Biology subjects

Cohen, I.

Publications and source records attributed to Cohen, I..

31 records · Page 2Linked to original sources

bioRxiv2021

MEK1/2 inhibition transiently alters the tumor immune microenvironment to enhance immunotherapy efficacy against head and neck cancer

Although the mitogen-activated protein kinases (MAPK) pathway is hyperactive in head and neck cancer (HNC), inhibition of MEK1/2 in HNC patients has not shown clinically meaningful activity. Using pre-clinical HNC models, we demonstrated that treatment with the MEK1/2 blocker trametinib delays HNC initiation and progression by reducing tumor cell proliferation and enhancing the anti-tumor immunity of CD8+ T cells. Further activation of CD8+ T cells by supplementation with anti-programmed death-1 (PD-1) antibody eliminated tumors and induced an immune memory in the cured mice. Mechanistically, an early response to trametinib treatment sensitized tumors to PD-1-supplementation by attenuating the expression of tumor-derived colony-stimulating factor-1 (CSF-1), which reduced the abundance of two CSF-1R+CD11c+ myeloid-derived suppressor cell (MDSC) populations in the tumor microenvironment (TME). In contrast, prolonged treatment with trametinib abolished the anti-tumor activity of PD-1, because tumor cells undergoing the epithelial to mesenchymal transition (EMT) in response to trametinib restored CSF-1 expression and re-created an immune-suppressive TME. These findings provide the rationale for testing the trametinib/PD-1 combination in HNC and highlight the importance of sensitizing tumors to immunotherapies by using targeted therapies to interfere with the host-tumor interaction. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=199 SRC="FIGDIR/small/457244v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@1c65286org.highwire.dtl.DTLVardef@1541becorg.highwire.dtl.DTLVardef@f557f3org.highwire.dtl.DTLVardef@16134dc_HPS_FORMAT_FIGEXP M_FIG Graphical abstract C_FIG

cancer biology↗

bioRxiv2021

Paramecium Polycomb Repressive Complex 2 physically interacts with the small RNA binding PIWI protein to repress transposable elements

Polycomb Repressive Complex 2 (PRC2) maintains transcriptionally silent genes in a repressed state via deposition of histone H3 K27 trimethyl (me3) marks. PRC2 has also been implicated in silencing transposable elements (TEs) yet how PRC2 is targeted to TEs remains unclear. To address this question, we performed tandem affinity purification combined with mass spectrometry and identified proteins that physically interact with the Paramecium Enhancer-of-zeste Ezl1 enzyme, which catalyzes H3K9me3 and H3K27me3 deposition at TEs. We show that the Paramecium PRC2 core complex comprises four subunits, each required in vivo for catalytic activity. We also identify PRC2 cofactors, including the RNA interference (RNAi) effector Ptiwi09, which are necessary to target H3K9me3 and H3K27me3 to TEs. We find that the physical interaction between PRC2 and the RNAi pathway is mediated by a RING finger protein and that small RNA recruitment of PRC2 to TEs is analogous to the small RNA recruitment of H3K9 methylation SU(VAR)3-9 enzymes.

molecular biology↗

bioRxiv2020

Climbing up and down binding landscapes: a high-throughput study of mutational effects in homologous protein-protein complexes

Each protein-protein interaction (PPI) has evolved to possess binding affinity that is compatible with its cellular function. As such, cognate enzyme/inhibitor interactions frequently exhibit very high binding affinities, while structurally similar non-cognate PPIs possess substantially weaker binding affinities. To understand how slight differences in sequence and structure could lead to drastic changes in PPI binding free energy ({Delta}{Delta}Gbind), we study three homologous PPIs that span nine orders of magnitude in binding affinity and involve a serine protease interacting with an inhibitor BPTI. Using state-of-the-art methodology that combines protein randomization and affinity sorting coupled to next-generation sequencing and data normalization, we report quantitative binding landscapes consisting of {Delta}{Delta}Gbind values for the three PPIs, gleaned from tens of thousands of single and double mutations in the BPTI binding interface. We demonstrate that the three homologous PPIs possess drastically different binding landscapes and lie at different points in respect to the landscape maximum. Furthermore, the three PPIs demonstrate distinct patterns of coupling energies between two simultaneous mutations that depend not only on positions involved but also on the nature of the mutation. Interestingly, we find that in all three PPIs positive epistasis is frequently observed at hot-spot positions where mutations lead to loss of high affinity, while conversely negative epistasis is observed at cold-spot positions, where mutations lead to affinity enhancement. The new insights on PPI evolution revealed in this study will be invaluable in understanding evolution of other biological complexes and can greatly facilitate design of novel high-affinity protein inhibitors. SignificanceProtein-protein interactions (PPIs) have evolved to display binding affinities that can support their function. As such, cognate and non-cognate PPIs could be highly similar structurally but exhibit huge differences in binding affinities. To understand this phenomenon, we studied the effect of tens of thousands of single and double mutations on binding affinity of three homologous protease-inhibitor complexes. We show that binding landscapes of the three complexes are strikingly different and depend on the PPI evolutionary optimality. We observe different patterns of couplings between mutations for the three PPIs with negative and positive epistasis appearing most frequently at hot-spot and cold-spot positions, respectively. The evolutionary trends observed here are likely to be universal to all biological complexes in the cell.

biophysics↗

bioRxiv2020

The impact of water deficit and heat stress combination on the molecular response, physiology and seed production of soybean

A combination of drought and heat stress, occurring at the vegetative or reproductive growth phase of many different crops, can have a devastating impact on yield. In soybean (Glycine max), a considerable effort has been made to develop genotypes with enhanced yield production under conditions of drought or heat stress. However, how these genotypes perform in terms of growth, physiological responses and most importantly seed production, under conditions of drought and heat combination is mostly unknown. Here, we studied the impact of water deficit and heat stress combination on the physiology, seed production and yield per plant of two soybean genotypes, Magellan and Plant Introduction (PI) 548313, that differ in their reproductive responses to heat stress. Our findings reveal that although PI 548313 produced more seeds than Magellan under conditions of heat stress, under conditions of water deficit and heat stress combination its seed production decreased. Because number of flowers and pollen germination of PI 548313 remained high under heat or water deficit and heat combination, the reduced seed production exhibited by PI 548313 under the stress combination could be a result of processes that occur at the stigma, ovaries and/or other parts of the flower following pollen germination. HighlightTolerance to heat stress was found not to confer tolerance to a combination of water deficit and heat stress in soybean, highlighting the need for breeding strategies targeting the stress combination.

plant biology↗

bioRxiv2020

Online modulation of brain hemodynamics despite stereotyped running

Theta and gamma rhythms coordinate large cell assemblies during locomotion. Their spread across temporal and spatial scales makes them challenging to observe. Additionally, the metabolic cost of these oscillations and their contribution to neuroimaging signals remains elusive. To finely characterize neurovascular interactions in running rats, we monitored brain hemodynamics with functional ultrasound and hippocampal local field potentials in running rats. Theta rhythm and running speed were strongly coupled to brain hemodynamics in multiple structures, with delays ranging from 0.8 seconds to 1.8 seconds. Surprisingly, hemodynamics was also strongly modulated across trials within the same recording session: cortical hemodynamics sharply decreased after 5-10 runs, while hippocampal hemodynamics strongly and linearly potentiated, particularly in the CA regions. This effect occurred while running speed and theta activity remained constant, and was accompanied by increased power in hippocampal high-frequency oscillations (100-150 Hz). Our findings reveal distinct vascular subnetworks modulated across fast and slow timescales and suggest strong adaptation processes despite stereotyped behavior.

neuroscience↗

bioRxiv2019

A dynamic pattern of local auxin sources is required for root regeneration

Following removal of its stem cell niche, the root meristem can regenerate by recruitment of remnant cells from the stump. Regeneration is initiated by rapid accumulation of auxin near the injury site but the source of this auxin is unknown. Here, we show that auxin accumulation arises from the activity of multiple auxin biosynthetic sources that are newly specified near the cut site and that their continuous activity is required for the regeneration process. Auxin synthesis is highly localized and PIN-mediate transport is dispensable for auxin accumulation and tip regeneration. Roots lacking the activity of the regeneration competence factor ERF115, or that are dissected at a zone of low-regeneration potential, fail to activate local auxin sources. Remarkably, restoring auxin supply is sufficient to confer regeneration capacity to these recalcitrant tissues. We suggest that regeneration competence relies on the ability to specify new local auxin sources in a precise spatio-temporal pattern.

plant biology↗

bioRxiv2019

Regulation of perineuronal nets in the adult cortex by the electrical activity of parvalbumin interneurons

Perineuronal net (PNN) accumulation around parvalbumin-expressing (PV) inhibitory interneurons marks the closure of critical periods of high plasticity, whereas PNN removal reinstates juvenile plasticity in the adult cortex. Using targeted chemogenetic in vivo approaches in the adult mouse visual cortex, we found that transient electrical silencing of PV interneurons, directly or through inhibition of local excitatory neurons, induced PNN regression. Conversely, excitation of either neuron types did not reduce the PNN. We also observed that chemogenetically inhibited PV interneurons exhibited reduced PNN compared to their untransduced neighbors, and confirmed that single PV interneurons express multiple genes enabling cell-autonomous control of their own PNN density. Our results indicate that PNNs are dynamically regulated in the adult by PV neurons acting as sensors of their local microcircuit activities. PNN regulation provides individual PV neurons with an activity-dependent mechanism to control the local remodeling of adult cortical circuits.

neuroscience↗

bioRxiv2018

Total lung capacity without plethysmography

BackgroundAmong the most basic measures of respiratory function is the total lung capacity (TLC). TLC is the pulmonary gas volume at maximal lung inflation, which is the sum of the volume of gas that can be exhaled -the vital capacity (VC)- and the volume of gas that cannot -the residual volume (RV). Determination of VC requires only spirometry whereas determination of RV or TLC requires body plethysmography, gas dilution or washout, or thoracic imaging, each of which is more complex than spirometry, and none of which is suited to routine office practice, population screening, or community medicine. To fill this gap, we describe here a new approach to determine TLC without plethysmography.\n\nMethodsIn a heterogeneous population of 434 volunteers (265 male, 169 female; 201 healthy, 170 with airflow obstruction, and 63 with ventilatory restriction), we determined TLC in the standard fashion using conventional body plethysmography (TLCpleth). In the same individuals, we also determined TLC in a novel fashion using the MiniBox (TLCMB). To obtain TLCMB, population-based data from traditional spirometry together with flow-interruption transients were subjected to data mining and machine-learning to create for each individual subject an unbiased statistical determination of TLC.\n\nResultsFor the combined heterogeneous population, we found TLCpleth = 1.02TLCMB -0.091 L, adjusted r2=0.824. For the heterogeneous population as a whole, and for each subpopulation, TLCMB closely tracked TLCpleth. For 26 healthy subjects measured on different days, the coefficient of variation for repeated measurements in was 3.3% for TLCpleth versus 1.6% for TLCMB.\n\nConclusionsThese results establish the validity and potential utility of a new method for rapid, accurate, and repeatable determination of TLC in a heterogeneous patient population, but without the need of a plethysmograph.

bioengineering↗

bioRxiv2018

Desiccation-induced changes in recombination rate and interference in fruit flies: an evidence for fitness-dependent plasticity

Meiotic recombination is evolutionarily ambiguous, giving both benefits and costs to its bearers, with the resultant depending on a variety of conditions. The existing theoretical models explain why recombination does exist while the existence of its essential features remains underexplored. Here we test whether recombination plasticity and interference plasticity are fitness dependent. We compare effects of desiccation on recombination and interference in chromosome 3 in desiccation-sensitive and desiccation-tolerant Drosophila lines. We show that in desiccation-sensitive genotypes the treatment causes significant segment-specific increase in single- and double-crossover frequencies while in desiccation-tolerant genotypes changes are less pronounced, if seen at all. These results indicate that desiccation is a recombinogenic factor, and that desiccation-induced changes in both recombination and interference are fitness dependent, with a negative-feedback pattern implying the tendency of less fitted individuals to produce more variable progeny. Such dependence may play an important role in regulation of genetic variation in population experiencing environmental challenges.

genetics↗

bioRxiv2018

Cortical control of kinematic primitives in mice performing a hold-still-center-out reach task

Motor sequences are constructed from primitives, hypothesized building blocks of movement, but mechanisms of primitive generation remain unclear. Using automated homecage training and a novel forelimb sensor, we trained freely-moving mice to initiate forelimb sequences with clearly resolved submillimeter-scale micromovements followed by millimeter-scale reaches to learned spatial targets. Hundreds of thousands of trajectories were decomposed into millions of kinematic primitives, while closed-loop photoinhibition was used to test roles of motor cortical areas. Inactivation of contralateral motor cortex reduced primitive peak speed but, surprisingly, did not substantially affect primitive direction, initiation, termination, or complexity, resulting in isomorphic, spatially contracted trajectories that undershot targets. Our findings demonstrate separable loss of a single kinematic parameter, speed, and identify conditions where loss of cortical drive reduces the gain of motor primitives but does not affect their generation, timing or direction. The combination of high precision forelimb sensing with automated training and neural manipulation provides a system for studying how motor sequences are constructed from elemental building blocks.

neuroscience↗

bioRxiv2018

Anterior thalamic excitation and feed-forward inhibition of presubicular neurons projecting to medial entorhinal cortex

The presubiculum contains head direction cells that are crucial for spatial navigation. Here, we examined the connectivity and strengths of thalamic inputs to presubicular layer 3 neurons projecting to the medial entorhinal cortex in the mouse. We recorded pairs of projection neurons and interneurons while optogenetically stimulating afferent fibers from the anterior thalamic nuclei (ATN). Thalamic input differentially affects presubicular neurons: layer 3 pyramidal neurons and fast-spiking parvalbumin expressing (PV) interneurons are directly and monosynaptically activated, with depressing dynamics, while somatostatin (SST) expressing interneurons are indirectly excited, during repetitive ATN activity. This arrangement ensures that the thalamic excitation of layer 3 cells is often followed by disynaptic inhibition. Feed-forward inhibition is largely mediated by PV interneurons which have a high probability of connection to presubicular pyramidal cells. Our data point to a specific role of presubicular microcircuits in shaping thalamic head-direction signals transmitted to medial entorhinal cortex: Short-latency PV cell activation may enforce temporally precise head direction tuning during fast turns. However, depression at ATN-PV synapses during repeated activation tends to facilitate pyramidal cell firing when head direction is maintained. Operations performed in presubicular layer 3 circuits seem well-adapted for spatial fine-tuning of head direction signals sent to the medial entorhinal cortex.\n\nSIGNIFICANCE STATEMENTHow microcircuits participate in shaping neural inputs is crucial to understanding information processing in the brain. Here, we show how the presubiculum may process thalamic head directional information before transmitting it to the medial entorhinal cortex. Synaptic inputs from the anterior thalamic nuclei (ATN) excite layer 3 pyramidal cells (PC) and parvalbumin (PV) interneurons, which mediate disynaptic feed-forward inhibition. Somatostatin (SST) interneurons are excited indirectly. Presubicular circuits may switch between two regimes according to the angular velocity of head movements. During immobility, SST-PC interactions support maintained head directional firing with attractor-like dynamics. During rapid head turns, in contrast, PV mediated feed-forward inhibition acts to tune the head direction signal transmitted to medial entorhinal cortex.

neuroscience↗

bioRxiv2017

Converting a broad matrix metalloproteinase family inhibitor into a specific inhibitor of MMP-9 and MMP-14

MMP-14 and MMP-9 are two well established cancer targets for which no specific clinically relevant inhibitor is available. Using a powerful combination of computational design and yeast surface display technology, we engineered such an inhibitor starting from a non-specific MMP inhibitor, N-TIMP2. The engineered purified N-TIMP2 variants showed enhanced specificity towards MMP-14 and MMP-9 relative to a panel of off-target MMPs. MMP-specific N-TIMP2 sequence signatures were obtained that could be understood from the structural perspective of MMP/N-TIMP2 interactions. Our MMP-9 inhibitor exhibited 1000-fold preference for MMP-9 vs. MMP-14, which is likely to translate into significant differences under physiological conditions. Our results provide new insights regarding evolution of promiscuous proteins and optimization strategies for design of inhibitors with single-target specificities.

biophysics↗

bioRxiv2017

How activity dependent feedback inhibition may maintain head direction signals in mouse presubiculum

Orientation in space is represented in specialized brain circuits. Persistent head direction signals are transmitted from anterior thalamus to the presubiculum, but the identity of the presubicular target neurons, their connectivity and function in local microcircuits are unknown. Here we examine how thalamic afferents recruit presubicular principal neurons and Martinotti interneurons and the ensuing synaptic interactions between these cells. Pyramidal neuron activation of Martinotti cells in superficial layers is strongly facilitating such that high frequency head directional stimulation efficiently unmutes synaptic excitation. Martinotti cell feedback plays a dual role: precisely timed spikes may not inhibit the firing of in-tune head direction cells, while exerting lateral inhibition. Autonomous attractor dynamics emerge from a modeled network implementing wiring motifs and timing sensitive synaptic interactions in the pyramidal - Martinotti cell feedback loop. This inhibitory microcircuit is therefore tuned to refine and maintain head direction information in the presubiculum.

neuroscience↗