Biology subjects

Baum, R.

Publications and source records attributed to Baum, R..

8 recordsLinked to original sources

bioRxiv2025

Activation of IL-17+ ILC subsets in IL-18R-deficient mice during fungal allergen exposure

Group 2 innate lymphoid cells (ILC2s) are critical players during type 2 inflammation present in most forms of asthma. ILC2s are tissue-resident cells that produce cytokines IL-5 and IL-13 critical to eosinophilic airway inflammation, mucus production, remodeling, and hyperresponsiveness. Though each ILC subset (ILC1s, ILC2s, ILC3s) is identified by specific transcription factors, cell surface receptors and cytokine profiles, functional plasticity between ILC subtypes occurs in various contexts. IL-18/IL-18R loci SNPs are linked to asthma in multiple genome-wide association studies and IL-18 has been shown to promote plasticity in ILC2s. Despite this, little is known about the in vivo role of IL-18/IL-18R on ILC2 responses in the lung. Within hours after mice were exposed to the fungal allergy Alternaria alternata, airway levels of IL-18 and IL-18 receptor expression increased on ST2+ ILCs. Single-cell RNA sequencing of lung cells from Alternaria-challenged mice showed that Il18 was largely expressed by alveolar macrophages, while IL-18R was highly expressed in IL-13+ILC2s. Utilizing IL-18 receptor knock-out mice (IL- 18R-/-), we observed increases in IL-17A production from both ST2+ and ST2-negative ILCs compared to control mice. We further observed an early increase in dual production of IL-5 and IL-17A in ST2+ ILCs followed by enhanced lung eosinophilia in the absence of IL-18R. Together, our findings suggest that IL-18 signaling prevents IL-17A production from ILC2s and subsequent eosinophilia in vivo. A further understanding of the regulation of ILC plasticity may lead to novel therapeutic targets in the treatment of ILC-driven asthma.

immunology↗

bioRxiv2024

Mapping the Genetic Architecture of the Adaptive Integrated Stress Response in S. cerevisiae

The integrated stress response (ISR) is a conserved eukaryotic signaling pathway that responds to diverse stress stimuli to restore proteostasis. The strength and speed of ISR activation must be tuned properly to allow protein synthesis while maintaining proteostasis. Here, we describe how genetic perturbations change the dynamics of the ISR in budding yeast. We treated ISR dynamics, comprising timecourses of ISR activity across different levels of stress, as a holistic phenotype. We profiled changes in ISR dynamics across thousands of genetic perturbations in parallel using CRISPR interference with barcoded expression reporter sequencing (CiBER-seq). We treated cells with sulfometuron methyl, a titratable inhibitor of branched-amino acid synthesis, and measured expression of an ISR reporter. Perturbations to translation such as depletion of aminoacyl-tRNA synthetases or tRNA biogenesis factors reduced cell growth and caused a strikingly proportionate activation of the ISR activation. In contrast, impaired ribosome biogenesis reduced basal ISR activity and weakened ISR dynamics. Reduced ribosome capacity may lower the demand for amino acids and thereby explain these changes. Our work illustrates how CiBER-seq enables high-throughput measurements of complex and dynamic phenotypes that shed light on adaptive and homeostatic mechanisms.

genomics↗

bioRxiv2022

A germline-targeting chimpanzee SIV envelope glycoprotein elicits a new class of V2-apex directed cross-neutralizing antibodies

HIV-1 and its SIV precursors share a broadly neutralizing antibody (bNAb) epitope in variable loop 2 (V2) at the envelope glycoprotein (Env) trimer apex. Here, we tested the immunogenicity of germline-targeting versions of a chimpanzee SIV (SIVcpz) Env in human V2-apex bNAb heavy-chain precursor-expressing knock-in mice and as chimeric simian-chimpanzee immunodeficiency viruses (SCIVs) in rhesus macaques (RMs). Trimer immunization of knock-in mice induced V2-directed NAbs, indicating activation of V2-apex bNAb precursor-expressing mouse B cells. SCIV infection of RMs elicited high-titer viremia, potent autologous tier 2 neutralizing antibodies, and rapid sequence escape in the canonical V2-apex epitope. Six of seven animals also developed low-titer heterologous plasma breadth that mapped to the V2-apex. Antibody cloning from two of these identified multiple expanded lineages with long heavy chain third complementarity determining regions that cross-neutralized as many as 7 of 19 primary HIV-1 strains, but with low potency. Negative stain electron microscopy (NSEM) of members of the two most cross-reactive lineages confirmed V2 targeting but identified an angle of approach distinct from prototypical V2-apex bNAbs, with antibody binding either requiring or inducing an occluded-open trimer. Probing with conformation-sensitive, non-neutralizing antibodies revealed that SCIV-expressed Envs as well as some primary HIV-1 Envs adopted a more open conformation, thereby exposing a conserved V2 epitope that is occluded in closed SIVcpz and HIV-1 Env trimers. These results expand the spectrum of V2-apex targeted antibodies that can contribute to neutralization breadth and identify novel SIV Env platforms for further development as germline-targeting and immunofocusing immunogens. One sentence summaryA cryptic V2 epitope in occluded-open HIV and SIV Env trimers is the target of a new class of V2-directed cross-neutralizing antibodies.

immunology↗

bioRxiv2022

GJA1-20k, an internally translated isoform of Connexin 43, is an actin capping protein

Previously, we identified that GJA1-20k, an internally translated isoform of Connexin 43, mediates an actin-dependent protective form of mitochondrial fission (Shimura, Nuebel et al. 2021). We found that when GJA1-20k is present, bands of actin surround mitochondria at locations enriched with GJA1-20k, inducing mitochondrial fission which generates less oxygen free radicals, protecting hearts subjected to ischemia-reperfusion injury. Here, we report that GJA1-20k is a direct actin binding protein and thereby identify the mechanism by which GJA1-20k is able to recruit and stabilize actin filaments around the mitochondria. Surprisingly, GJA1-20k functions as a canonical actin capping protein, producing both truncated actin puncta and stabilized actin filaments. GJA1-20k contains an RPEL-like actin binding motif, and we confirm with both computational modeling and biochemistry, that this domain is crucial for actin capping. The actin capping functionality of GJA1-20k adds GJA1-20k to the family of proteins that regulate actin dynamics. As a stress responsive protein, GJA1-20k can help explain cytoskeletal dependent responses to cellular stress, from delivery of channels to affecting mitochondrial size and function.

cell biology↗

bioRxiv2021

LncRNA Miat and Metadhedrin maintain a treatment resistant stem-like niche of medulloblastoma cells

Medulloblastoma is a pediatric brain tumor arising from the cerebellum and brainstem that is curable with surgery and aggressive cytotoxic therapy including craniospinal irradiation and chemotherapy. Preclinical models indicate that a small pool of de-differentiated, stem cell-like medulloblastoma cells are resistant to cytotoxic treatment and contribute to medulloblastoma relapse after aggressive therapy. Here, we identify Miat as a Shh and Myc regulated long noncoding RNA (lncRNA) that is required for maintenance of a treatment-resistant medulloblastoma stem-like phenotype. Loss of Miat delays medulloblastoma formation in genetically defined mouse models of Shh medulloblastoma and enforces differentiation of tumorigenic stem-like medulloblastoma cells into a non-tumorigenic cell state. Miat facilitates treatment resistance in part by downregulating p53 signaling and impairing radiation induced cell death in stem-like MB cells, which can be reversed by therapeutic inhibition of Miat with antisense oligonucleotides. The RNA binding protein Metadhedrin (Mtdh), which is associated with resistance to cytotoxic therapy in numerous types of cancer, co-localizes with Miat in stem-like medulloblastoma cells. Further, loss of Mtdh activates p53 signaling and reduces tumorigenicity in stem-like medulloblastoma cells. Taken together, these data reveal a critical role for the lncRNA Miat in sustaining a treatment resistant pool of tumorigenic stem-like medulloblastoma cells.

cancer biology↗

bioRxiv2021

Protective mitochondrial fission induced by stress responsive protein GJA1-20k

The Connexin43 gap junction gene GJA1 has one coding exon, but its mRNA undergoes internal translation to generate N-terminal truncated isoforms of Connexin43 with the predominant isoform being only 20 kDa in size (GJA1-20k). Endogenous GJA1-20k protein is not membrane bound, and has been found to increase in response to ischemic stress, localize to mitochondria, and mimic ischemic preconditioning protection in the heart. However, it is not known how GJA1-20k benefits mitochondria to provide this protection. Here, we identify that GJA1-20k polymerizes actin around mitochondria which induces focal constriction sites. Mitochondrial fission events occur within about 45 seconds of GJA1-20k recruitment of actin. Interestingly, GJA1-20k mediated fission is independent of canonical Dynamin Related Protein 1 (DRP1). We find that GJA1-20k induced smaller mitochondria have decreased reactive oxygen species (ROS) generation and, in hearts, provide potent protection against ischemia-reperfusion injury. The results indicate that stress responsive internally translated GJA1-20k stabilizes polymerized actin filaments to stimulate non-canonical mitochondrial fission which limits ischemic-reperfusion induced myocardial infarction.

cell biology↗

bioRxiv2020

A genome-scale CRISPR interference guide library enables comprehensive phenotypic profiling in yeast

CRISPR/Cas9-mediated transcriptional interference (CRISPRi) enables programmable gene knock-down, yielding interpretable loss-of-function phenotypes for nearly any gene. Effective, inducible CRISPRi has been demonstrated in budding yeast, but no genome-scale guide libraries have been reported. We present a comprehensive yeast CRISPRi library, based on empirical design rules, containing 10 distinct guides for most genes. Competitive growth after pooled transformation revealed strong fitness defects for most essential genes, verifying that the library provides comprehensive genome coverage. We used the relative growth defects caused by different guides targeting essential genes to further refine yeast CRISPRi design rules. In order to obtain more accurate and robust guide abundance measurements in pooled screens, we link guides with random nucleotide barcodes and carry out linear amplification by in vitro transcription. Taken together, we demonstrate a broadly useful platform for comprehensive, high-precision CRISPRi screening in yeast.

genomics↗

bioRxiv2019

Muntjac chromosome evolution and architecture

AbstractDespite their recent divergence, muntjac deer show striking karyotype differences. Here we describe new chromosome-scale genome assemblies for the Chinese and Indian muntjacs, Muntiacus reevesi (2n=46) and Muntiacus muntjak (2n=6/7), and analyze their evolution and architecture. We identified six fusion events shared by both species relative to the cervid ancestor and therefore present in the muntjac common ancestor, six fusion events unique to the M. reevesi lineage, and twenty-six fusion events unique to the M. muntjak lineage. One of these M. muntjak fusions reverses an earlier fission in the cervid lineage. Although comparative Hi-C analysis revealed differences in long-range genome contacts and A/B compartment structures, we discovered widespread conservation of local chromatin contacts between the muntjacs, even near the fusion sites. A small number of genes involved in chromosome maintenance show evidence for rapid evolution, possibly associated with the dramatic changes in karyotype. Analysis of muntjac genomes reveals new insights into this unique case of rapid karyotype evolution and the resulting biological variation.

genomics↗