Biology subjects

Nelson, D. C.

Publications and source records attributed to Nelson, D. C..

16 recordsLinked to original sources

bioRxiv2026

A synthetic phageome reveals widespread and diverse human antibody responses to bacteriophages

Bacteriophages colonize humans, yet their recognition by humoral immunity remains unclear. We developed PhageScan, a PhIP- Seq platform comprising 277,051 phage and 49,630 human viral oligopeptides. Profiling sera from 192 healthy donors without prior phage therapy characterized anti-phage response specificity and revealed widespread, strong IgG reactivity to phages, comparable in prevalence and magnitude to that against human viruses. Recognition patterns were however highly individualized. Among phage sequences, 12,576 peptides were significantly enriched in at least two individuals. Strongly immunogenic epitopes preferentially originated from phages infecting pathogenic bacteria, whereas widely shared ''public'' epitopes were enriched among phages of environmental and food-associated bacteria. ImmE, a lactococcal phage endolysin recognized in 59.9% of individuals, showed reduced antibacterial activity in the presence of sera identified as ImmE-reactive by PhageScan, demonstrating functional relevance. These findings establish phage-directed antibodies as a widespread component of human immunity and PhageScan as a scalable platform for defining phage--antibody interactions.

immunology↗

bioRxiv2026

Viral Burden: New insights into Estimating IgG Recognition Across Human Populations

Assessment of viral burden in populations is essential for understanding virus epidemiology and herd immunity potential. While serological profiling is straightforward at the individual level, it remains challenging at scale. We propose analysis advancing in serological technologies for broader population-level analysis and comparison. We used an epitope library in Phage Display ImmunoPrecipitation (PhIP) technology (VirScan type library) to assess IgG recognition of 49,630 representative viral oligopeptides in 134 serum samples from two populations (Poland and the US). Only 5.9% of oligopeptides were immunogenic, yet IgG recognition of viruses was consistent across populations - over 90% of virus species, 99% of genera, and 97% of families were detected. Shannon Diversity Index analysis supported these findings. Among immunogenic peptides, 9.1% were significantly more frequently recognized, though not correlated with recognition strength. We further proposed a normalization method to account for differences in viral proteome representation when assessing immune burden: the burden score. Finally, we demonstrated how this approach facilitates serological comparisons between populations. These observations show that while people are exposed to similar viruses, they produce antibodies against different viral epitopes. This individual variability, combined with broad virus recognition, likely strengthens population-level protection. Epitope recognition frequency seems to be shaped more by population exposure than by magnitude of response that an epitope can induce. Accurately measuring viral burden can inform healthcare planning, and antiviral technologies development.

immunology↗

bioRxiv2026

HDAC Inhibition Sensitizes Pancreatic Tumors to DNA Damage by Global Redistribution of the Transcriptional Machinery

The DNA damage response (DDR) is critical for pancreatic ductal adenocarcinoma (PDAC) development and therapeutic responses, including to genotoxic agents. While epigenetic modulators have been shown to contribute to the DDR, how chromatin regulation dictates responses to DNA damage in PDAC remains incompletely understood. Here, we identify Class I histone deacetylases (HDACs) as critical regulators of the DDR. HDAC1/2 directs the genomic distribution of H3K27ac, ensuring sufficient BRD4 and RNA polymerase II (Pol II) occupancy at DDR gene promoters. HDAC inhibition by entinostat shifts the balance of H3K27 acetylation preferentially towards intergenic regions, diverting BRD4 and Pol II from promoters, thereby suppressing DDR gene expression. In line with this, HDAC inhibition heightens DNA damage and sensitizes PDAC to diverse DNA-damaging and DDR-targeting agents. Since the clinical development of HDAC inhibitors has been limited by systemic toxicity, we developed bottlebrush prodrug (BPD) nanoparticles for tumor-selective entinostat delivery. Entinostat-BPD achieved tumor-specific HDAC inhibition while displaying potent efficacy and reduced systemic toxicity. These findings reveal an HDAC-dependent DDR vulnerability and offer combinational and precision targeting strategies to facilitate clinical translation and improve PDAC patient outcomes. SIGNIFICANCE STATEMENTThe ability of tumor cells to tolerate DNA damage limits the efficacy of many anticancer therapies. Our study reveals that pancreatic cancer cells enforce this resistance by sustaining expression of DNA damage response (DDR) genes through Class I histone deacetylases (HDACs). HDACs maintain genome-wide acetylation patterns required for efficient recruitment of the transcriptional machinery to DDR genes. Pharmacological HDAC inhibition disrupts this process and sensitizes pancreatic cancer cells to diverse DNA-damaging agents. To overcome systemic toxicity that limits translational potential, we further establish a bottlebrush prodrug nanoparticle platform that enables tumor-selective HDAC inhibition. Given the central role of the DDR in cancer, targeting HDAC-mediated DDR regulation through drug combinations and precision delivery may have broad therapeutic relevance across cancer types.

cancer biology↗

bioRxiv2024

An N-terminal domain specifies developmental control by the SMAX1-LIKE family of transcriptional co-repressors in Arabidopsis thaliana

SMAX1-LIKE (SMXL) proteins are transcriptional co-repressors that regulate many aspects of plant growth and development. Proteins from the SMAX1- and SMXL78-clades of this family are targeted for degradation after karrikin or strigolactone perception, triggering downstream responses. We investigated how SMXL proteins control development. SMXL7 can partially replicate SMAX1 function in seeds and seedlings, but SMAX1 cannot replace SMXL7 in shoot branching control. Therefore, the distinct roles of these genes arise from differences in protein activity more than expression. Analysis of chimeras and domain deletions of SMAX1 and SMXL7 proteins revealed that an N-terminal domain is necessary and sufficient to specify developmental functions. We screened 158 transcription factors for interactions with SMAX1. The N-terminal domain is necessary and/or sufficient for the majority of candidate interactions. These discoveries enable cross-wiring of karrikin and strigolactone control of plant development and lay a foundation for understanding how SMXL proteins evolved functional differences.

plant biology↗

bioRxiv2023

Inhibiting Stromal Class I HDACs Curbs Pancreatic Cancer Progression

Oncogenic lesions in pancreatic ductal adenocarcinoma (PDAC) hijack the epigenetic machinery in stromal components to establish a desmoplastic and therapeutic resistant tumor microenvironment (TME). Here we identify Class I histone deacetylases (HDACs) as key epigenetic factors facilitating the induction of pro-desmoplastic and pro-tumorigenic transcriptional programs in pancreatic stromal fibroblasts. Mechanistically, HDAC-mediated changes in chromatin architecture enable the activation of pro-desmoplastic programs directed by serum response factor (SRF) and forkhead box M1 (FOXM1). HDACs also coordinate fibroblast pro-inflammatory programs inducing leukemia inhibitory factor (LIF) expression, supporting paracrine pro-tumorigenic crosstalk. HDAC depletion in cancer-associated fibroblasts (CAFs) and treatment with the HDAC inhibitor entinostat (Ent) in PDAC mouse models reduce stromal activation and curb tumor progression. Notably, HDAC inhibition (HDACi) enriches a lipogenic fibroblast subpopulation, a potential precursor for myofibroblasts in the PDAC stroma. Overall, our study reveals the stromal targeting potential of HDACi, highlighting the utility of this epigenetic modulating approach in PDAC therapeutics.

cancer biology↗

bioRxiv2022

Escaping antibody responses to bacteriolytic enzymes Pal and Cpl-1 by epitope scanning and engineering

Bacteriolytic enzymes are promising antibacterial agents, but they can cause a typical immune response in vivo. In this study, we used a targeted modification of two antibacterial endolysins, Pal and Cpl-1. We identified the key immunogenic amino-acids, and designed and tested new, bacteriolytic variants with altered immunogenicity. One new variant of Pal (257-259 MKS [->] TFG) demonstrated decreased immunogenicity while a similar mutant (257-259 MKS [->] TFK) demonstrated increased immunogenicity. A third variant (280-282 DKP [->] GGA) demonstrated significantly increased antibacterial activity and it was not cross-neutralized by antibodies induced by the wild-type enzyme. We propose this variant as a new engineered endolysin with increased antibacterial activity that is capable of escaping cross-neutralization by antibodies induced by wild-type Pal. We show that efficient antibacterial enzymes that avoid cross-neutralization by IgG can be developed by epitope scanning, in silico modelling, and substitutions of identified key amino acids with a high rate of success. Importantly, this universal approach can be applied to many proteins beyond endolysins and has the potential for design of numerous biological drugs.

bioengineering↗

bioRxiv2022

An Orobanche cernua x Orobanche cumana segregating population provides insight into the regulation of germination specificity in a parasitic plant

Germination of seeds of Orobanche species requires specific chemicals exuded by host roots. A family of "divergent" KARRIKIN INSENSITIVE2 (KAI2d) genes encode proteins that recognize strigolactone (SL) class germination simulants. We explored specificity of germination stimulant detection by analyzing interspecific segregants of a cross between Orobanche cernua and O. cumana, closely related species that differ in stimulant response. O. cernua parasitizes tomato and germinates in response to the SL orobanchol, while O. cumana parasitizes sunflower and responds to dehydrocostus lactone (DCL). KAI2d genes were catalogued in parents and in segregants that showed stimulant specificity. KAI2d genes were also functionally assayed in the Arabidopsis kai2 mutant background. We identified five full-length KAI2d genes in O. cernua and eight in O. cumana. The O. cernua KAI2d2, as well as its ortholog in O. cumana, are associated with SL perception. A cluster of O. cumana KAI2d genes was genetically linked to DCL perception, although no specific receptor gene was identified by heterologous complementation. These findings support the KAI2d-mediated perception of SLs, but fall short of explaining how O. cumana perceives DCL. The ability of some O. cumana KAI2d genes to detect SLs points to the involvement of additional factors in regulating stimulant specificity.

plant biology↗

bioRxiv2022

Strigolactones are chemoattractants for host tropism in Orobanchaceae parasitic plants

Parasitic plants are worldwide threats that damage major agricultural crops. To initiate infection, parasitic plants have developed the ability to locate hosts and grow towards them. This ability, called host tropism, is critical for parasite survival, but its underlying mechanism remains mostly unresolved. To characterize host tropism, we used the model facultative root parasite Phtheirospermum japonicum, a member of the Orobanchaceae. Here, we show that strigolactones (SLs) function as host-derived chemoattractants. Chemotropism to SLs is also found in Striga hermonthica, a parasitic member of the Orobanchaceae, but not in non-parasites. Intriguingly, chemotropism to SLs in P. japonicum is attenuated in ammonium ion-rich conditions, where SLs are perceived, but the resulting asymmetrical accumulation of the auxin transporter PIN2 is diminished. P. japonicum encodes putative receptors that sense exogenous SLs, whereas expression of a dominant-negative form reduces its chemotropic ability. We propose a new function for SLs as navigators for parasite roots.

plant biology↗

bioRxiv2022

Thermal characterization of the multimeric bacteriophage endolysin PlyC

Bacteriophage endolysins degrade the bacterial peptidoglycan and are considered enzymatic alternatives to small molecule antibiotics. In particular, the multimeric streptococcal endolysin PlyC has appealing antibacterial properties. However, a comprehensive thermal analysis of PlyC is lacking, which is necessary for evaluating long-term stability and downstream therapeutic potential. Biochemical and kinetic-based methods were used in combination with differential scanning calorimetry to investigate the structural, kinetic and thermodynamic stability of PlyC and its various subunits and domains. The PlyC holoenzyme structure is irreversibly compromised due to partial unfolding and aggregation at 46{degrees}C. Unfolding of the catalytic subunit, PlyCA, instigates this event, resulting in the kinetic inactivation of the endolysin. In contrast to PlyCA, the PlyCB octamer (the cell wall binding domain) is thermostable, denaturing at [~]75{degrees}C. Isolation of PlyCA or PlyCB alone altered their thermal properties. Contrary to the holoenzyme, PlyCA alone unfolds uncooperatively and is thermodynamically destabilized whereas the PlyCB octamer reversibly dissociates into monomers and forms an intermediate state at 74{degrees}C in phosphate buffered saline, with each subunit subsequently denaturing at 92{degrees}C. Adding folded PlyCA to an intermediate state PlyCB, followed by cooling, allowed for in vitro reconstitution of the active holoenzyme.

biophysics↗

bioRxiv2021

A KARRIKIN INSENSITIVE2 paralog in lettuce mediates highly sensitive germination responses to karrikinolide

Karrikins (KARs) are chemicals in smoke that can enhance germination of many plants. Lactuca sativa cv. Grand Rapids (lettuce), germinates in the presence of nanomolar karrikinolide (KAR1). We found that lettuce is much less responsive to KAR2 or a mixture of synthetic strigolactone analogs, rac-GR24. We investigated the molecular basis of selective and sensitive KAR1 perception in lettuce. The lettuce genome contains two copies of KARRIKIN INSENSITIVE2 (KAI2), a receptor that is required for KAR responses in Arabidopsis thaliana. LsKAI2b is more highly expressed than LsKAI2a in dry achenes and during early stages of seed imbibition. Through cross-species complementation assays in Arabidopsis we found that LsKAI2b confers robust responses to KAR1, but LsKAI2a does not. Therefore, LsKAI2b likely mediates KAR1 responses in lettuce. We compared homology models of the ligand-binding pockets of KAI2 proteins from lettuce and a fire follower, Emmenanthe penduliflora. This identified pocket residues 96, 124, 139, and 161 as candidates that influence the ligand-specificity of KAI2. Further support for the significance of these residues was found through a broader comparison of pocket residue conservation among 324 asterid KAI2 proteins. We tested the effects of substitutions at these four positions in Arabidopsis thaliana KAI2 and found that a broad array of responses to KAR1, KAR2, and rac-GR24 could be achieved.

plant biology↗

bioRxiv2021

Activation Mechanism of Strigolactone Receptors And Its Impact On Ligand Selectivity Between Host And Parasitic Plants

Parastic weeds such as Striga have led to significant losses in agricultural productivity worldwide. These weeds use the plant hormone strigolactone as a germination stimulant. Strigolactone signaling involves substrate binding and hydrolysis followed by a large conformational change of the receptor to a "closed" or "active" state that is able to associate with a downstream signaling partner MAX2/D3. The crystal structure of the active and inactive AtD14 receptor have helped in elucidating the structural changes involved in activation. However, the mechanism by which the receptor activates remains unknown. The ligand dependence of AtD14 activation has been disputed by mutagenesis studies showing that enzymatically inactive receptors are able to form a complex with MAX2 proteins. Furthermore, activation differences between strigolactone receptor in Striga, ShHTL7 and textitAtD14 could contribute to the high sensitivity to strigolactones exhibited by parasitic plants. Using molecular dynamics simulations, we demonstrate that both AtD14 and ShHTL7 could adopt an active conformation in absence of ligand. However, the ShHTL7 receptor exhibits higher population in the inactive apo state as compared to the AtD14 receptor. We demonstrate that this difference in inactive state population is caused by sequence differences between their D-loops and its interactions with the catalytic histidine that prevents full binding pocket closure in ShHTL7. These results indicate that hydrolysis of a strigolactone ligand would enhance the active state population by destabilizing the inactive state in ShHTL7 as compared to AtD14. We also show that the mechanism of activation is more concerted in AtD14 than in ShHTL7 and that the main barrier to activation in ShHTL7 is closing of the binding pocket.

plant biology↗

bioRxiv2021

Rapid analysis of strigolactone receptor activity in a Nicotiana benthamiana dwarf14 mutant

DWARF14 (D14) is an [a]/{beta}-hydrolase and receptor for the plant hormone strigolactone (SL) in angiosperms. Upon SL perception, D14 works with MORE AXILLARY GROWTH2 (MAX2) to trigger polyubiquitination and degradation of DWARF53(D53)-type proteins in the SUPPRESSOR OF MAX2 1-LIKE (SMXL) family. We used CRISPR-Cas9 to generate knockout alleles of the two homoeologous D14 genes in the Nicotiana benthamiana genome. The Nbd14a,b double mutant had several phenotypes that are consistent with the loss of SL perception in other plants, including increased axillary bud outgrowth, reduced height, shortened petioles, and smaller leaves. A ratiometric fluorescent reporter system was used to monitor degradation of SMXL7 from Arabidopsis thaliana (AtSMXL7) after transient expression in N. benthamiana and treatment with the strigolactone analog GR24. AtSMXL7 was degraded after treatment with GR245DS, which has the stereochemical configuration of SLs, as well as its enantiomer GR24ent-5DS . In Nbd14a,b leaves, AtSMXL7 abundance was unaffected by GR24. Transient coexpression of AtD14 with the AtSMXL7 reporter in Nbd14a,b restored the degradation response to GR24, but required an active catalytic triad. With this platform, we evaluated the ability of several AtD14 mutants that had not been characterized in plants to target AtSMXL7 for degradation.

plant biology↗

bioRxiv2021

Molecular basis for recognition of the Group A Carbohydrate backbone by the PlyC streptococcal bacteriophage endolysin

Endolysins are peptidoglycan (PG) hydrolases that function as part of the bacteriophage (phage) lytic system to release progeny phage at the end of a replication cycle. Notably, endolysins alone can produce lysis without phage infection, which offers an attractive alternative to traditional antibiotics. Endolysins from phage that infect Gram-positive bacterial hosts contain at least one enzymatically active domain (EAD) responsible for hydrolysis of PG bonds and a cell wall binding domain (CBD) that binds a cell wall epitope, such as a surface carbohydrate, providing some degree of specificity for the endolysin. Whilst the EADs typically cluster into conserved mechanistic classes with well-defined active sites, relatively little is known about the nature of the CBDs and only a few binding epitopes for CBDs have been elucidated. The major cell wall components of many streptococci are the polysaccharides that contain the polyrhamnose (pRha) backbone modified with species-specific and serotype-specific glycosyl side chains. In this report, using molecular genetics, microscopy, flow cytometry and lytic activity assays, we demonstrate the interaction of PlyCB, the CBD subunit of the streptococcal PlyC endolysin, with the pRha backbone of the cell wall polysaccharides, Group A Carbohydrate (GAC) and serotype c-specific carbohydrate (SCC) expressed by the Group A Streptococcus and Streptococcus mutans, respectively.

biochemistry↗

bioRxiv2020

Role of substrate recognition in modulating strigolactone receptor selectivity in witchweed.

Witchweed, or Striga hermonthica, is a parasitic weed that destroys billions of dollars worth of crops globally every year. Its germination is stimulated by strigolactones exuded by its host plants. Despite high sequence, structure, and ligand binding site conservation across different plant species, one strigolactone receptor in witchweed (Sh HTL7) uniquely exhibits a picomolar EC50 for downstream signaling. Previous biochemical and structural analyses have hypothesized that this unique ligand sensitivity can be attributed to a large binding pocket volume in Sh HTL7 resulting in enhanced ability to bind substrates. Additional structural details of the substrate binding process can help explain its role in modulating the ligand selectivity. Using long-timescale molecular dynamics simulations, we demonstrate that mutations at the entrance of the binding pocket facilitate a more direct ligand binding pathway to Sh HTL7, whereas hydrophobicity at the binding pocket entrance results in a stable "anchored" state. We also demonstrate that several residues on the D-loop of At D14 stabilize catalytically inactive conformations. Finally, we show that strigolactone selectivity is not modulated by binding pocket volume. Our results indicate that while ligand binding is not the sole modulator of strigolactone receptor selectivity, it is a significant contributing factor. These results can be used to inform the design of selective antagonists for strigolactone receptors in witchweed.

plant biology↗

bioRxiv2020

A series of dual-reporter vectors for ratiometric analysis of protein abundance in plants

Ratiometric reporter systems enable comparisons of the abundance of a protein of interest, or "target," relative to a reference protein. Both proteins are encoded on a single transcript but are separated during translation. This arrangement bypasses the potential for discordant expression that can arise when the target and reference proteins are encoded by separate genes. We generated a set of 18 Gateway-compatible vectors termed pRATIO that combine a variety of promoters, fluorescent and bioluminescent reporters, and 2A "self-cleaving" peptides. These constructs are easily modified to produce additional combinations or introduce new reporter proteins. We found that mScarlet-I provides the best signal-to-noise ratio among several fluorescent reporter proteins during transient expression experiments in Nicotiana benthamiana. Firefly and Gaussia luciferase also produce high signal-to-noise in N. benthamiana. As proof of concept, we used this system to investigate whether degradation of the receptor KAI2 after karrikin treatment is influenced by its subcellular localization. KAI2 is normally found in the cytoplasm and the nucleus of plant cells. In N. benthamiana, karrikin-induced degradation of KAI2 was only observed when it was retained in the nucleus. These vectors are tools to easily monitor in vivo the abundance of a protein that is transiently expressed in plants, and will be particularly useful for investigating protein turnover in response to different stimuli.

plant biology↗

bioRxiv2017

The complex origins of strigolactone signalling in land plants

Strigolactones (SLs) are a class of plant hormones that control many aspects of plant growth. The SL signalling mechanism is homologous to that of karrikins (KARs), smoke-derived compounds that stimulate seed germination. In angiosperms, the SL receptor is an /{beta} hydrolase known as DWARF14 (D14); its close homologue, KARRIKIN INSENSITIVE2 (KAI2), functions as a KAR receptor, and likely recognizes an uncharacterized, endogenous signal. Previous phylogenetic analyses have suggested that the KAI2 lineage is ancestral in land plants, and that canonical D14-type SL receptors only arose in seed plants; this is paradoxical, however, as non-vascular plants synthesize and respond to SLs. Here, we have used a combination of phylogenetic and structural approaches to re-assess the evolution of the D14/KAI2 family in land plants. We analyzed 339 members of the D14/KAI2 family from land plants and charophyte algae. Our phylogenetic analyses show that the divergence between the eu-KAI2 lineage and the DDK (D14/DLK2/KAI2) lineage that includes D14 occurred very early in land plant evolution. We identify characteristic structural features of D14 and KAI2 proteins, and use homology modelling to show that the earliest members of the DDK lineage structurally resemble KAI2, and not D14 proteins. Furthermore, we show that probable SL receptors in non-seed plants do not have D14-like structure. Our results suggest that SL perception has relatively relaxed structural requirements, and that the evolution from KAI2-like to D14-like protein structure in the DDK lineage may have been driven by interactions with protein partners, rather than being required for SL perception itself.

plant biology↗