Biology subjects

Michel, K.

Publications and source records attributed to Michel, K..

15 recordsLinked to original sources

bioRxiv2026

Reversible Opto-Chemical activation of KRASG12V signaling with near single-cell precision

KRAS mutations drive some of the most lethal carcinomas, and genomic and inducible systems have established many of the cellular and tissue-level consequences. However, these approaches operate at the level of oncogene expression, allowing for cellular adaptation that masks the individual role of KRAS oncoprotein signaling. Here, we developed a reversible Opto-Chemical system to activate KRAS signaling by chemically translocating a cytosolic mutant KRASG12V G-domain to the plasma membrane upon light or small-molecule input. In MDCK cells, the G-domain plasma membrane recruitment activated downstream signaling and reduced collective migration. In mouse small Intestinal Organoids, G-domain recruitment promoted increased crypt size and number under Epidermal Growth Factor-deprived conditions. We further showed that the increased number of crypts depended on continuous KRASG12V signaling. Finally, under the same deprived conditions, localized activation in just one budding crypt promoted crypt formation compared to controls. This system decouples oncoprotein activity from oncogene expression, allowing to investigate the KRAS signaling contribution to early epithelial transformation.

Systems Biology↗

bioRxiv2026

Proliferation and differentiation in intestinal organoids as a balance of ligand-modulated the EGFR trafficking

Epidermal Growth factor (EGF) signaling is associated with (oncogenic) proliferation. Conversely, EGF-family ligands are able to trigger a differentiation program in cultured cells, an effect attributed to ligand affinity and EGFR phosphorylation. How EGF/EGFR driven proliferation-differentiation dynamics underlie tissue self-renewal has not been addressed. We show that culturing mouse small intestinal organoids (mSIOs) without EGF enhanced EGFR expression and base phosphorylation while maintaining a balanced development of proliferative crypts and differentiated villi. Addition of EGF or EREG triggers receptor endocytosis, reducing cell-surface and expression levels. While EGF promoted crypt proliferation, EREG promoted both proliferation and villus differentiation compared to untreated controls. Removal or re-introduction of EGF or EREG proved sufficient to induce development comparable to constant presence of ligands over 96h. Sub-saturating concentrations of EGF led to increased villus differentiation, resembling EREG treatments, suggesting that control over EGFR endocytic cycle ultimately regulates the balance of proliferation and differentiation in mSIOs SummaryExpression and signaling competency at the plasma membrane of EGFR drives crypt proliferation vs villus differentiation by medium ligand-composition, aiding mouse intestinal organoids self-renewal and regeneration.

developmental biology↗

bioRxiv2025

The potential of Clonostachys rosea as a biolarvicide against Anopheles gambiae

Mosquitoes vector multiple disease-causing pathogens detrimental to humans, imposing a high public health and economic burden. Chemical and biological larvicides are used in some settings to reduce mosquito populations as a supplement to adult population control. In this study, we assessed the larvicidal potential of a newly isolated strain of the hypocrealean fungus, Clonostachys rosea, by modifying a previously established laboratory-based experimental design. This design allowed us to distinguish between the larvicidal impact of the fungal culture supernatant and conidia. Our data show that C. rosea efficiently kills larvae of the African malaria mosquito, Anopheles gambiae, at an LC50 of 7.78 x 106 for culture supernatant and 7.13 x 107 conidia/mL for washed conidia. Fungal culture supernatant retained over 95% of its activity upon autoclaving, suggesting a temperature stable non-proteinaceous mycotoxin is responsible for larvicidal activity in the supernatant. Conidia were readily ingested, remained in the gut lumen, and did not germinate or translocate to the ectoperitrophic space nor the hemocoel. UV and heat-killed conidia retained larvicidal activity, demonstrating that no active secretion of toxins, fungal growth, or infection is required for killing mosquito larvae. However, autoclaving conidia abolished larvicidal activity, suggesting that larvae may not be killed via a mechanical mechanism, such as occlusion. Instead, our data demonstrate that C. rosea conidia kill An. gambiae larvae via the passive release of intracellular mycotoxins and have potential as a fungal biolarvicide to control An. gambiae populations. HighlightsO_LIClonostachys rosea is a potent larvicide of Anopheles gambiae mosquitoes. C_LIO_LILarvicidal activity from C. rosea culture supernatant is heat stable. C_LIO_LIC. rosea conidia are ingested but remain in the gut lumen and do not germinate. C_LIO_LIC. rosea conidia passively release toxins ultimately leading to larval death. C_LI

microbiology↗

bioRxiv2025

Larvicidal activity of Trichoderma atroviride (Hypocreales: Hypocreaceae) against Aedes albopictus (Diptera: Culicidae)

Larviciding is an important part of effective integrated mosquito management. However, growing resistance to chemical and bacterial-based insecticides requires biocontrol agents with novel modes of action. Entomopathogenic fungi are good candidates for larval control due to their capability to infect mosquito larvae and their production of larvicidal compounds. In this study, we isolated a strain of Trichoderma atroviride from Aedes albopictus larvae collected in Manhattan, KS, USA. We used a laboratory-based microcosm assay to expose L3 Ae. albopictus larvae to T. atroviride conidia and culture supernatant treatments. Larvae were monitored daily for survival and development to pupae and adults. In addition, adult survival was monitored for ten-days post pupation, and wing lengths were measured to assess mosquito size. Our results revealed that T. atroviride culture supernatant was a potent larvicide towards Ae. albopictus. However, conidia by themselves were not larvicidal, indicating the major mode of killing was through toxicity exerted by the culture supernatant. We further showed that larval exposure to T. atroviride supernatant delayed larval development to pupae. Sex-specific adult survival was not affected by larval exposure to T. atroviride. However, wing length of male and female mosquitoes were reduced, indicating a reduction in adult mosquito body size as compared to the control. Taken together, this study identifies the culture supernatant from a novel strain of T. atroviride as a potent larvicide of Ae. albopictus, potentially expanding our toolbox for biological control of mosquitoes.

microbiology↗

bioRxiv2024

The molecular Toll pathway repertoire in anopheline mosquitoes

Innate immunity in mosquitoes has received much attention due to its potential impact on vector competence for vector-borne disease pathogens, including malaria parasites. The nuclear factor (NF)-{kappa}B-dependent Toll pathway is a major regulator of innate immunity in insects. In mosquitoes, this pathway controls transcription of the majority of the known canonical humoral immune effectors, mediates anti-bacterial, anti-fungal and anti-viral immune responses, and contributes to malaria parasite killing. However, besides initial gene annotation of putative Toll pathway members and genetic analysis of the contribution of few key components to immunity, the molecular make-up and function of the Toll pathway in mosquitoes is largely unexplored. To facilitate functional analyses of the Toll pathway in mosquitoes, we report here manually annotated and refined gene models of Toll-like receptors and all putative components of the intracellular signal transduction cascade across 19 anopheline genomes, and in two culicine genomes. In addition, based on phylogenetic analyses, we identified differing levels of evolutionary constraint across the intracellular Toll pathway members, and identified a recent radiation of TOLL1/5 within the An. gambiae complex. Together, this study provides insight into the evolution of TLRs and the putative members of the intracellular signal transduction cascade within the genus Anopheles.

immunology↗

bioRxiv2024

The expanded immunoregulatory protease network in mosquitoes is governed by gene co-expression

Serine protease cascades regulate key innate immune responses. In mosquitoes, these cascades involve clip-domain serine proteases and their non-catalytic homologs (CLIPs), forming a complex network whose make-up and structural organization is not fully understood. This study assessed the impact of 85 CLIPs on humoral immunity in Anopheles gambiae. By coupling RNAi with assays measuring antimicrobial activity and melanization, we identified 27 CLIPs as immunoregulators that together form two distinct subnetworks. CLIPs regulating antimicrobial activity were found to control infection resistance, as knockdowns reduced bacterial load and improved survival. Furthermore, our analysis of CLIP gene expression unveiled a novel immunoregulatory mechanism reliant on protease baseline co-expression rather than infection-induced upregulation. These findings underscore that despite its complexity mosquito immune regulation may be targeted for malaria interventions.

immunology↗

bioRxiv2023

The transcriptional response in mosquitoes distinguishes between fungi and bacteria but not Gram types

Mosquitoes are prolific vectors of human pathogens; a clear and accurate understanding of the organization of their antimicrobial defenses is crucial for informing the development of transmission control strategies. The canonical infection response in insects, as described in the insect model Drosophila melanogaster, is pathogen type-dependent, with distinct stereotypical responses to Gram-negative bacteria and Gram-positive bacteria/fungi mediated by the activation of the Imd and Toll pathways, respectively. To determine whether this pathogen-specific discrimination is shared by mosquitoes, we used RNAseq to capture the genome-wide transcriptional response of Aedes aegypti and Anopheles gambiae (s.l.) to systemic infection with Gram-negative bacteria, Gram-positive bacteria, yeasts, and filamentous fungi, as well as challenge with heat-killed Gram-negative, Gram-positive, and fungal pathogens. From the resulting data, we found that Ae. aegypti and An. gambiae both mount a core response to all categories of infection, and this response is highly conserved between the two species with respect to both function and orthology. When we compared the transcriptomes of mosquitoes infected with different types of bacteria, we observed that the intensity of the transcriptional response was correlated with both the virulence and growth rate of the infecting pathogen. Exhaustive comparisons of the transcriptomes of Gram-negative-challenged versus Gram-positive-challenged mosquitoes yielded no difference in either species. In Ae. aegypti, however, we identified transcriptional signatures specific to bacterial infection and to fungal infection. The bacterial infection response was dominated by the expression of defensins and cecropins, while the fungal infection response included the disproportionate upregulation of an uncharacterized family of glycine-rich proteins. These signatures were also observed in Ae. aegypti challenged with heat-killed bacteria and fungi, indicating that this species can discriminate between molecular patterns that are specific to bacteria and to fungi.

immunology↗

bioRxiv2023

Insight into the structural hierarchy of the protease cascade that regulates the mosquito melanization response

Serine protease cascades regulate important insect immune responses, including melanization and Toll pathway activation. In the context of melanization, central components of these cascades are clip domain serine proteases (CLIPs) including the catalytic, clip domain serine proteases (cSPs) and their non-catalytic homologs (cSPHs). Here, we define partially the structural hierarchy of An. gambiae cSPs of the CLIPB family, central players in melanization, and characterize their relative contributions to bacterial melanization and to mosquito susceptibility to bacterial infections. Using in vivo genetic analysis we show that the protease cascade branches downstream of the cSPs CLIPB4 and CLIPB17 into two branches one converging on CLIPB10 and the second on CLIPB8. We also show that the contribution of key cSPHs to melanization in vivo in response to diverse microbial challenges is more significant than any of the individual cSPs, possibly due to partial functional redundancy among the latter. Interestingly, we show that the key cSPH CLIPA8 which is essential for the efficient activation cleavage of CLIPBs in vivo is efficiently cleaved itself by several CLIPBs in vitro, suggesting that cSPs and cSPHs regulate signal amplification and propagation in melanization cascades by providing positive reinforcement upstream and downstream of each other.

immunology↗

bioRxiv2023

CLIPB4 is a central node in the protease network that regulates humoral immunity in Anopheles gambiae mosquitoes

Insect humoral immune responses are regulated in part by protease cascades, whose components circulate as zymogens in the hemolymph. In mosquitoes, these cascades consist of clip domain serine proteases (cSPs) and/or their non-catalytic homologs (cSPHs), which form a complex network, whose molecular make-up is not fully understood. Using a systems biology approach, based on a co-expression network of gene family members that function in melanization and co-immunoprecipitation using the serine protease inhibitor (SRPN)2, a key negative regulator of the melanization response in mosquitoes, we identify the cSP CLIPB4 from the African malaria mosquito Anopheles gambiae as a central node in this protease network. CLIPB4 is tightly co-expressed with SRPN2 and forms protein complexes with SRPN2 in the hemolymph of immune-challenged female mosquitoes. Genetic and biochemical approaches validate our network analysis and show that CLIPB4 is required for melanization and antibacterial immunity, acting as a prophenoloxidase (proPO)-activating protease, which is inhibited by SRPN2. In addition, we provide novel insight into the structural organization of the cSP network in An. gambiae, by demonstrating that CLIPB4 is able to activate proCLIPB8, a cSP upstream of the proPO-activating protease CLIPB9. These data provide the first evidence that, in mosquitoes, cSPs provide branching points in immune protease networks and deliver positive reinforcement in proPO activation cascades.

immunology↗

bioRxiv2022

The potential of the Beauveria bassiana MHK isolate for mosquito larval control

The African malaria mosquito, Anopheles gambiae Giles (Diptera: Culicidae), and the Asian tiger mosquito, Aedes albopictus Skuse (Diptera: Culicidae) are of public health concern due to their ability to transmit disease-causing parasites and pathogens. Current mosquito control strategies to prevent vector-borne diseases rely mainly on the use of chemicals. However, insecticide resistance in mosquito populations necessitates alternative control measures, including biologicals such as entomopathogenic fungi. Here we report the impact of a Beauveria bassiana (Balsamo) Vuillemin (Hyprocreales: Cordycipitaeceae) isolate, MHK, isolated from field-collected Ae. albopictus larvae on mosquito survival and development. Larval infection bioassays using three B. bassiana conidial doses were performed on the second and third larval instars of An. gambiae and Ae. albopictus mosquitoes. Larvae were monitored daily for survival and development to pupae and adults. Our results show that B. bassiana MHK was more effective in killing An. gambiae than Ae. albopictus larvae. We further observed delays in development to pupae and adults in both mosquito species exposed the varying doses of B. bassiana as compared to the water control. In addition, larval exposure to B. bassiana reduced adult male and female survival in both mosquito species, further contributing to mosquito population control. Thus, this study identifies the locally isolated fungus, B. bassiana MHK, as a possible biological control agent of two mosquito species of public health concern, increasing the arsenal for integrated mosquito control.

microbiology↗

bioRxiv2022

RPTPγ is a redox-regulated suppressor of promigratory EGFR signaling

Spatially-organized interaction dynamics between proto-oncogenic epidermal growth factor receptor (EGFR) and protein tyrosine phosphatases (PTPs) determine EGFRs phosphorylation response to growth factors and thereby cellular behavior within developing tissues. We show here, that and how the coupling between EGFR and RPTP{gamma} activity leads to migratory signaling responses to very low, physiological growth factor stimuli while suppressing aberrant, spontaneous signaling. Single cell imaging of EGFR phosphorylation and PTP oxidation revealed that RPTP{gamma} fully suppresses spontaneous EGFR phosphorylation, while EGF-induced NADPH-oxidase activity enables promigratory signaling responses at the plasma membrane by H2O2-mediated oxidative inhibition of RPTP{gamma}s phosphatase activity. The EGF-dependent toggle switch dynamics between interacting EGFR monomers and RPTP{gamma} thereby enables autocatalytically amplified phosphorylation responses to very low, physiological, EGF levels even at sparse receptor expression. This signaling mechanism is distinct from the proliferative signaling stemming from liganded endosomal EGFR complexes at high growth factor concentrations. Accordingly, RPTP{gamma} knock-out results in spontaneous promigratory EGFR signaling but loss of proliferative signaling. We thereby provide evidence of RPTP{gamma}s suppressor function of oncogenic, promigratory EGFR-signaling from the plasma membrane.

systems biology↗

bioRxiv2022

A global Anopheles gambiae gene co-expression network constructed from hundreds of experimental conditions with missing values

Gene co-expression networks can be used to determine gene regulation and attribute gene function to biological processes. Different high throughput technologies, including one and two-channel microarrays and RNA-sequencing, allow evaluating thousands of gene expression data simultaneously, but these methodologies provide results that cannot be directly compared. Thus, it is complex to analyze coexpression relations between genes, especially when there are missing values arising for experimental reasons. Networks are a helpful tool for studying gene co-expression, where nodes represent genes and edges represent co-expression of pairs of genes. In this paper, we propose a method for constructing a gene co-expression network for the Anopheles gambiae transcriptome from 257 unique studies obtained with different methodologies and experimental designs. We introduce the sliding threshold approach to select node pairs with high Pearson correlation coefficients. The robustness of the method was verified by comparing edge weight distributions under random removal of conditions. The properties of the constructed network are studied in this paper, including node degree distribution, coreness, and community structure. The network core is largely comprised of genes that encode components of the mitochondrial respiratory chain and the ribosome, while different communities are enriched for genes involved in distinct biological processes. This suggests that the overall network structure is driven to maximize the integration of essential cellular functions, possibly allowing the flexibility to add novel functions.

bioinformatics↗

bioRxiv2020

Host-environment interplay shapes fungal diversity in mosquitoes

Mosquito larvae encounter diverse assemblages of bacteria (i.e. microbiota) and fungi (i.e. mycobiota) in the aquatic environments they develop in. However, while a number of studies have addressed the diversity and function of microbiota in mosquito life history, relatively little is known about mosquito-mycobiota interactions outside of several key fungal entomopathogens. In this study, we used high-throughput sequencing of ITS2 gene amplicons to provide the first simultaneous characterization of the mycobiota in field-collected Aedes albopictus larvae and their associated aquatic environments. Our results reveal unprecedented variation in mycobiota among adjacent but discrete larval breeding habitats. Our results also reveal distinct mycobiota assembly in the mosquito gut versus other tissues, with gut-associated fungal communities being most similar to those present in the environment where larvae feed. Altogether, our results identify the environment as the dominant factor shaping mosquito mycobiota with no evidence of environmental filtering of the gut mycobiota. These results also identify mosquito feeding behavior and fungal mode of nutrition as potential drivers of tissue-specific mycobiota assembly after environmental acquisition. IMPORTANCEThe Asian tiger mosquito, Aedes albopictus, is the dominant mosquito species in the USA and an important vector of arboviruses of major public health concern. One aspect of mosquito control to curb mosquito-borne diseases has been the use of biological control agents such as fungal entomopathogens. Recent studies also demonstrate the impact of mosquito-associated microbial communities on various mosquito traits, including vector competence. However, while much research attention has been dedicated to understanding the diversity and function of mosquito-associated bacterial communities, relatively little is known about mosquito-associated fungal communities. A better understanding of the factors that drive mycobiota diversity and assembly in mosquitoes will be essential for future efforts to target mosquito micro- and mycobiomes for mosquito and mosquito-borne disease control.

microbiology↗

bioRxiv2020

Neuron-class specific responses govern adaptive remodeling of myelination in the neocortex

Myelination plasticity plays a critical role in neurological function, including learning and memory. However, it is unknown whether this plasticity is enacted through uniform changes across all neuronal subtypes, or whether myelin dynamics vary between neuronal classes to enable fine-tuning of adaptive circuit responses. We performed in vivo two-photon imaging to investigate the dynamics of myelin sheaths along single axons of both excitatory callosal projection neurons and inhibitory parvalbumin+ interneurons in layer 2/3 of adult mouse visual cortex. We find that both neuron types show dynamic, homeostatic myelin remodeling under normal vision. However, monocular deprivation results in experience-dependent adaptive myelin remodeling only in parvalbumin+ interneurons, but not in callosal projection neurons. Monocular deprivation induces an initial increase in elongation events in myelin segments of parvalbumin+ interneurons, followed by a contraction phase affecting a separate cohort of segments. Sensory experience does not alter the generation rate of new myelinating oligodendrocytes, but can recruit pre-existing oligodendrocytes to generate new myelin sheaths. Parvalbumin+ interneurons also show a concomitant increase in axonal branch tip dynamics independent from myelination events. These findings suggest that adaptive myelination is part of a coordinated suite of circuit reconfiguration processes, and demonstrate that distinct classes of neocortical neurons individualize adaptive remodeling of their myelination profiles to diversify circuit tuning in response to sensory experience.

neuroscience↗

bioRxiv2019

Innate Antifungal Immune Receptor, Dectin-1, Undergoes Ligand-Induced Oligomerization with Highly Structured β-Glucans and at Fungal Cell Contact Sites

Dectin-1A is a C-type Lectin innate immunoreceptor that recognizes {beta}-(1,3;1,6)-glucan, a structural component of Candida species cell walls. The higher order structure of {beta}-glucans ranges from random coil to insoluble fiber due to varying degrees of tertiary (helical) and quaternary structure. Model Saccharomyces cerevisiae {beta}-glucans of medium and high molecular weight (MMW and HMW, respectively) are highly structured. In contrast, low MW glucan (LMW) is much less structured. Despite similar affinity for Dectin-1A, the ability of glucans to induce Dectin-1A mediated calcium influx and Syk phosphorylation positively correlates with their degree of higher order structure. Chemical denaturation and renaturation of MMW glucan showed that glucan structure determines agonistic potential, but not binding affinity, for Dectin-1A. We explored the role of glucan structure on Dectin-1A oligomerization, which is thought to be required for Dectin-1 signaling. Glucan signaling decreased Dectin-1A diffusion coefficient in inverse proportion to glucan structural content, which was consistent with Dectin-1A aggregation. Forster Resonance Energy Transfer (FRET) measurements revealed that molecular aggregation of Dectin-1 occurs in a manner dependent upon glucan higher order structure. Number and Brightness analysis specifically confirmed an increase in the Dectin-1A dimer and oligomer populations that is correlated with glucan structure content. Comparison of receptor modeling data with FRET measurements confirms that in resting cells, Dectin-1A is predominantly in a monomeric state. Super Resolution Microscopy revealed that glucan-stimulated Dectin-1 aggregates are very small (<15 nm) collections of a few engaged receptors. Finally, FRET measurements confirmed increased molecular aggregation of Dectin-1A at fungal particle contact sites in a manner that positively correlated with the degree of exposed glucan on the particle surface. These results indicate that Dectin-1A senses the solution conformation of {beta}-glucans through their varying ability to drive receptor dimer/oligomer formation and activation of membrane proximal signaling events.

immunology↗