bluuebunny/crossref_metadata_2025_split huggingface dataset:
A strain of
Streptococcus mitis
inhibits biofilm formation of caries pathogens via abundant hydrogen peroxide production
Authors: Williams Isabella , Tuckerman Jacob S. , Peters Daniel I. , Bangs Madisen , Williams Emily , Shin Iris J. , Kaspar Justin R. 
Published: February 2025

ABSTRACT

Commensal oral streptococci that colonize supragingival biofilms deploy mechanisms to combat competitors within their niche. Here, we determined that
Streptococcus mitis
more effectively inhibited biofilm formation of
Streptococcus mutans
compared to other oral streptococci. This phenotype was common among all isolates of
S. mutans
, but was specific to a single strain of
S. mitis
, ATCC 49456. We documented ATCC 49456 to accumulate four to five times more hydrogen peroxide (H
2
O
2
) than other
Streptococcus
species tested, and 5–18 times more than other
S. mitis
strains assayed.
S. mutans
biofilm formation inhibition was dependent on cell contact/proximity and reduced when grown in media containing catalase or with a
S. mitis
mutant of pyruvate oxidase (
spxB; pox
), confirming that SpxB-dependent H
2
O
2
production was a major antagonistic factor. Addition of
S. mitis
within hours after
S. mutans
inoculation was effective at reducing biofilm biomass, but not for 24 h pre-formed biofilms in an SpxB-dependent manner. Transcriptome analysis revealed responses for both
S. mitis
and
S. mutans
, with several
S. mutans
differentially expressed genes following a gene expression pattern we have previously described, while others being unique to the interaction with
S. mitis
. Finally, we show that
S. mitis
also affected coculture biofilm formation of several other commensal streptococci as well as cariogenic
Streptococcus sobrinus
. Our study shows that strains with abundant H
2
O
2
production are effective at inhibiting initial growth of caries pathogens like
S. mutans
, but are less effective at disrupting pre-formed biofilms and have the potential to influence the stability of other oral commensal strains.

IMPORTANCE

Antagonistic properties displayed by oral bacteria have been sought as therapeutic approaches against dental caries pathogens like
Streptococcus mutans
. An emergent theme has been the ability of select strains that produce high amounts of hydrogen peroxide to effectively inhibit the growth of
S. mutans
within
in vitro
and
in vivo
models. Our study builds on these previous findings by determining that
Streptococcus mitis
ATCC 49456 is a high hydrogen peroxide producer, compared to other
Streptococcus
species as well as additional strains of
S. mitis
. In addition to
S. mutans
, we show that ATCC 49456 also affects biofilm formation of other oral streptococci, a non-desirable trait that should be weighed heavily for strains under consideration as probiotics. Further phenotypic characterization of strains like
S. mitis
ATCC 49456 in mixed-species settings will allow us to hone in on qualities that are optimal for probiotic strains that are intended to prevent the emergence of odontopathogens.
Single-cell delineation of strain-specific HIV-1 Vif activities using dual reporter sensor cells and live cell imaging
Authors: Guerrero Jorge F. , Zimdars Laraine L. , Bruce James W. , Becker Jordan T. , Evans Edward L. , Torabi Soroosh , Striker Rob , Berry Scott M. , Sherer Nathan M. 
Published: February 2025

ABSTRACT

Human immunodeficiency virus type 1 (HIV-1) genome diversification is a key determinant of viral evolution and the pathogenesis of HIV/AIDS. Antiretroviral therapy is non-curative, and in the context of monitoring the latent reservoir, precision tools are needed to detect and enumerate HIV-1 genomes as well as to assess their heterogeneity, replication potential, and predict responses to therapy. Current sequencing-based methodologies are often unable to confirm intact genomes and most cell-based reporters provide limited information pertaining to viral fitness. In this study, we describe dual reporter sensor cells (DRSCs), an imaging-based reporter system designed to detect HIV-1 infection and measure several independent attributes of the virus in a single-cell high-content assay. We show that the DRSC assay can be used to measure infection, viral gene activation kinetics, and quantify viral circumvention of host antiviral responses. Using the DRSCs, we confirmed markedly different functional heterogeneity for
vif
alleles derived from diverse HIV-1 strains and subtypes affecting both rates of APOBEC3G degradation and the cell cycle. Furthermore, the assay allowed for the delineation of virus co-receptor preference (X4- vs R5-tropism) and visualization of virion assembly. Overall, our study illustrates proof-of-principle for a multivariate imaging-based cell-based system capable of detecting HIV-1 and studying viral genetic variability with greater data richness relative to prior available modalities.

IMPORTANCE

Human immunodeficiency virus type 1 (HIV-1) is highly heterogeneous and constantly mutating. These changes drive immune evasion and can cause treatment efforts to fail. Here, we describe the “dual reporter sensor cell” (DRSC) assay; a novel imaging-based approach that allows for the detection of HIV-1 infection coupled with a multivariate definition of several independent phenotypic aspects of viral genome activity in a single integrated assay. We validate the DRSC system by studying lab-adapted and patient isolate-derived versions of the viral Vif accessory protein, confirming marked differences in the capacity of diverse
vif
alleles to mediate downregulation of antiviral APOBEC3G proteins and dysregulate the cell cycle.
Quorum sensing employs a dual regulatory mechanism to repress T3SS gene expression
Authors: Paul Payel , Podicheti Ram , Geyman Logan J. , Baker Elizabeth N. , Papenfort Kai , Rusch Douglas B. , van Kessel Julia C. 
Published: February 2025

ABSTRACT

The type III secretion system (T3SS) is a needle-like complex used by numerous bacterial pathogens in host infection to inject exotoxins into the host cell cytoplasm. The T3SS is a known virulence factor in the shrimp pathogen
Vibrio campbellii
. The ~40 genes comprising the
V. campbellii
T3SS are regulated by a network of transcription factors in response to changes in the cell’s environment: cell density (quorum sensing; QS), temperature, calcium, and host cell contact. Under positive environmental stimuli, the master T3SS transcription factor ExsA activates the expression of the four structural T3SS operons required for needle formation. Previous studies identified a key role of the master QS transcription factor LuxR: repression of
exsA
transcription via DNA binding at the
exsBA
promoter. Here, we uncovered a new regulatory role of LuxR: post-translational repression of ExsA activity via transcriptional repression of the gene encoding the anti-anti-activator ExsC. In
V. campbellii
, ExsC is a positive regulator of T3SS transcription; deletion of
exsC
decreases ExsA-dependent transcription activation of the T3SS structural promoters. Through genetic epistasis and
in vitro
biochemical assays, we show that LuxR directly binds the
exsC
promoter upstream of ExsA and represses transcription of
exsC
. Our findings collectively show that
V. campbellii
responds to high cell density signals to shut down ExsA-dependent expression of the T3SS via two mechanisms. We postulate that this dual regulatory mechanism by LuxR enables both the rapid inactivation of existing ExsA protein and blocks its further synthesis, leading to a rapid shutdown of T3SS activity at high cell density.

IMPORTANCE

Vibrio campbellii
utilizes the type III secretion system (T3SS) as a mechanism of pathogenesis, which is a highly studied “injectisome” complex that delivers exotoxins into host cells during infection. The T3SS pathogenicity island in
V. campbellii
comprises ~40 genes that are organized into four structural operons. In this study, we determined that quorum sensing—a method of bacterial communication—regulates T3SS genes both at the transcriptional and post-translational levels to shut down T3SS gene expression at high population densities.
Fission yeast metabolome dynamics during phosphate starvation and replenishment
Authors: Sanchez Ana M. , Kyaw Aye K. , Violante Sara Nunes , Garg Angad , Cross Justin R. , Shuman Stewart 
Published: February 2025

ABSTRACT

Inorganic phosphate is an essential nutrient acquired by cells from their environment and assimilated into myriad intracellular metabolites and macromolecules. Here, we characterize the metabolic responses of fission yeast to a 24 h interval of phosphate starvation, during which cells enter a state of G0 quiescence. Time-resolved profiling revealed that many key phosphometabolites were progressively depleted, including (i) NTPs, NDPs, and dNTPs; (ii) coenzyme A, NAD
+
, NADP
+
, NADH, and ADP-ribose; (iii) glycolysis pathway intermediates upstream of pyruvate; (iv) pentose phosphate pathway intermediates from 6-phosphogluconate to sedoheptulose-7-phosphate; (v) nucleotide sugars GDP-hexose, UDP-glucose/galactose, and UDP-GalNAc/GlcNAc; and (vi) phospholipid precursors glycerol-3-phosphate, CDP-choline, and glycerophosphocholine. By contrast, early Krebs cycle intermediates accumulated during phosphate starvation. Other metabolic changes included the following: (i) interdiction of
de novo
pyrimidine synthesis; (ii) depletion of
S
-adenosylmethionine and
S
-adenosylhomocysteine; (iii) transient accumulation of polyamine biosynthetic intermediates putrescine and 5-methylthioadenosine; (iv) accumulation of betaine (correlating with an increase in expression of
atd1
mRNA encoding aldehyde dehydrogenase); and (v) depletion of aminoadipate pathway intermediates 2-oxoadipate, 2-aminoadipate, and saccharopine. Replenishing phosphate after 24 h of starvation resulted in restoration of the pre-starvation metabolome (over 2 to 12 h) as cells exited quiescence and resumed growth.

IMPORTANCE

Fission yeast
Schizosaccharomyces pombe
is a valuable model system to study cellular phosphate homeostasis and the adaptive responses to chronic phosphate starvation. Previous analyses focused on changes in the fission yeast transcriptome and proteome during phosphate starvation-induced durable G0 quiescence. Here, we deployed metabolomics to survey the scope and temporal order of metabolite changes during 24 h of phosphate starvation and the kinetics of metabolic recovery after cells starved for 24 h are replenished with phosphate. These results contribute to a multi-omics understanding of how phosphate status impacts cell cycle, gene expression, metabolism, and chronological lifespan.
The HOPS and vCLAMP protein Vam6 connects polyphosphate with mitochondrial function and oxidative stress resistance in
Cryptococcus neoformans
Authors: Sánchez-León Eddy , Bhalla Kabir , Hu Guanggan , Lee Christopher W. J. , Lagace Melissa , Jung Won Hee , Kronstad James W. 
Published: February 2025

ABSTRACT

Cryptococcus neoformans
is considered one of the most dangerous fungal threats to human health, and the World Health Organization recently ranked it in the critical priority group for perceived public health importance. Proliferation of
C. neoformans
within mammalian hosts is supported by its ability to overcome nutritional limitations and endure stress conditions induced by the host immune response. Previously, we reported that the Vam6/Vps39/TRAP1-domain protein Vam6 was crucial for vacuolar morphology, iron acquisition, and virulence. However, the molecular mechanisms underlying the pleiotropic phenotypes resulting from loss of Vam6 remain poorly understood. In this study, we determined that Vam6 has roles in the HOPS complex for endomembrane trafficking to the vacuole and in the vCLAMP membrane contact site between the vacuole and mitochondria. Importantly, both of these roles regulate polyphosphate (polyP) metabolism, as demonstrated by a defect in trafficking of the VTC complex subunit Vtc2 for polyphosphate synthesis and by an influence on mitochondrial functions. In the latter case, Vam6 was required for polyP accumulation in response to electron transport chain inhibition and for overcoming oxidative stress. Overall, this work establishes connections between endomembrane trafficking, mitochondrial functions, and polyP homeostasis in
C. neoformans
.

IMPORTANCE

A detailed understanding of stress resistance by fungal pathogens of humans may provide new opportunities to improve antifungal therapy and combat life-threatening diseases. Here, we used a
vam6
deletion mutant to investigate the role of the homotypic fusion and vacuole protein sorting (HOPS) complex in mitochondrial functions and polyphosphate homeostasis in
Cryptococcus neoformans
, an important fungal pathogen of immunocompromised people including those suffering from HIV/AIDS. Specifically, we made use of mutants defective in late endocytic trafficking steps to establish connections to oxidative stress and membrane trafficking with mitochondria. In particular, we found that mutants lacking the Vam6 protein had altered mitochondrial function, and that the mutants were perturbed for additional mitochondria and vacuole-related phenotypes (e.g., membrane composition, polyphosphate accumulation, and drug sensitivity). Overall, our study establishes connections between endomembrane trafficking components, mitochondrial functions, and polyphosphate homeostasis in an important fungal pathogen of humans.
A proteomic and functional view of intrabacterial lipid inclusion biogenesis in mycobacteria
Authors: Dargham Tonia , Aguilera-Correa John Jairo , Avellan Romain , Mallick Ivy , Celik Léa , Santucci Pierre , Brasseur Gael , Poncin Isabelle , Point Vanessa , Audebert Stéphane , Camoin Luc , Daher Wassim , Cavalier Jean-François , Kremer Laurent , Canaan Stéphane 
Published: February 2025

ABSTRACT

During infection and granuloma formation, pathogenic mycobacteria store triacylglycerol as intrabacterial lipid inclusions (ILIs). This accumulation of nutrients provides a carbon source for bacterial persistence and slows down intracellular metabolism.
Mycobacterium abscessus
(
Mab
), a rapidly growing non-tuberculous actinobacterium, produces ILI throughout its infection cycle. Here,
Mab
was used as a model organism to identify proteins associated with ILI accumulation on a global scale. By using the APEX2 proximity labeling method in an
in vitro
model for ILI accumulation, we identified 228 proteins possibly implicated in ILI biosynthesis. Fluorescence microscopy of strains overexpressing eight ILI-associated proteins (IAP) candidates fused to superfolder green fluorescent protein showed co-localization with ILI. Genetic inactivation of these potential IAP-encoding genes and subsequent lipid analysis emphasized the importance of MAB_3486 and MAB_4532c as key enzymes influencing triacylglycerol storage. This study underscores the dynamic process of ILI biogenesis and advances our understanding of lipid metabolism in pathogenic mycobacteria. Identifying major IAP in lipid accumulation offers new therapeutic perspectives to control the growth and persistence of pathogenic mycobacteria.

IMPORTANCE
This study sheds light into the complex process of intracellular lipid accumulation and storage in the survival and persistence of pathogenic mycobacteria, which is of clinical relevance. By identifying the proteins involved in the formation of intrabacterial lipid inclusions and revealing their impact on lipid metabolism, our data may lead to the development of new therapeutic strategies to target and control pathogenic mycobacteria, potentially improving outcomes for patients with mycobacterial infections.
Blanking on blanks: few insect microbiota studies control for contaminants
Authors: Williamson Elisabeth M. , Hammer Tobin J. , Hogendoorn Katja , Eisenhofer Raphael 
Published: February 2025

ABSTRACT

Research on insect-microbe relationships is booming, with DNA sequencing being the most commonly used method to describe insect microbiota. However, sequencing is vulnerable to contamination, especially when the sample has low microbial biomass. Such low-biomass samples are common across insect taxa, developmental stages, and tissue types. Identifying putative contaminants is essential to distinguish between true microbiota and introduced contaminant DNA. It is therefore important that studies control for contamination, but how often this is done is unknown. To investigate the status quo of contamination control, we undertook a systematic literature review to quantify the prevalence of negative control usage and contamination control across the literature on insect microbiota (specifically bacterial communities) over a 10 year period. Two-thirds of the 243 insect microbiota studies evaluated had not included blanks (negative controls), and only 13.6% of the studies sequenced these blanks and controlled for contamination in their samples. Our findings highlight a major lack of contamination control in the field of insect microbiota research. This result suggests that a number of microbes reported in the literature may be contaminants as opposed to insect-associated microbiota and that more rigorous contamination control is needed to improve research reliability, validity, and reproducibility. Based on our findings, we recommend the previously developed guidelines outlined in the RIDE checklist, with the addition of one more guideline. We refer to this as the RIDES checklist, which stands for Report methodology, Include negative controls, Determine the level of contamination, Explore contamination downstream, and State the amount of off-target amplification.

IMPORTANCE
Our systematic review reveals a major lack of methodological rigor within the field of research on insect-associated microbiota. The small percentage of studies that control for contamination suggests that an unknown but potentially considerable number of bacteria reported in the literature could be contaminants. The implication of this finding is that true microbiota may be masked or misrepresented, especially in insects with low microbial biomass.
Inter-protomer opening cooperativity of envelope trimers positively correlates with HIV-1 entry stoichiometry
Authors: Katte Revansiddha H. , Xu Wang , Han Yang , Hong Xinyu , Lu Maolin 
Published: February 2025

ABSTRACT

HIV-1 entry to host cells is fulfilled by fusion, mediated by surface glycoprotein envelope (Env) trimers upon interaction with host receptors. The entry stoichiometry (T) defining the number of Env trimers required for fusion remains elusive. Stoichiometry was previously estimated using mathematical modeling of infectivity curves of pseudoviruses surface-decorated with heterotrimers containing wild-type and entry-deficient Env. Nevertheless, previous models rarely co-considered inter-protomer opening cooperativity (S, reflecting how CD4-induced conformational changes in one protomer affect the opening of adjacent protomers) and virion trimer number distributions, while experiments were limited to pseudoviruses. Here, we factored these two parameters into our models and included replication-competent virions. We provided simultaneous estimates of T and S under varying trimer number distributions and offered 2D stoichiometry maps for different Env strains. Our results depicted the interplay between viral infectivity and stoichiometry tuned by the number of trimers per virion. The estimates for all tested Env strains were prevalently higher (T ≥ 7 for BG505 or JR-FL, T ≥ 13 for NL4-3) than reported. A high degree of inter-protomer opening cooperativity was observed for the neutralization-sensitive NL4-3, while neutralization-resistant BG505 and JR-FL showed a low to intermediate degree. Entry stoichiometry and opening cooperativity were strikingly positive-correlated, implying tied inter-protomer and inter-Env cooperative interactions. Our findings provided an in-depth view of Env cooperativities during HIV-1 entry.

IMPORTANCE
The sparsely distributed envelope (Env) trimers on the surface of HIV-1 work collaboratively to mediate viral entry into the host, the early step of infection. The number of interacting trimers with host receptors required for entry awaits elucidation. Here, we explored the cooperative interplay among and within Env trimers, shedding light on a previously overlooked dimension of HIV-1 entry. For the first time, we presented distributions of estimated parameters depicting the number of Env trimers and degrees of inter-protomer opening cooperativities using biologically relevant mathematic models combined with virion infectivity measurements. Our results demonstrated that the quantity of required functional trimers positively correlates with inter-protomer opening cooperativity, a feature conserved across various strains. Our findings underscore cooperative behavior as an inherent characteristic of Env dynamics during HIV-1 entry. These insights enhance our understanding of HIV-1 infection mechanisms and could inform strategies for developing effective inhibitors or neutralizing agents.
Heparanase, a host gene that potently restricts retrovirus transcription
Authors: Waxman Brandon , Salka Kyle , Timilsina Uddhav , Umthong Supawadee , Shukla Deepak , Stavrou Spyridon 
Published: February 2025

ABSTRACT

Heparanase (HPSE) is a heterodimeric β-D-glucuronidase that is critical in mammalian cells for the enzymatic cleavage of membrane-associated heparan sulfate moieties. Apart from its enzymatic function, HPSE has important non-enzymatic functions, which include transcriptional regulation, chromatin modification, and modulation of various signaling pathways. Interestingly, while
HPSE
is an interferon-stimulated gene, past reports have shown that it has proviral properties for many different viruses, including herpes simplex virus 1, as it assists virus release from infected cells. However, as of yet, no antiviral functions associated with HPSE have been described. Here, we show that HPSE utilizes a hitherto unknown mechanism to restrict retroviruses by targeting the step of proviral transcription. Moreover, we demonstrate that HPSE blocks transcription initiation by targeting the SP1 transcription factor. Finally, we illustrate that the antiretroviral effect of HPSE is independent of its enzymatic activity. This report describes a novel antiviral mechanism utilized by HPSE to inhibit retrovirus infection.

IMPORTANCE
Heparanase (HPSE) has emerged as an important factor that has proviral functions for a number of viruses, including herpes simplex virus and hepatitis C virus, by assisting in virus egress. However, HPSE is an interferon-stimulated gene and, thus, is a part of the host antiviral defense. Nothing is known about the antiviral functions of HPSE. Here, we examine in depth the role of HPSE during retrovirus infection using two retroviruses, human immunodeficiency virus type 1 (HIV-1) and murine leukemia virus. In this report, we show that mouse, but not human, HPSE blocks retrovirus infection by targeting provirus transcription. HPSE sequesters the SP1 transcription factor away from the proviral promoter, thereby inhibiting transcription initiation. In conclusion, our findings identify a novel antiviral function of HPSE and its potential role as an inhibitor of zoonotic transmission of retroviruses.