Biology subjects

McIntyre, A. B. R.

Publications and source records attributed to McIntyre, A. B. R..

7 recordsLinked to original sources

bioRxiv2026

Fibroblasts neurotrophin signaling sustains pathological vascular maturation in rheumatoid arthritis.

Treatment failures in rheumatoid arthritis (RA) leads to undesirable morbidity associated with immunosuppression. Recent studies of synovial tissue from refractory RA patients highlight the role of synovial fibroblasts and vascular endothelium in driving treatment failure. Utilizing high-dimensional spatial transcriptomics, we uncovered a crucial role for neurotrophin signaling in driving abnormal vascular maturation in RA synovia. Neurotrophins, including nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT3), induce differentiation of synovial fibroblasts into mural cells - pericytes and vascular smooth muscle cells. Mechanistically, NOTCH3 signaling activates a cascade of neurotrophin signaling through transcriptional induction of NGFR, a co-receptor for NGF. In RA synovial tissue explants, stimulation with NGF, BDNF, or NT3 leads to a dramatic increase in maturation of synovial tissue vasculature. Conversely, pharmacologic inhibition with neurotrophin inhibitors drastically abolished maturation of vascularization in RA synovial explants. Notably, the FDA-approved tropomyosin receptor kinase (TRK) inhibitors larotrectinib and entrectinib effectively reverse synovial vascular maturation in human RA tissue explants.Our findings suggest that fibroblast-derived neurotrophin signaling is a critical pathway in sustaining mature blood vessels in RA synovia, and that neurotrophin inhibitors reverse abnormal vascular maturation in RA. One Sentence SummaryIn rheumatoid arthritis, fibroblast neurotrophin signaling drives abnormal vascular maturation by inducing differentiation of fibroblasts into vascular mural cells.

molecular biology↗

bioRxiv2023

Phosphorylation-controlled cohesion of a nuclear condensate regulates mRNA retention

Nuclear speckles are membraneless organelles that associate with active transcription sites and participate in post-transcriptional mRNA processing. During the cell cycle, nuclear speckles dissolve following phosphorylation of their protein components. Here, we identify the PP1 family as the phosphatases that counteract kinase-mediated dissolution. PP1 overexpression increases speckle cohesion and leads to retention of polyadenylated RNA within speckles and the nucleus. Using APEX2 proximity labeling combined with RNA-sequencing, we characterized the relationship between the cohesion of nuclear speckles and the recruitment of specific RNAs. We find that many transcripts are preferentially enriched within nuclear speckles compared to the nucleoplasm, particularly chromatin- and nucleus-associated transcripts. While total polyadenylated RNA retention increased with nuclear speckle cohesion, the ratios of most mRNA species to each other were constant, indicating non-selective, or proportional, retention. We then explored whether nuclear speckle cohesion changes in response to environmental perturbations associated with changes in kinase or phosphatase activity. We found that cellular responses to heat shock, oxidative stress, and hypoxia include changes to the cohesion of nuclear speckles and mRNA retention. Our results demonstrate that tuning the material properties of nuclear speckles provides a mechanism for the acute control of mRNA localization.

cell biology↗

bioRxiv2020

Post-Transcriptional Regulation of Antiviral Gene Expression by N6-Methyladenosine

Type I interferons (IFN) induce hundreds of IFN-stimulated genes (ISGs) in response to viral infection. These ISGs require strict regulation for an efficient and controlled antiviral response, but post-transcriptional controls of these genes have not been well defined. Here, we identify a new role for the RNA base modification N6-methyladenosine (m6A) in the regulation of ISGs. Using ribosome profiling and quantitative mass spectrometry, coupled with m6A-immunoprecipitation and sequencing, we identified a subset of ISGs, including IFITM1, whose translation is enhanced by m6A and the m6A methyltransferase proteins METTL3 and METTL14. We further determined that the m6A reader YTHDF1 increases the expression of IFITM1 in an m6A binding-dependent manner. Importantly, we found that the m6A methyltransferase complex promotes the antiviral activity of type I IFN. Thus, these studies identify m6A as a post-transcriptional control of ISG translation during the type I IFN response for antiviral restriction.

immunology↗

bioRxiv2019

Direct RNA sequencing reveals m6A modifications on adenovirus RNA are necessary for efficient splicing

Adenovirus is a nuclear replicating DNA virus reliant on host RNA processing machinery. Processing and metabolism of cellular RNAs can be regulated by METTL3, which catalyzes the addition of N6-methyladenosine (m6A) to mRNAs. While m6A-modified adenoviral RNAs have been previously detected, the location and function of this mark within the infectious cycle is unknown. Since the complex adenovirus transcriptome includes overlapping spliced units that would impede accurate m6A mapping using short-read sequencing, we profiled m6A within the adenovirus transcriptome using a combination of meRIP-seq and direct RNA long-read sequencing to yield both nucleotide and transcript-resolved m6A detection. Although both early and late viral transcripts contain m6A, depletion of m6A writer METTL3 specifically impacts viral late transcripts by reducing their splicing efficiency. These data showcase a new technique for m6A discovery within individual transcripts at nucleotide resolution, and highlight the role of m6A in regulating splicing of a viral pathogen.

molecular biology↗

bioRxiv2019

Altered m6A modification of specific cellular transcripts affects Flaviviridae infection

The RNA modification N6-methyladenosine (m6A) can modulate mRNA fate and thus affect many biological processes. We analyzed m6A modification across the transcriptome following infection by dengue virus (DENV), Zika virus (ZIKV), West Nile virus (WNV), and hepatitis C virus (HCV). We found that infection by these viruses in the Flaviviridae family alters m6A modification of specific cellular transcripts, including RIOK3 and CIRBP. During viral infection, the addition of m6A to RIOK3 promotes its translation, while loss of m6A in CIRBP promotes alternative splicing. Importantly, we found that activation of innate immune sensing or the endoplasmic reticulum (ER) stress response by viral infection contributes to the changes in m6A modification in RIOK3 and CIRBP, respectively. Further, several transcripts with infection-altered m6A profiles, including RIOK3 and CIRBP, encode proteins that influence DENV, ZIKV, and HCV infection. Overall, this work reveals that cellular signaling pathways activated during viral infection lead to alterations in m6A modification of host mRNAs to regulate infection.

molecular biology↗

bioRxiv2019

Limits in the detection of m6A changes using MeRIP/m6A-seq

Many cellular mRNAs contain the modified base m6A, and recent studies have suggested that various stimuli can lead to changes in m6A. The most common method to map m6A and to predict changes in m6A between conditions is methylated RNA immunoprecipitation sequencing (MeRIP-seq), through which methylated regions are detected as peaks in transcript coverage from immunoprecipitated RNA relative to input RNA. Here, we generated replicate controls and reanalyzed published MeRIP-seq data to estimate reproducibility across experiments. We found that m6A peak overlap in mRNAs varies from [~]30 to 60% between studies, even in the same cell type. We then assessed statistical methods to detect changes in m6A peaks as distinct from changes in gene expression. However, from these published data sets, we detected few changes under most conditions and were unable to detect consistent changes across studies of similar stimuli. Overall, our work identifies limits to MeRIP-seq reproducibility in the detection both of peaks and of peak changes and proposes improved approaches for analysis of peak changes.

bioinformatics↗

bioRxiv2017

Nanopore detection of bacterial DNA base modifications

The common bacterial base modification N6-methyladenine (m6A) is involved in many pathways related to an organisms ability to survive and interact with its environment. Recent research has shown that nanopore sequencing can detect m5C with per-read accuracy of upwards of 80% but m6A with significantly lower accuracy. Here we use a binary classifier to improve m6A classification by marking adenines as methylated or unmethylated based on differences between measured and expected current values as each adenine travels through the nanopore. We also illustrate the importance of read quality for base modification detection and compare to PacBio methylation calls. With recent demonstrations of nanopore sequencing in Antarctica and onboard the International Space Station, the ability to reliably characterize m6A presents an opportunity to further examine the role of methylation in bacterial adaptation to extreme or very remote environments.

genomics↗