Biology subjects

Medina, M.

Publications and source records attributed to Medina, M..

32 records · Page 2Linked to original sources

bioRxiv2022

Endosomal removal and disposal of dysfunctional, immunostimulatory mitochondrial DNA

Maternally inherited mitochondrial DNA (mtDNA) encodes essential subunits of the mitochondrial oxidative phosphorylation system, but is also a major damage-associated molecular pattern (DAMP) that engages innate immune sensors when released into the cytoplasm, outside of cells or into circulation1. This function of mtDNA contributes to antiviral resistance, but unfortunately also causes pathogenic inflammation in many disease contexts2. Cells experiencing mtDNA stress due to depletion of the mtDNA-packaging protein, Transcription Factor A, Mitochondrial (TFAM), or HSV-1 infection exhibit elongated mitochondria, mtDNA depletion, enlargement of nucleoids (mtDNA-protein complexes), and activation of cGAS/STING innate immune signaling via mtDNA released into the cytoplasm3. However, the relationships between altered mitochondrial dynamics and mtDNA-mediated activation of the cGAS-STING pathway remain unclear. Here, we show that entire enlarged nucleoids are released from mitochondria that remain bound to TFAM and colocalize with cGAS. These nucleoids arise at sites of mtDNA replication due to a block in mitochondrial fission at a stage when endoplasmic reticulum (ER) actin polymerization would normally commence, which we propose is a fission checkpoint to ensure that mtDNA has completed replication and is competent for segregation into daughter mitochondria. Released nucleoids also colocalize with the early endosomal marker RAB5 as well as the late endosomal marker RAB7 in TFAM-deficient cells and in response to mtDNA stress caused by the HSV-1 UL12.5 protein. Loss of RAB7 increases interferon stimulated gene (ISG) expression. Thus, we propose that defects in mtDNA replication and/or segregation enact a late mitochondrial fission checkpoint that, if persistent, leads to selective removal of dysfunctional nucleoids by a mitochondrial-endosomal pathway. Early steps in this pathway are prone to mtDNA release and cGAS-STING activation, but the immunostimulatory mtDNA is ultimately disposed of through a mechanism involving RAB7-containing late endosomes to prevent excessive innate immune signaling. This mtDNA quality control pathway might represent a therapeutic target to prevent mtDNA-mediated inflammation and associated pathology.

cell biology↗

bioRxiv2022

A meta-analysis of the stony coral tissue loss disease microbiome finds key bacteria in lesions and unaffected tissue of diseased colonies

Stony coral tissue loss disease (SCTLD) has been causing significant whole colony mortality on reefs in Florida and the Caribbean. The cause of SCTLD remains unknown, with limited concurrence of SCTLD-associated bacteria among studies. We conducted a meta-analysis of SSU 16S ribosomal RNA gene datasets generated by 16 field and laboratory SCTLD studies to find consistent bacteria associated with SCTLD across disease zones (vulnerable, endemic, and epidemic), coral species, coral compartments (mucus, tissue, and skeleton), and disease states (apparently healthy colony tissue [AH], and unaffected [DU] and lesion [DL] tissue from diseased colonies). We also evaluated bacteria in seawater and sediment, which may be sources of SCTLD transmission. Although AH colonies in endemic and epidemic zones harbor bacteria associated with SCTLD lesions, and aquaria and field samples had distinct microbial compositions, there were still clear differences in the microbial composition among AH, DU, and DL in the combined dataset. Alpha diversity between AH and DL was not different; however, DU showed increased alpha diversity compared to AH, indicating that, prior to lesion formation, corals may undergo a disturbance to the microbiome. This disturbance may be driven by Flavobacteriales, which were especially enriched in DU. While Rhodobacterales and Peptostreptococcales-Tissierellales were prominent in structuring microbial interactions in DL. Peptostreptococcales-Tissierellales specifically may contribute to lesion progression through an alpha-toxin. We provide a consensus of SCTLD-associated bacteria both prior to and during lesion progression and identify how these taxa vary across studies, coral species, coral compartments, seawater, and sediment.

microbiology↗

bioRxiv2022

Symbiosis-driven development in an early branching metazoan

Microbes can initiate developmental gene regulatory cascades in animals. The molecular mechanisms underlying microbe-induced animal development and the evolutionary steps to integrate microbial signals into regulatory programs remain poorly understood. In the upside-down jellyfish Cassiopea xamachana, a dinoflagellate endosymbiont initiates the life stage transition from the sessile polyp to the sexual medusa. We found that metabolic products derived from symbiont carotenoids may be important to initiate C. xamachana development, in addition to expression of conserved genes involved in medusa development of non-symbiotic jellyfish. We also revealed the transcription factor COUP is expressed during metamorphosis, potentially as a co-regulator of nuclear receptor RXR. These data suggest relatively few steps may be necessary to integrate symbiont signals into gene regulatory networks and cements the role of the symbiont as a key trigger for life history transition in C. xamachana.

developmental biology↗

bioRxiv2022

Comparative genomic insights into bacterial induction of larval settlement and metamorphosis in the upside-down jellyfish Cassiopea

Bacterial biofilm is crucial in inducing the larval transition from pelagic to benthic environments for marine organisms. Bacteria can therefore dictate species distribution and success of the individual. Despite the importance of marine bacteria to animal ecology, the identity of inductive microbes for many invertebrates are unknown. We isolated bacteria belonging to multiple phyla are capable of inducing settlement and metamorphosis in the upside-down jellyfish Cassiopea xamachana. The most inductive isolates belonged to the genus Pseudoalteromonas, a marine bacterium known to induce the pelago-benthic transition in other marine invertebrates. In sequencing the genome of the isolated Pseudoalteromonas and an inductive Vibrio, we found biosynthetic pathways previously implicated in larval settlement were absent in these Cassiopea inducing taxa. Comparative analysis of the Pseudoalteromonas and Vibrio revealed shared genes that could underlie the inductive capacity of these two bacteria. Thus, C. xamachana are capable of responding to multiple bacterial species, but they may be responding to a common cue produced by multiple taxa. These findings could provide hints to the ecological success of C. xamachana compared to sympatric congeneric species within mangrove environments and provide avenues to investigate the evolution of animal-microbe interactions.

microbiology↗

bioRxiv2022

A surface morphometrics toolkit to quantify organellar membrane ultrastructure using cryo-electron tomography

Cellular cryo-electron tomography (cryo-ET) enables 3-dimensional reconstructions of organelles in their native cellular environment at subnanometer resolution. However, quantifying ultrastructural features of pleomorphic organelles in three dimensions is challenging, as is defining the significance of observed changes induced by specific cellular perturbations. To address this challenge, we established a semi-automated workflow to segment organellar membranes and reconstruct their underlying surface geometry in cryo-ET. To complement this workflow, we developed an open source suite of ultrastructural quantifications, integrated into a single pipeline called the surface morphometrics toolkit. This toolkit allows detailed mapping of spacing, curvature, and orientation onto reconstructed membrane meshes, highlighting subtle organellar features that are challenging to detect in three dimensions and allowing for statistical comparison across many organelles. To demonstrate the advantages of this approach, we combine cryo-ET with cryo-fluorescence microscopy to correlate bulk mitochondrial network morphology (i.e., elongated versus fragmented) with membrane ultrastructure of individual mitochondria in the presence and absence of endoplasmic reticulum (ER) stress. Using our toolkit, we demonstrate ER stress promotes adaptive remodeling of ultrastructural features of mitochondria including spacing between the inner and outer membranes, local curvature of the inner membrane, and spacing between mitochondrial cristae. We show that differences in membrane ultrastructure correlate to mitochondrial network morphologies, suggesting that these two remodeling events are coupled. Our toolkit offers opportunities for quantifying changes in organellar architecture on a single-cell level using cryo-ET, opening new opportunities to define changes in ultrastructural features induced by diverse types of cellular perturbations.

cell biology↗

bioRxiv2022

APOE ε4 effects on hippocampal atrophy in the healthy elderly reflect future cognitive decline

The APOE {varepsilon}4 allele is the primary genetic risk factor for late onset Alzheimers disease (AD). A cardinal problem in determining APOE {varepsilon}4s effect on cognition and brain structure in older individuals is dissociating prodromal changes - linked to increased AD risk - from potential phenotypic differences. To address this, we used cognitive and neuroimaging data from a large cohort of cognitively normal 69-86 year-olds with up to 8 yearly follow-ups to investigate cross-sectional and longitudinal differences between APOE {varepsilon}3/{varepsilon}3 homozygotes and {varepsilon}3/{varepsilon}4 heterozygotes. Although we found a significant age-by-genotype interaction in right hippocampal volume, once our analyses were conditionalised by future diagnosis to account for prodromal mild cognitive impairment (MCI) and AD, this effect was no longer observed. Likewise, longitudinally, rate of hippocampal atrophy was determined not by genotype, but by future diagnosis. Thus, we provide direct evidence in support of the prodromal hypothesis of APOE {varepsilon}4 on brain structure.

neuroscience↗

bioRxiv2021

Linking photoacclimation responses and microbiome shifts between depth-segregated sibling species of reef corals

Metazoans host complex communities of microorganisms that include dinoflagellates, fungi, bacteria, archaea, and viruses. Interactions among members of these complex assemblages allow hosts to adjust their physiology and metabolism to cope with environmental variation and occupy different habitats. Here, using reciprocal transplantation across depths, we studied adaptive divergence in the Caribbean corals Orbicella annularis and O. franksi. When transplanted from deep to shallow, O. franksi experienced fast photoacclimation, low mortality, and maintained a consistent bacterial community. In contrast, O. annularis experienced higher mortality, and limited photoacclimation when transplanted from shallow to deep. The photophysiological collapse of O. annularis in the deep environment was associated with an increased microbiome variability and reduction of some bacterial taxa. Differences in the symbiotic algal community were more pronounced between coral species than between depths. Our study suggests that these sibling species are adapted to distinctive light environments partially driven by the algae photoacclimation capacity and the microbiome robustness, highlighting the importance of niche specialization in symbiotic corals for the maintenance of species diversity. Our findings have implications for the management of these threatened Caribbean corals and the effectiveness of coral reef restoration efforts.

microbiology↗

bioRxiv2021

Humoral immune responses against seasonal coronaviruses predict efficiency of SARS-CoV-2 spike targeting, FcγR activation, and corresponding COVID-19 disease severity

Despite SARS-CoV-2 being a "novel" coronavirus, several studies suggest that detection of anti-spike IgG early in infection may be attributable to the amplification of humoral memory responses against seasonal hCoVs in severe COVID-19 patients. In this study, we examined this concept by characterizing anti-spike IgG from a cohort of non-hospitalized convalescent individuals with a spectrum of COVID-19 severity. We observed that anti-spike IgG levels positively correlated with disease severity, higher IgG cross-reactivity against betacoronaviruses (SARS-CoV-1 and OC43), and higher levels of proinflammatory Fc gamma receptor 2a and 3a (Fc{gamma}R2a & Fc{gamma}R3a) activation. In examining the levels of IgG targeting betacoronavirus conserved and immunodominant epitopes versus disease severity, we observed a positive correlation with the levels of IgG targeting the conserved S2FP region, and an inverse correlation with two conserved epitopes around the heptad repeat (HR) 2 region. In comparing the levels of IgG targeting non-conserved epitopes, we observed that only one of three non-conserved immunodominant epitopes correlated with disease severity. Notably, the levels of IgG targeting the receptor binding domain (RBD) were inversely correlated with severity. Importantly, targeting of the RBD and HR2 regions have both been shown to mediate SARS-CoV-2 neutralization. These findings show that, aside from antibody (Ab) targeting of the RBD region, humoral memory responses against seasonal betacoronaviruses are potentially an important factor in dictating COVID-19 severity, with anti-HR2-dominant Ab profiles representing protective memory responses, while an anti-S2FP dominant Ab profiles indicate deleterious recall responses. Though these profiles are masked in whole antigen profiling, these analyses suggest that distinct Ab memory responses are detectable with epitope targeting analysis. These findings have important implications for predicting severity of SARS-CoV-2 infections (primary and reinfections), and may predict vaccine efficacy in subpopulations with different dominant antibody epitope profiles.

microbiology↗

bioRxiv2021

Physiological and ecological consequences of the water optical properties degradation on reef corals

Degradation of water optical properties due to anthropogenic disturbances is a common phenomenon in coastal waters globally. Although this condition is associated with multiple drivers that affect corals health in multiple ways, its effect on light availability and photosynthetic energy acquisition has been largely neglected. Here, we describe how declining the water optical quality in a coastal reef exposed to a turbid plume of water originating from a man-made channel compromise the functionality of the keystone coral species Orbicella faveolata. We found highly variable water optical conditions with significant effects on the light quantity and quality available for corals. Reduction of light penetration into the water column elicits the development of low-light phenotypes close to theoretical limits of photoacclimation despite their occurrence at shallow depths. Predicted photosynthetic energy depletion with increasing depth is associated with patterns of colony mortality and contraction of the habitable space for the population. A numerical model illustrates the potential effect the progressive degradation of water optical properties on the gradual mortality and population decline of O. faveolata. Our findings suggest that preserving the water optical properties seeking to maximize light penetration into the water column may have an extraordinary impact on coral reefs conservation, mostly toward the deeper portions of reefs.

physiology↗

bioRxiv2021

A metagenomic-based stufy of two sites from the Barbadian reef system

We study the microbiome of sea water collected from two locations of the Barbadian coral reefs. The two sites differ in several environmental and ecological variables including their endogenous benthic community in addition to their proximity to urban development and runoffs from inland watersheds. The composition of the microbial community was estimated using whole genome DNA shotgun sequencing. Although both sites exhibit a similar degree of richness, the less urbanized site (Maycocks reef at Hangmans Bay) is strongly concentrated with phototrophs. In comparison, the more urbanized location (Bellairs Research Institute) is enriched for copiotrophs, macroalgal symbionts and marine-related disease-bearing organisms from taxa scattered across the tree of life. Overall, our samples and associated measurements of chemical and environmental qualities of the water are in line with previous marine microbiome profiles of warm ocean surface waters. This suggests our approach captures salient information regarding the state of each coral reef site and sets the stage for larger longitudinal studies of coral reef dynamics in Barbados.

microbiology↗

bioRxiv2020

Predicting the future development of mild cognitive impairment in the cognitively healthy elderly

Identifying measures that predict future cognitive impairment in healthy individuals is necessary to inform treatment strategies for candidate dementia-preventative and modifying interventions. Here, we derive such measures by studying "converters" who transitioned from cognitively normal at baseline to mild-cognitive impairment (MCI) in a longitudinal study of 1213 elderly participants. We first establish reduced grey matter density (GMD) in left entorhinal cortex (EC) as a biomarker for impending cognitive decline in healthy individuals, employing a matched sampling control for several dementia risk-factors, thereby mitigating the potential effects of bias on our statistical tests. Next, we determine the predictive performance of baseline demographic, genetic, neuropsychological and MRI measures by entering these variables into an elastic net-regularized classifier. Our trained statistical model classified converters and controls with validation Area-Under-the-Curve>0.9, identifying only delayed verbal memory and left EC GMD as relevant predictors for classification. This performance was maintained on test classification of out-of-sample converters and controls. Our results suggest a parsimonious but powerful predictive model for MCI development in the cognitively healthy elderly.

neuroscience↗

bioRxiv2020

Genomic adaptations to an endolithic lifestyle in the coral-associated alga Ostreobium

The green alga Ostreobium is an important coral holobiont member, playing key roles in skeletal decalcification and providing photosynthate to bleached corals that have lost their dinoflagellate endosymbionts. Ostreobium lives in the corals skeleton, a low-light environment with variable pH and O{square} availability. We present the Ostreobium nuclear genome and a metatranscriptomic analysis of healthy and bleached corals to improve our understanding of Ostreobiums adaptations to its extreme environment and its roles as a coral holobiont member. The Ostreobium genome has 10,663 predicted protein-coding genes and shows adaptations for life in low and variable light conditions and other stressors in the endolithic environment. This alga presents a rich repertoire of light-harvesting complex proteins but lacks many genes for photoprotection and photoreceptors. It also has a large arsenal of genes for oxidative stress response. An expansion of extracellular peptidases suggests that Ostreobium may supplement its energy needs by feeding on the organic skeletal matrix, and a diverse set of fermentation pathways allow it to live in the anoxic skeleton at night. Ostreobium depends on other holobiont members for vitamin B12, and our metatranscriptomes identify potential bacterial sources. Metatranscriptomes showed Ostreobium becoming a dominant agent of photosynthesis in bleached corals and provided evidence for variable responses among coral samples and different Ostreobium genotypes. Our work provides a comprehensive understanding of the adaptations of Ostreobium to its extreme environment and an important genomic resource to improve our comprehension of coral holobiont resilience, bleaching and recovery.

genomics↗

bioRxiv2020

In silico discovery and biological validation of ligands of FAD synthase, a promising new antimicrobial target

New treatments for diseases caused by antimicrobial-resistant microorganisms can be developed by identifying unexplored therapeutic targets and by designing efficient drug screening protocols. In this study, we have screened a library of compounds to find ligands for the flavin-adenine dinucleotide synthase (FADS) -a potential target for drug design against tuberculosis and pneumonia- by implementing a new and efficient virtual screening protocol. The protocol has been developed for the in silico search of ligands of unexplored therapeutic targets, for which limited information about ligands or ligand-receptor structures is available. It implements an integrative funnel-like strategy with filtering layers that increase in computational accuracy. The protocol starts with a pharmacophore-based virtual screening strategy that uses ligand-free receptor conformations from molecular dynamics (MD) simulations. Then, it performs a molecular docking stage using several docking programs and an exponential consensus ranking strategy. The last filter, samples the conformations of compounds bound to the target using MD simulations. The MD conformations are scored using several traditional scoring functions in combination with a newly-proposed score that takes into account the fluctuations of the molecule with a Morse-based potential. The protocol was optimized and validated using a compound library with known ligands of the Corynebacterium ammoniagenes FADS. Then, it was used to find new FADS ligands from a compound library of 14,000 molecules. A small set of 17 in silico filtered molecules were tested experimentally. We identified five inhibitors of the activity of the flavin adenylyl transferase mononucleotide of the FADS, and some of them were able to inhibit growth of three bacterial species: Corynebacterium ammoniagenes, Mycobacterium tuberculosis, and Streptococcus pneumoniae, where the former two are human pathogens. Overall, the results show that the integrative VS protocol is a cost-effective solution for the discovery of ligands of unexplored therapeutic targets. Author summaryDeveloping cures for antimicrobial-resistant microorganisms is a pressing necessity. Addressing this problem requires the discovery of novel therapeutic targets -for example, bacterial proteins with no human homologues- and the development of cost-effective drug screening protocols. In this work, we tackled the problem on both sides. We developed an efficient and successful integrative computational protocol for screening inhibitory-molecules for unexplored targets. We used it to discover five novel inhibitors of flavin-adenine dinucleotide synthase (FADS), a promising protein target of pathogens causing tuberculosis and pneumonia.

biophysics↗

bioRxiv2020

Doxycycline Significantly Enhances Induction of iPSCs to Endoderm by Enhancing survival via AKT Phosphorylation

Induced pluripotent stem cells (iPSCs) provide an important tool for the generation patient-derived cells including hepatocyte-like cells via developmental cues through an endoderm intermediate. However, most iPSCs fail to differentiate into endoderm, with induction resulting in apoptosis. To address this issue, we built upon published methods to develop an improved protocol with our discovery that doxycycline dramatically enhances the iPSC to endoderm differentiation efficiency by inhibiting apoptosis and promoting proliferation via the AKT pathway. We tested this new protocol in more than 70 iPSC lines with consistent formation of complete sheets of endoderm in 90%. Endoderm generated by our method achieves similar transcriptomic profiles, including FOXA2, HNF1{beta}, CXCR4, and SOX17 positive cells, and the ability to be further differentiated. Furthermore this method achieves a four-fold increase in endoderm cell number and will accelerate studies of human diseases in vitro and facilitate the expansion of iPSC-derived cells for transplantation studies.

developmental biology↗