Biology subjects

Jacobson, K.

Publications and source records attributed to Jacobson, K..

7 recordsLinked to original sources

bioRxiv2024

Single-dose avian influenza A(H5N1) Clade 2.3.4.4b hemagglutinin-Matrix-M nanoparticle vaccine induces neutralizing responses in nonhuman primates

With the recent rise in cases of highly pathogenic avian influenza (HPAI) A(H5N1) clade 2.3.4.4b infection in humans and animals, there is an associated increase in the risk of human-to-human transmission. In this study, we characterize recombinant A(H5N1) A/American Wigeon/South Carolina/22/000345-001/2021 (A/AW/SC/2021) clade 2.3.4.4b vaccine. Purified recombinant A/AW/SC/2021 HA trimers upon formulation with Matrix-M adjuvant, saponin-cholesterol-phospholipid icosahedral particles, non-covalently anchored to the vertices of the Matrix-M forming A(H5N1) HA-Matrix-M nanoparticles (H5-MNPs). In naive mice, two intranasal (IN) or intramuscular (IM) doses of A/AW/SC/2021 H5-MNP vaccine induced robust antibody- and cell-mediated immune responses, including neutralizing antibodies against A(H5N1). In non-human primates (NHPs) primed with seasonal influenza vaccine, a single IM or IN dose of the A/AW/SC/2021 H5-MNP vaccine induced geometric mean serum A(H5N1) clade 2.3.4.4b pseudovirus neutralizing titers of 1:1160 and 1:54, respectively; above the generally accepted seroconverting neutralizing titer of 1:40. Immunization with H5-MNP vaccine induced antibody responses against conserved epitopes in the A(H5N1) HA stem, vestigial esterase subdomain, and receptor binding site. This novel A(H5N1) H5-MNP IN and IM vaccine was immunogenic in rodents and NHPs as a potential A(H5N1) pandemic single-dose vaccine.

microbiology↗

bioRxiv2022

COP9 signalosome component CSN-5 stabilizes PUF proteins FBF-1 and FBF-2 in C. elegans germline stem cells

RNA-binding proteins FBF-1 and FBF-2 (FBFs) are required for germline stem cell maintenance and the sperm/oocyte switch in Caenorhabditis elegans, though the mechanisms controlling FBF protein levels remain unknown. We identified an interaction between both FBFs and CSN-5, a component of the COP9 (constitutive photomorphogenesis 9) signalosome. Here, we find that the MPN (Mpr1/Pad1 N terminal) metalloprotease domain of CSN-5 interacts with the PUF (Pumilio and FBF) RNA-binding domain of FBFs and the interaction is conserved for human homologs PUM1 and CSN5. The interaction between FBF-2 and CSN-5 can be detected in vivo by proximity ligation. csn-5 mutation results in destabilization of FBF proteins, a decrease in the numbers of germline stem cells, and disruption of the switch from spermatogenesis to oogenesis. The loss of csn-5 does not decrease the levels of a related PUF protein PUF-3 and csn-5(lf) phenotype is not enhanced by fbf-1/2 depletion, suggesting that the effect is specific to FBFs. The effect of csn-5 on germline sex determination is largely independent of the COP9 signalosome and is cell autonomous. Surprisingly, regulation of FBF protein levels involves a combination of COP9-dependent and -independent mechanisms differentially affecting FBF-1 and FBF-2. This work supports a previously unappreciated role for CSN-5 in stabilization of germline stem cell regulatory proteins FBF-1 and FBF-2. Author SummaryGerm cell development and reproductive success in the nematode C. elegans rely on the function of germline stem cells. Continued maintenance of these cells is supported by the activity of conserved RNA-binding proteins FBF-1 and FBF-2 (FBFs). However, it is unknown how FBF protein levels are regulated. Here, we identify a direct interaction between FBFs and CSN-5, a component of the COP9 signalosome best known for its role in regulating protein degradation. We find that CSN-5 promotes FBF stability and allows for accumulation of steady-state protein levels, thereby promoting FBF function. In csn-5 mutants, we find a significant reduction of FBF proteins, decrease of stem cells, and failure to promote oogenesis consistent with compromised FBF function. Furthermore, CSN-5 contributes to FBF protein stability through two mechanisms. This work demonstrates a previously unappreciated role for CSN-5 in stabilization of FBF proteins. Based on our finding that the FBF/CSN-5 interaction is conserved and detectable between homologous human proteins, we speculate this relationship might be relevant for understanding stem cell maintenance in a range of species, from nematodes to humans.

developmental biology↗

bioRxiv2021

AN INTEGRATIVE BIOLOGY APPROACH TO QUANTIFY THE BIODISTRIBUTION OF AZIDOHOMOALANINE IN VIVO

Identification and quantitation of newly synthesized proteins (NSPs) are critical to understanding protein dynamics in development and disease. Probing the nascent proteome can be achieved using non-canonical amino acids (ncAAs) to selectively label the NSPs utilizing endogenous translation machinery, which can then be quantitated with mass spectrometry. Since its conception, ncAA labeling has been applied to study many in vitro systems and more recently the in vivo proteomes of complex organisms such as rodents. In vivo labeling is typically achieved by introducing ncAAs into diet, which requires extended labeling times. We have previously demonstrated that labeling the murine proteome is feasible via injection of azidohomoalanine (Aha), a ncAA and methionine (Met) analog, without the need for Met depletion. With the ability to isolate NSPs without applying stress from dietary changes, Aha labeling can address biological questions wherein temporal protein dynamics are significant. However, accessing this temporal resolution requires a more complete understanding of Aha distribution kinetics in tissues. Furthermore, studies of physiological effects of ncAA administration have been limited to gross observation of animal appearance. To address these gaps, we created a deterministic, compartmental model of the biokinetic transport and incorporation of Aha in mice. Parameters were informed from literature and experimentally. Model results demonstrate the ability to predict Aha distribution and labeling under a variety of dosing paradigms and confirms the use of the model as a tool for design of future studies. To establish the suitability of the method for in vivo studies, we investigated the impact of Aha administration on normal physiology by analyzing the plasma metabolome following Aha injection. We show that Aha administration does not significantly perturb cellular functions as reflected by an unchanged plasma metabolome compared to non-injected controls. Author SummaryAs the machinery of life, proteins play a key role in dynamic processes within an organism. As such, the response of the proteome to perturbation is increasingly becoming a critical component of biological and medical studies. Dysregulation of protein mechanisms following exposure to experimental treatment conditions can implicate physiological mechanisms of health and disease, elucidate toxin/drug response, and highlight potential targets for novel therapies. Traditionally, these questions have been probed by studying perturbations in total proteins following an experimental treatment. However, the proteome is expansive and noisy, often an early response can be indiscernible against the background of unperturbed proteins. Here, we apply a technique to selectively label newly synthesized proteins, which enables capturing early changes in protein behavior. We utilize an amino acid analog that naturally incorporates into proteins, and investigate the tissue distribution, protein labeling efficiency, and potential physiological impact of this analog in mice. Our results demonstrate that we can reproducibly predict protein labeling and that the administration of this analog does not significantly alter in vivo physiology over the course of our experimental study. We further present a computational model that can be used to guide future experiments utilizing this technique to study proteomic responses to stimuli.

bioengineering↗

bioRxiv2021

AN INULIN-TYPE FRUCTAN ENRICHED EXCLUSIVE ENTERAL NUTRITION FORMULA SUPPRESSES COLITIS THROUGH GUT MICROBIOME MODULATION AND PROMOTING EXPANSION OF ANTI-INFLAMMATORY T CELL SUBSETS

Background & AimsExclusive enteral nutrition (EEN) is used to treat pediatric Crohns disease (CD), but therapeutic benefits are not long lasting. Due to reported lower efficacy EEN is not routinely used to treat pediatric ulcerative colitis (UC). Inulin-type fructans (IN) beneficially modulate the gut microbiome and promote expansion of anti-inflammatory immune cells. We hypothesized that enriching EEN with IN (EENIN) would enhance treatment efficacy. To test this, we examined the effects of EEN-IN on colitis development, the gut microbiome and CD4+ T cells using an adoptive T cell transfer model of colitis. MethodsTCR-{beta} deficient mice were randomized to one of four groups: 1) Control, 2) Chow, 3) EEN and 4) EEN-IN, and naive CD4+ T cells were adoptively transferred into groups 2-4, after which mice were monitored for 5-weeks prior to experimental endpoint. ResultsMice fed EEN-IN showed greater colitis protection, with colonic shortening, goblet cell and crypt density loss reduced over that of EEN fed mice and reduced disease activity and immune cell infiltration compared to chow fed mice, and less crypt hyperplasia and higher survival compared to both groups. EENIN mice maintained colonic mucus layer thickness and had increased levels of Foxp3+IL-10+ and Ror{gamma}t+IL- 22+ and reduced levels of Tbet+IFN{gamma}+ and Tbet+TNF+ CD4+ T cells. EEN-IN also lead to higher butyrate, Bifidobacterium spp. and Bacteroides spp. concentrations. ConclusionThe EEN-IN group showed reduced colitis development as compared to the chow and EEN groups. This highlights the potential benefits of EEN-IN as a novel induction therapy for pediatric CD and UC patients. SynopsisWe demonstrated that inulin-type fructan enriched exclusive enteral nutrition formula reduced colitis development likely due to butyrate-dependent pathways that helped preserve the mucus layer and promote an anti-inflammatory intestinal environment via expansion of regulatory T cells.

microbiology↗

bioRxiv2020

Flexible RSV prefusogenic fusion glycoprotein exposes multiple neutralizing epitopes that may collectively contribute to protective immunity

Human respiratory syncytial virus (RSV) is a significant cause of lower respiratory tract infection in infants, young children, and older adults. There is no licensed vaccine and prophylactic treatment options are limited and not widely available in developing countries with the greatest disease burden. The RSV fusion (F) glycoprotein is a primary target of host immunity and thus a major focus for vaccine development. The native F glycoprotein structure is flexible and undergoes significant rearrangements from the metastable prefusion to a stable postfusion structure with neutralizing epitopes on intermediate structures. We hypothesize trimeric vaccine strategies that recapitulate the breathable F quaternary structure, and provide accessibility of B-cells to epitopes on intermediate conformations, may collectively contribute to protective immunity, while ridge prefusion F structures restrict access to key protective epitopes. To test this hypothesis, we used the near full-length native prefusogenic F as a backbone to construct three prefusion F variants with substitutions in the hydrophobic head cavity: 1) disulfide bond double mutant (DS), 2) space filling hydrophobic amino acid substitutions (Cav1), and 3) DS plus Cav1 substitutions (DS-Cav1). In this study, we compared the immunogenicity of prefusogenic F to the immunogenicity of the prefusion F variants in two animal models. Native prefusogenic F was significantly more immunogenic producing high titer antibodies to prefusogenic, prefusion, and postfusion F structures compared to animals immunized with prefusion F DS or DS-Cav1. Importantly, native prefusogenic F elicited antibodies that targeted neutralizing epitopes including prefusion-specific site zero (O) and V as well as conformation-independent neutralizing sites II and IV. Immunization with prefusion F DS or DS-Cav1 elicited antibodies primarily targeting antigenic sites O and V with little or no detectable antibodies to other key neutralizing sites. Animals immunized with native prefusogenic F also had significantly higher neutralizing antibodies that cross-neutralized RSV A and B subtypes while immunization with DS or DS-Cav1 elicited neutralizing antibodies primarily to the A subtype. We conclude that breathable trimeric vaccines that closely mimic the native F-structure, and incorporate strategies for B-cell accessibility to protective epitopes, are important considerations for vaccine design. F structures locked in a single conformation restrict B-cell access to neutralizing epitopes that may collectively contribute to destabilizing F-trimers important for broad protection. These results also have implications for vaccine strategies targeting other type 1 integral membrane proteins.

immunology↗

bioRxiv2020

SARS-CoV-2 spike glycoprotein vaccine candidate NVX-CoV2373 elicits immunogenicity in baboons and protection in mice

The COVID-19 pandemic continues to spread throughout the world with an urgent need for a safe and protective vaccine to effectuate herd immunity to control the spread of SARS-CoV-2. Here, we report the development of a SARS-CoV-2 subunit vaccine (NVX-CoV2373) produced from the full-length spike (S) protein, stabilized in the prefusion conformation. Purified NVX-CoV2373 S form 27.2nm nanoparticles that are thermostable and bind with high affinity to the human angiotensin-converting enzyme 2 (hACE2) receptor. In mice and baboons, low-dose NVX-CoV2373 with saponin-based Matrix-M adjuvant elicits high titer anti-S IgG that is associated with blockade of hACE2 receptor binding, virus neutralization, and protection against SARS-CoV-2 challenge in mice with no evidence of vaccine-associated enhanced respiratory disease (VAERD). NVX-CoV2373 vaccine also elicits multifunctional CD4+ and CD8+ T cells, CD4+ T follicular helper T cells (Tfh), and the generation of antigen-specific germinal center (GC) B cells in the spleen. These results support the ongoing phase 1/2 clinical evaluation of the safety and immunogenicity of NVX-CoV2327 with Matrix-M (NCT04368988).

microbiology↗

bioRxiv2019

Non-rainfall moisture: a key driver of carbon flux from standing litter in arid, semiarid, and mesic grasslands

Models assume that rainfall is the major source of moisture driving decomposition. Non-rainfall moisture (NRM: high humidity, dew, and fog) can also induce standing litter decomposition, but there have been few standard measurements of NRM-mediated decompositions across sites, and no efforts to extrapolate the contribution of NRM to larger scales to assess whether this mechanism can improve model predictions. Here we show that NRM is an important, year-round source of moisture in grassland sites with contrasting moisture regimes using field measurements and modeling. We first characterized NRM frequency and measured NRM-mediated decomposition in sites on the extreme dry and wet end of grassland systems: at two sites in the Namib Desert, Namibia (hyperarid desert) and at one site in Iowa, USA (tallgrass prairie). NRM was frequent at all sites (85-99% of hours that litter was likely to be wet were attributed to NRM) and tended to occur in cool, high-humidity periods for several hours or more at a time. NRM also caused respiration of standing litter at all sites when litter became sufficiently wet (>5% for fine litter and >13% for coarse), and contributed to mass loss, even in the Namib West site that had almost no rain. When we modeled annual mass loss induced by NRM and rain, and extrapolated our characterization of NRM decomposition to a final site with intermediate rainfall (Sevilleta, New Mexico, semiarid grassland), we found that models driven by rainfall alone underestimated mass loss, while including NRM produced estimates within the range of observed mass loss. Together these findings suggest that NRM is an important missing component in quantitative and conceptual models of litter decomposition, but there is nuance involved in modeling NRM at larger scales. Specifically, temperature and physical features of the substrate emerge as factors that affect the common microbial response to litter wetting under NRM across grasslands sites, and require further study. Hourly humidity can provide an adequate proxy of NRM frequency, but site-specific calibration with litter wetness is needed to accurately attribute decomposition to periods when NRM wets litter. Greater recognition of NRM-driven decomposition and its interaction with other processes (e.g. photodegradation) is needed, especially since fog, dew, and humidity are likely to shift under future climates.\n\nManuscript highlightsO_LINon-rainfall moisture (NRM; humidity, fog, dew) induces decomposition in grasslands\nC_LIO_LINRM decomposition depends on substrate type, and occurs at colder times than rain\nC_LIO_LIIncluding NRM (instead of rain alone) improved predictions of litter decomposition\nC_LI

ecology↗