Biology subjects

Roberts, R.

Publications and source records attributed to Roberts, R..

29 records · Page 2Linked to original sources

bioRxiv2022

Development of genome-driven, lifestyle-informed primers for identification of the cereal infecting pathogens Xanthomonas translucens pathovars undulosa and translucens.

Bacterial leaf streak, blight and black chaff caused by Xanthomonas translucens pathovars are major diseases affecting small grains. Xanthomonas translucens pv. translucens and X. translucens pv. undulosa are seedborne pathogens that cause similar symptoms on barley, but only X. translucens pv. undulosa causes bacterial leaf streak of wheat. Recent outbreaks of X. translucens have been a concern for wheat and barley growers in the Northern Great Plains and Upper Midwest; however, there are limited diagnostic tools for pathovar differentiation. We developed a multiplex PCR based on whole-genome differences to distinguish X. translucens pv. translucens and X. translucens pv. undulosa. We validated the primers across different Xanthomonas and non-Xanthomonas strains. To our knowledge, these are the first multiplex PCR to distinguish X. translucens pv. translucens and X. translucens pv. undulosa. These molecular tools will support disease management strategies enabling detection and pathovar incidence analysis of X. translucens.

microbiology↗

bioRxiv2022

Visible blue light inactivates SARS-CoV-2 variants and inhibits Delta replication in differentiated human airway epithelia

The emergence of SARS-CoV-2 variants that evade host immune responses has prolonged the COVID-19 pandemic. Thus, the development of an efficacious, variant-agnostic therapeutic for the treatment of early SARS-CoV-2 infection would help reduce global health and economic burdens. Visible light therapy has the potential to fill these gaps. In this study, visible blue light centered around 425 nm efficiently inactivated SARS-CoV-2 variants in cell-free suspensions and in a translationally relevant well-differentiated tissue model of the human large airway. Specifically, 425 nm light inactivated cell-free SARS-CoV-2 variants Alpha, Beta, Delta, Gamma, Lambda, and Omicron by up to 99.99% in a dose-dependent manner, while the monoclonal antibody bamlanivimab did not neutralize the Beta, Delta, and Gamma variants. Further, we observed that 425 nm light reduced virus binding to host ACE-2 receptor and limited viral entry to host cells in vitro. Further, the twice daily administration of 32 J/cm2 of 425 nm light for three days reduced infectious SARS-CoV-2 Beta and Delta variants by >99.99% in human airway models when dosing began during the early stages of infection. In more established infections, logarithmic reductions of infectious Beta and Delta titers were observed using the same dosing regimen. Finally, we demonstrated that the 425 nm dosing regimen was well-tolerated by the large airway tissue model. Our results indicate that blue light therapy has the potential to lead to a well-tolerated and variant-agnostic countermeasure against COVID-19.

microbiology↗

bioRxiv2021

ChAdOx1 nCoV-19 (AZD1222) vaccine elicits monoclonal antibodies with potent cross-neutralizing activity against SARS-CoV-2 viral variants

Although the antibody response to COVID-19 vaccination has been studied extensively at the polyclonal level using immune sera, little has been reported on the antibody response at the monoclonal level. Here we isolate a panel of 44 anti-SARS-CoV-2 monoclonal antibodies (mAbs) from an individual who received two doses of the ChAdOx1 nCoV-19 (AZD1222) vaccine at a 12-week interval. We show that despite a relatively low serum neutralization titre, mAbs with potent neutralizing activity against the current SARS-CoV-2 variants of concern (B.1.1.7, P.1, B.1.351 and B.1.617.2) were obtained. The vaccine elicited neutralizing mAbs form 8 distinct competition groups and bind epitopes overlapping with neutralizing mAbs elicited following SARS-CoV-2 infection. AZD1222 elicited mAbs are more mutated than mAbs isolated from convalescent donors 1-2 months post infection. Spike reactive IgG+ B cells were still detectable 9-months post boost. These findings give molecular insights into AZD1222 elicited antibody response.

immunology↗

bioRxiv2020

COPB2 haploinsufficiency causes a coatopathy with osteoporosis and developmental delay

Coatomer complexes function in the sorting and trafficking of proteins between subcellular organelles. Pathogenic variants in coatomer subunits or associated factors have been reported in multi-systemic disorders, i.e., coatopathies, that can affect the skeletal and central nervous systems. We have identified loss-of-function variants in COPB2, a component of the coatomer complex I (COPI), in individuals presenting with osteoporosis, fractures and developmental delay of variable severity. Because the role of COPB2 in bone has not been characterized, we studied the effect of COPB2 deficiency on skeletal development in mice and zebrafish. Copb2+/- mice showed low bone mass and decreased bone strength. In zebrafish, larvae carrying a copb2 heterozygous frameshift variant showed delayed mineralization. copb2-null embryos showed endoplasmic reticulum (ER) and Golgi disorganization, and embryonic lethality. COPB2 siRNA-treated fibroblasts showed delayed collagen trafficking with retention of type I collagen in the ER and Golgi, and altered distribution of Golgi markers. Our data suggest that COPB2 haploinsufficiency leads to disruption of intracellular collagen trafficking and osteoporosis, which may improve with ascorbic acid supplementation. This work highlights the role of COPI complex as a critical regulator of bone mass and identifies a new form of coatopathy due to COPB2 deficiency.

genetics↗

bioRxiv2020

Region-Specific PSD-95 Interactomes Contribute to Functional Diversity of Excitatory Synapses in Human Brain.

The overarching goal of this exploratory study is to link subcellular microdomain specific protein-protein interactomes with big data analytics. We isolated postsynaptic density-95 (PSD-95) complexes from four human brain regions and compared their protein interactomes using multiple bioinformatics techniques. We demonstrate that human brain regions have unique postsynaptic protein signatures that may be used to interrogate perturbagen databases. Assessment of our hippocampal signature using the iLINCS database yielded several compounds with recently characterized "off target" effects on protein-protein interactions in the posynaptic density compartment.

neuroscience↗

bioRxiv2020

Molecular characterization of differences between the tomato immune receptors Fls3 and Fls2

Plants mount defense responses by recognizing indications of pathogen invasion, including microbe-associated molecular patterns (MAMPs). Flagellin from the bacterial pathogen Pseudomonas syringae pv. tomato (Pst) contains two MAMPs, flg22 and flgII-28, that are recognized by tomato receptors Flagellin sensing 2 (Fls2) and Flagellin sensing 3 (Fls3), respectively. It is unknown to what degree each receptor contributes to immunity and if they promote immune responses using the same molecular mechanisms. Characterization of CRISPR/Cas9-generated Fls2 and Fls3 tomato mutants revealed that the two receptors contribute equally to disease resistance both on the leaf surface and in the apoplast. However, striking differences were observed in certain host responses mediated by the two receptors. Compared to Fls2, Fls3 mediated a more sustained production of reactive oxygen species (ROS) and an increase in transcript abundance of 44 tomato genes, with two genes serving as reporters for Fls3. Fls3 had greater in vitro kinase activity and interacted differently with the Pst effector AvrPtoB as compared to Fls2. Using chimeric Fls2/Fls3 proteins, we found that no receptor domain was solely responsible for the Fls3 sustained ROS, suggesting involvement of multiple structural features. This work reveals differences in the immunity outputs between Fls2 and Fls3, suggesting they use distinct molecular mechanisms to activate pattern-triggered immunity in response to flagellin-derived MAMPs.

plant biology↗

bioRxiv2019

Spinal V2b neurons reveal a role for ipsilateral inhibition in speed control

The spinal cord contains a diverse array of interneurons that govern motor output. Traditionally, models of spinal circuits have emphasized the role of inhibition in enforcing reciprocal alternation between left and right sides or flexors and extensors. However, recent work has shown that inhibition also increases coincident with excitation during contraction. Here, using larval zebrafish, we investigate the V2b (Gata3+) class of neurons, which contribute to flexor-extensor alternation but are otherwise poorly understood. Using newly generated transgenic lines we define two stable subclasses with distinct neurotransmitter and morphological properties. These two V2b subclasses make direct synapses onto motor neurons with differential targeting to slower and faster circuits. In vivo, optogenetic suppression of V2b activity leads to increases in locomotor speed. We conclude that V2b neurons exert speed-specific influence over axial motor circuits throughout the rostrocaudal axis. Together, these results indicate a new role for ipsilateral inhibition in speed control.

neuroscience↗

bioRxiv2019

Mai1 protein acts between host recognition of pathogen effectors and MAPK signaling

The molecular mechanisms acting between host recognition of pathogen effectors by NOD-like receptor (NLR) proteins and mitogen-activated protein kinase (MAPK) signaling cascades are unknown. MAPKKK (M3K) activates MAPK signaling leading to programmed cell death (PCD) associated with NLR-triggered immunity. We identified a tomato M3K-interacting protein, SlMai1, that has 80% amino acid identity with Arabidopsis brassinosteroid kinase 1 (AtBsk1). SlMai1 has a protein kinase domain and a C-terminal tetratricopeptide repeat domain which interacts with the kinase domain of M3K. Virus-induced gene silencing of Mai1 homologs in Nicotiana benthamiana increased susceptibility to Pseudomonas syringae and compromised PCD induced by four NLR proteins. PCD was restored by expression of a synthetic SlMai1 gene that resists silencing. Expression of AtBsk1 did not restore PCD in Mai1-silenced plants, suggesting SlMai1 is functionally divergent from AtBsk1. PCD caused by overexpression of M3K or MKK2 was unaffected by Mai1 silencing indicating Mai1 acts upstream of these proteins. Co-expression of Mai1 with M3K in leaves enhanced MAPK phosphorylation and accelerated PCD. These findings reveal Mai1 as a molecular link acting between host recognition of pathogens and MAPK signaling. Author SummaryPlants use intracellular immune receptors to detect and respond to specific effector proteins which pathogens translocate into the host cell as part of their infection process. Localized programmed cell death (PCD) involving a mitogen-activated protein kinase (MAPK) cascade is an important host response associated with effector-triggered immunity, although the molecular connections between immune receptors and MAPK signaling is unknown. The Mai1 protein was found to act downstream of multiple immune receptors in Nicotiana benthamiana and to physically interact with MAPKKK. The Mai1-MAPKKK interaction enhances MAPK phosphorylation, triggers PCD and promotes disease resistance.

plant biology↗

bioRxiv2019

Natural variation for unusual host responses and flagellin-mediated immunity against Pseudomonas syringae in genetically diverse tomato accessions

The interaction between tomato and Pseudomonas syringae pv. tomato (Pst) is a well-developed model for investigating the molecular basis of the plant immune system. There is extensive natural variation in Solanum lycopersicum (tomato) but it has not been fully leveraged to enhance our understanding of the tomato-Pst pathosystem. We screened 216 genetically diverse accessions of cultivated tomato and a wild tomato species for natural variation in their response to three strains of Pst. The screen uncovered a broad range of previously unseen host symptoms in response to Pst, and one of these, stem galls, was found to be simply inherited. The screen also identified tomato accessions that showed enhanced responses to flagellin in bacterial population assays and in reactive oxygen species assays upon exposure to flagellin-derived peptides, flg22 and flgII-28. Reporter genes confirmed that the host responses were due primarily to pattern recognition receptor-immunity. This study revealed extensive natural variation in tomato for susceptibility and resistance to Pst and will enable elucidation of the molecular mechanisms underlying these host responses.

plant biology↗

bioRxiv2018

Variation in pigmentation gene expression is associated with distinct aposematic color morphs in the poison frog, Dendrobates auratus

Color and pattern phenotypes have clear implications for survival and reproduction in many species. However, the mechanisms that produce this coloration are still poorly characterized, especially at the genomic level. Here we have taken a transcriptomics-based approach to elucidate the underlying genetic mechanisms affecting color and pattern in a highly polytypic poison frog. We sequenced RNA from the skin from four different color morphs during the final stage of metamorphosis and assembled a de novo transcriptome. We then investigated differential gene expression, with an emphasis on examining candidate color genes from other taxa. Overall, we found differential expression of a suite of genes that control melanogenesis, melanocyte differentiation, and melanocyte proliferation (e.g., tyrpl, lefl, leol, and mitf) as well as several differentially expressed genes involved in purine synthesis and iridophore development (e.g., arfgapl, arfgap2, airc, and gairt). Our results provide evidence that several gene networks known to affect color and pattern in vertebrates play a role in color and pattern variation in this species of poison frog.

evolutionary biology↗

bioRxiv2018

Isoform and protein region abnormalities of dysbindin and copper transporter proteins in postmortem schizophrenia substantia nigra.

ObjectiveDysbindin is downregulated in several schizophrenia brain regions and modulates copper transport required for myelination and monoamine metabolism. We sought to determine dysbindin and copper transporter protein expression in schizophrenia subjects.\n\nMethodsWe studied the substantia nigra (which exhibits one of the highest copper contents of the human brain) using Western blot analysis. We characterized specific protein domains of copper transporters ATP7A, CTR1, ATP7B, and dysbindin isoforms 1A and 1B/C in postmortem substantia nigra in schizophrenia subjects (n=15) and matched controls (n=11). As a preliminary investigation, we examined medication status in medicated (n=11) versus unmedicated schizophrenia subjects (n=4).\n\nResultsThe combined schizophrenia group exhibited increased levels of C-terminus, but not N-terminus, ATP7A. Schizophrenia subjects expressed less transmembrane CTR1 and dysbindin 1B/C than controls. When subdivided, the increased C-terminus ATP7A protein was present only in medicated subjects versus controls. Unmedicated subjects exhibited less N-terminus ATP7A protein than controls and medicated subjects, suggesting medication-induced rescue of the ATP7A N-terminus. Transmembrane CTR1 was decreased to a similar extent in both treatment groups versus controls, suggesting no medication effect.\n\nConclusionsThese results provide the first evidence of disrupted copper transport into and within schizophrenia nigral cells that may be modulated by specific dysbindin isoforms and antipsychotic treatment.

neuroscience↗