monoclonal B cell expansions. Allotransplantation of the oligoclonal populations into immunodeficient mice resulted in the preferential expansion of one of the parental clones. The analysis of the IGHV sequences indicated that 15% were mutated (M) and 85% unmutated (UM). Furthermore, while the Traf2DN/BCL2-tg-/- (wild-type), -/+ (BCL2 single-tg) and +/- (Traf2DNDN single-tg) littermates showed the expression of various IGHV gene subgroups, the CLL/SLL expanded clones from the Traf2DN/BCL2-tg+/+ (double-transgenic) mice showed a more restricted IGHV gene subgroup usage and an overrepresentation of particular IGHV genes. In addition, the HCDR3-encoded protein sequence indicates the existence of stereotyped immunoglobulin (Ig) in the BCRs and strong similarities with BCR recognizing autoantigens and pathogen-associated antigens. Altogether, these results highlight the remarkable similarities between the CLL/SLL developed by the Traf2DN/BCL2-tg+/+ mice and its human counterpart.