2026-04-01
Phillip Wulfridge, Nathaniel Rell, John Doherty, Kuo-Chen Fang, Michelle Lee Lynskey, Kavitha Sarma
by Phillip Wulfridge, Nathaniel Rell, John Doherty, Kuo-Chen Fang, Michelle Lee Lynskey, Kavitha Sarma Transition from a pluripotent to a differentiated cell state is accompanied by significant changes in genome organization. Activity dependent neuroprotective protein (ADNP) is a chromatin regulator with critical roles in neurodevelopment and limits the genomic occupancy of CTCF, a master architectural protein in genome organization, in embryonic stem cells. However, ADNP localization, function, and relationship with CTCF in differentiated neural lineages are not well studied. Here we develop a dual degron model which allows us to acutely deplete ADNP in neural progenitor cells (NPCs). We find that ADNP depletion does not impact NPC survival in the short term, but results in a genome organization switch, which favors the formation of short-range chromatin looping interactions coinciding with CTCF accumulation. Furthermore, ADNP localizes to active gene promoters in NPCs that are unoccupied by CTCF, where it prevents over-expression of genes that are activated upon neurodifferentiation and represses those involved in commitment to other lineages. Our findings uncover CTCF-dependent as well as CTCF-independent regulatory mechanisms of ADNP in NPC-specific chromatin organization and gene expression programs that may underlie its essential function in neurodevelopment.
DOI: 10.1371/journal.pgen.10120812026-03-31
Anne Geertje Oostlander, Marcel René Schumann, Lina Strzelczyk, Lucas Well, Ulrike Brandt, Marco Leiterholt, Stefan Jakschies, Tina Rietschel, Josef Wissing, Lothar Jänsch, André Fleißner
by Anne Geertje Oostlander, Marcel René Schumann, Lina Strzelczyk, Lucas Well, Ulrike Brandt, Marco Leiterholt, Stefan Jakschies, Tina Rietschel, Josef Wissing, Lothar Jänsch, André Fleißner Cell fusion is a fundamental process essential for the development and proliferation of eukaryotic organisms. In the ascomycete fungus Neurospora crassa , germinating spores undergo chemotropic interactions and fusion to merge into a supracellular unit, which gives rise to the mycelial colony. Within mature colonies, hyphal branches fuse to form anastomoses between leading hyphae, enhancing the overall connectivity of the mycelium. Both germling and hyphal fusion rely on the same molecular machinery. The MAP kinase MAK-2 and the fungal-specific protein SO have been identified as key regulators of these processes, and their alternating recruitment to the plasma membrane at interacting cell tips suggests a dialog-like cell communication mechanism involving dynamic switches between signal sending and receiving. However, the mechanisms that trigger the onset of this intercellular communication are still not understood. This study identifies EOP-1 as an interaction partner of the SO protein and functionally characterizes its role in cell communication and fusion. Deletion of the eop-1 gene abolished germling fusion and chemotropic interactions, while live-cell imaging showed EOP-1 oscillating at interacting cell tips, coinciding with SO recruitment. Intriguingly, EOP-1 displayed a similar dynamic, oscillatory tip recruitment also in isolated, non-interacting germlings, setting it apart from previously characterized fusion factors in N. crassa . This observation suggests for the first time that spore germlings of N. crassa exhibit fusion related cell-autonomous oscillatory behavior and implicates EOP-1 in initiating intercellular communication. The oscillatory recruitment pattern of EOP-1 was dependent on the presence of SO, MAK-1, MAK-2, BEM1 and HAM-14 in the cell. Loss of EOP-1 strongly reduced MAK-1 phosphorylation, placing EOP-1 upstream of MAK-1 pathway activation. This work offers new insight into how genetically and developmentally identical cells initiate and coordinate their communication and mutual attraction.
DOI: 10.1371/journal.pgen.10120872026-03-31
Ying-Yu Chen, Lea S. Steglich, Nadja Spasovski, Marcel H. W. Franzius, Merle Aden, Isabel Maurus, Rebekka Harting, Gerhard H. Braus
by Ying-Yu Chen, Lea S. Steglich, Nadja Spasovski, Marcel H. W. Franzius, Merle Aden, Isabel Maurus, Rebekka Harting, Gerhard H. Braus Filamentous fungi produce a wide range of secondary metabolites to adapt to changing environments. RNA sequencing revealed that nine biosynthetic gene clusters (BGCs) of the phytopathogenic Verticillium dahliae react to different nutrient environments. The adapt-to-nutrient NRPS-like ( ANN ) cluster contributes to antibacterial activity and developmental processes important for the early biotrophic life cycle, but is dispensable for virulence on tomato ( Solanum lycopersicum ). Transcription of the core biosynthetic enzyme-encoding ANN3 is highly induced in nutrient-poor environment. ANN3 is transcriptionally controlled by global and in-cluster transcription factors. ANN3 is activated by early colonisation transcription factors Som1 and Vta2, but repressed by Mtf1, which governs late stages of disease progression. The in-cluster transcription factor Ann1, which represses ANN3 , is less abundant in nutrient-poor environment or when V. dahliae encounters antagonists. Ann1 promotes resting structure formation but suppresses conidiation and antibacterial activity. Possible products of the ANN cluster were revealed by comparing metabolites extracted from ANN3 regulator mutants and from the bacterial-fungal interaction zone. Our findings revealed that V. dahliae perceives different nutrient environments and changes its survival strategy by differential expression of the ANN secondary metabolite gene cluster.
DOI: 10.1371/journal.pgen.10119302026-03-30
Amamah Farzlin Farnaz, Sameer Joshi, Praseetha Sarath, Girija Jogwar, Koodali T. Nishant
by Amamah Farzlin Farnaz, Sameer Joshi, Praseetha Sarath, Girija Jogwar, Koodali T. Nishant Meiotic crossovers are generated from the repair of programmed DNA double-strand breaks (DSBs). In the budding yeast Saccharomyces cerevisiae and mammals, most crossovers are generated through the Class I pathway, involving the mismatch-repair related complex Msh4-Msh5, while a smaller fraction is produced by the Mms4-Mus81 endonuclease (Class II pathway). We present the first report on the genome-wide localization of the Mms4 protein during meiosis in S. cerevisiae . Surprisingly, Mms4 localization showed a trend towards weak DSB sites, unlike the localization of the Class I crossover protein -Msh5, which is biased towards strong DSB sites. This preference for weaker DSB hotspots was retained in a msh5 ∆ mutant, arguing against competitive models of Mms4 and Msh5 association on meiotic chromosomes. The chromosomal association of Mms4 does not require the formation of meiotic DNA breaks but is facilitated by chromosome axis assembly. These results suggest Mms4 is primarily associated with chromosomal axis regions positioned near recombination intermediates. Mms4 binding is also largely insensitive to heterozygosity, unlike Msh5, consistent with its independence from recombination for localization. Together, these findings support a model in which Mms4-Mus81 enhances the robustness of meiotic recombination with a trend towards binding DSB hotspots that are weaker or are located in regions with sequence divergence that may be processed less efficiently by the Class I pathway.
DOI: 10.1371/journal.pgen.10120972026-03-30
Logan B. Suits, Sebastian J. Khan, Dipanwita Bhattacharya, Silviya Dimitrova, Prahathees J. Eswara
by Logan B. Suits, Sebastian J. Khan, Dipanwita Bhattacharya, Silviya Dimitrova, Prahathees J. Eswara The metabolic networks of most life forms integrate cost-benefit analysis to properly budget carbon and other essential nutrients. Bacillus subtilis is a Gram-positive model bacterium found in diverse ecological niches such as soil, marine environments, and the human gut. As such, B. subtilis cells fine-tune metabolic pathways by monitoring signals indicating the presence of nutrients and stressors. A highly conserved protein, GlmR, is a key player in rationing carbon for the production of cell envelope precursors. This function of GlmR can be attributed to its role in cell shape regulation and antibiotic resistance. Given its central position in carbon utilization, GlmR is under post-translational regulation by phosphorylation and UDP-N-acetylglucosamine (UDP-GlcNAc) binding. GlmR is also linked to cyclic-di-AMP (c-di-AMP), a nucleotide second messenger involved in osmotic and cell wall stress response. In this study, we probed the importance of GlmR in cell morphogenesis, c-di-AMP signaling, and investigated the physiological significance of post-translational regulation. Our results reveal that cells lacking glmR exhibit: (i) increased susceptibility to tunicamycin, a cell envelope targeting antibiotic; (ii) impaired division site positioning; and (iii) reduced intracellular c-di-AMP concentration. Furthermore, we show that the function of GlmR is fine-tuned by UDP-GlcNAc binding, phosphorylation, and acetylation. Additionally, we provide evidence showing that the recently discovered uridyltransferase activity of GlmR is integral for its function. We show that GlmR is a cell width determinant and propose a model suggesting close cooperation with an actin-like protein, MreB. Overall, our studies highlight the importance of the enzymatic function of GlmR and elucidate the mechanism behind the multiple post-translational means to regulate this crucial protein which is at the crux of carbon flux with an important role in maintaining cell envelope integrity.
DOI: 10.1371/journal.pgen.10120962026-03-24
Maddy Comerford, Davide M. Vespasiani, Navya Shukla, Laura E. Cook, Danat Yermakovich, Michael Dannemann, Matthew Leavesley, Christopher Kinipi, François-Xavier Ricaut, Nicolas Brucato, Murray P. Cox, Irene Gallego Romero
by Maddy Comerford, Davide M. Vespasiani, Navya Shukla, Laura E. Cook, Danat Yermakovich, Michael Dannemann, Matthew Leavesley, Christopher Kinipi, François-Xavier Ricaut, Nicolas Brucato, Murray P. Cox, Irene Gallego Romero Interbreeding between anatomically modern humans and archaic hominins has contributed to the genomes of present-day human populations. However, our understanding of the specific gene regulatory consequences of Neanderthal, and particularly, Denisovan introgression is limited. Here, we used a massively parallel reporter assay to investigate the regulatory effects of 25,869 high-confidence introgressed SNPs segregating in present-day individuals of Papuan genetic ancestry in immune cell types. Overall, 8.22% of Denisovan and 8.58% of Neanderthal sequences showed active regulatory activity, and 9.22% of these displayed differential activity between archaic and modern alleles. We found no association between introgressed allele frequency on activity regardless of introgression source, but introgressed Denisovan alleles at higher frequencies were less likely to be differentially active than expected, suggesting introgression is under some degree of selective constraint. Both activity and differentially activity were associated with distance to the nearest transcription start site, while differential activity was additionally associated with differential transcription factor binding. Genes predicted to be regulated by differentially active sequences included IFIH1 and TNFAIP3 , key immune genes and known examples of archaic introgression. Overall, this work provides experimental validation of regulatory activity for thousands of archaic variants in populations with the highest levels of Denisovan ancestry worldwide, revealing how human evolutionary history actively shapes present-day genetic diversity and immune function.
DOI: 10.1371/journal.pgen.10120672026-03-17
Kyle Leix, Candilianne Serrano-Zayas, Hitarthi S. Vyas, Sarah E. Graham, Brian T. Emmer
by Kyle Leix, Candilianne Serrano-Zayas, Hitarthi S. Vyas, Sarah E. Graham, Brian T. Emmer Regulation of LDLR gene expression plays an important role in the development of atherosclerotic diseases including heart attack and stroke. Although LDLR regulation by sterol response elements has been well characterized, the functional significance of other noncoding regions at the LDLR locus remains poorly defined. In this study, we developed and applied a high throughput CRISPR screen to test the functional importance of candidate LDLR cis -regulatory elements (CREs) in their native genomic context. In total, we found 25 discrete regions to exhibit a significant impact on LDLR expression. For one of these regions with particularly strong activity in the first intron, we validated the presence of an enhancer by confirming that its disruption reduced endogenous LDLR expression while its insertion upstream of a minimal promoter augmented reporter gene expression. We then applied a massively parallel reporter assay to fine map enhancer activity within this region to a 129 bp interval that is highly conserved among vertebrates, exhibits biochemical hallmarks of enhancer activity, is enriched for transcription factor binding motifs, and contains a common genetic variant (rs57217136) that has been associated with human LDL cholesterol levels by genome-wide association studies. Overall, these findings demonstrate the power of CRISPR screening to interrogate candidate CREs and clarify the functional landscape of noncoding sequences at the LDLR locus.
DOI: 10.1371/journal.pgen.10120822026-03-17
Marleen T. Aarts, Anna Nordin, Claudio Cantù, Antonius L. van Boxtel, Renée van Amerongen
by Marleen T. Aarts, Anna Nordin, Claudio Cantù, Antonius L. van Boxtel, Renée van Amerongen Gene expression is controlled by complex transcriptional networks in which transcription factors and their cognate enhancer elements integrate developmental and environmental cues. The progesterone receptor (PR), a hormone-activated transcription factor, is essential for breast development and physiology, yet how it engages with the chromatin and lineage-specific cofactors remains unclear. Using an unbiased approach, we identify the epithelial transcription factor grainyhead-like 2 (GRHL2) as a key co-regulator of PR activity in hormone responsive breast cancer cells. We show that GRHL2 interacts with PR in a progesterone-independent manner. Upon progesterone stimulation, GRHL2 and PR are both recruited to distal enhancer elements of target genes. Furthermore, GRHL2- and PR-bound elements connect spatially through chromatin looping to regulate shared targets. These findings uncover a previously unrecognized mechanism by which GRHL2 and PR coordinate gene regulation through both chromatin binding and 3D genome architecture modification, positioning GRHL2 as a crucial modulator of steroid hormone receptor function.
DOI: 10.1371/journal.pgen.10120882026-03-13
Ying-Ju Lin, Ting-Yuan Liu, Jai-Sing Yang, Ju-Pi Li, Jian-Shiun Chiou, Hsing-Fang Lu, Kuyuri Ariyoshi, Keiko Hikino, Chikashi Terao, Chen-Hsing Chou, Wen-Miin Liang, I-Ching Chou, Ting-Hsu Lin, Chiu-Chu Liao, Shao-Mei Huang, Fuu-Jen Tsai
by Ying-Ju Lin, Ting-Yuan Liu, Jai-Sing Yang, Ju-Pi Li, Jian-Shiun Chiou, Hsing-Fang Lu, Kuyuri Ariyoshi, Keiko Hikino, Chikashi Terao, Chen-Hsing Chou, Wen-Miin Liang, I-Ching Chou, Ting-Hsu Lin, Chiu-Chu Liao, Shao-Mei Huang, Fuu-Jen Tsai Both genetic and environmental factors affect human stature, including overall height and familial short stature (FSS), and it is associated with various health outcomes. However, the study of genetic connections between stature and health conditions remains lacking in East Asian populations. Hence, we conducted parallel genome-wide association studies (GWAS) of body height and FSS in the Han Taiwanese population, aiming to elucidate the genetic influences of stature on health and facilitate the formulation of precision-health strategies. We analyzed large-scale GWAS data on adult height (120,301 Han Taiwanese) and FSS (FSS; 2,050 cases, 27,966 controls) to examine cross-trait genetic correlations across five East Asian biobanks, and applied phenome-wide association studies (PheWAS) and polygenic risk score (PRS) analyses to assess clinical outcomes using Cox proportional hazard models and Kaplan–Meier analyses. We identified 293 loci for height and five for FSS, with cross-biobank genetic correlations linking stature to body size, lung function, and cardiovascular/reproductive traits (atrial flutter/fibrillation [AF], menarche, and endometriosis). PheWAS showed that height PRS increased risks of AF and endometriosis, while FSS PRS had a protective effect against endometriosis. MR analyses showed that taller stature increased AF risk independently and endometriosis risk through menarche/weight, while shorter stature had a weak protective effect against endometriosis. Survival analyses showed the association of higher height PRS with greater AF risk and an earlier divergence of cumulative incidence curves. These time-to-event patterns were consistently replicated using meta-analysis–derived PRSs. The findings highlight stature-related genetic determinants, associated health outcomes, and polygenic risk scores as effective tools for early risk prediction and precision health strategies in East Asian populations.
DOI: 10.1371/journal.pgen.10120302026-03-13
Jian Yi Kok, Zachary H. Harvey, Elin Axelsson, Frédéric Berger
by Jian Yi Kok, Zachary H. Harvey, Elin Axelsson, Frédéric Berger Maintaining transcriptional fidelity is essential for precise gene regulation and genome stability. Despite this, cryptic antisense transcription, occurring opposite to canonical coding sequences, is a pervasive feature across all domains of life. How such potentially harmful cryptic sites are regulated remains incompletely understood. Here, we show that nucleosome arrays within gene bodies play a key role in suppressing cryptic transcription. Using the fission yeast Schizosaccharomyces pombe as a model, we demonstrate that the CHD-family chromatin remodeler Hrp3 coordinates with the transcription elongation machinery, via the transcriptional regulator Prf1/RTF1, to position nucleosomes at sites of cryptic transcription initiation within gene bodies. In the absence of Hrp3, AT-rich sequences within gene bodies lose nucleosome occupancy, exposing promoter-like sequences that drive cryptic initiation. While cryptic transcription is generally detrimental, we identify a subset of antisense transcripts that encode critical meiotic genes, suggesting that cryptic transcription can also serve as a source of regulatory innovation. These findings define an elongation‑coupled chromatin pathway that preserves transcriptional fidelity and reveal how nucleosome remodeling shapes antisense transcription, cellular homeostasis, and adaptive potential.
DOI: 10.1371/journal.pgen.10120782026-03-13
Daniel M. Weinreich, Tom Sgouros, Yevgeniy Raynes, Hlib Burtsev, Edison Chang, Sanyu Rajakumar, Ignacio G. Bravo, Csenge Petak
by Daniel M. Weinreich, Tom Sgouros, Yevgeniy Raynes, Hlib Burtsev, Edison Chang, Sanyu Rajakumar, Ignacio G. Bravo, Csenge Petak Classical population genetics provides a robust, quantitative framework for modeling how natural selection acts on alleles that influence phenotypes with invariant fitness consequences for their carriers, such as running speed or drug resistance. By contrast, modifier theory considers the evolution of alleles that influence population genetic parameter values in their carriers, such as mutation or recombination rates. This is a more complicated problem. First, the fitness effects of modifier alleles reflect independently realized stochastic phenotype perturbations they induce in their carriers. And second, the association between modifier alleles and their induced phenotypes can decay over generations. Consequently, general results in modifier theory have been few. Here, we propose recasting modifier theory as exploring the evolution of alleles that influence the amount of stochasticity in inheritance, be it genetic, epigenetic, cytoplasmic or somatic transmission. We then present a toy model that predicts the existence of a selectively optimal amount of such “reproductive noise,” which depends on the rate of environment change, the timescale of association between noise allele and induced phenotype, and population size. Next, we suggest that the same framework can be applied to the evolution of alleles that influence “developmental noise,” i.e., the amount of stochastic phenotypic variation among genetically identical organisms reared in identical environments. This theoretical connection is timely, because high throughput assays are now demonstrating widespread heritability in the amount of developmental noise. Our approach also resolves the long-standing teleological criticism of the hypothesis that evolvability can evolve by natural selection. Taken together, this work demonstrates the opportunities for a robust, quantitative population genetic theory of alleles that influence the amount of biological noise.
DOI: 10.1371/journal.pgen.10120662026-03-12
Jun Huang, Anna E. Lehmann, Patricia P. Peterson, Ziyan Xu, Liping Xiong, Sheng Sun, Joseph Heitman
by Jun Huang, Anna E. Lehmann, Patricia P. Peterson, Ziyan Xu, Liping Xiong, Sheng Sun, Joseph Heitman Sex-specific homeodomain (HD) proteins are key regulators of cell identity and sexual development in fungi, typically functioning as heterodimers to govern transcription. In the human fungal pathogens Cryptococcus neoformans and Cryptococcus deneoformans , the HD proteins Sxi1α and Sxi2a (Sex-inducer 1α and 2a) have been characterized as interacting components that play critical roles in sexual development during α x a sexual reproduction. α cells are the predominant mating type in natural populations of Cryptococcus , and unisexual (same-sex) mating can also occur in certain genetic backgrounds. The roles of Sxi1α and Sxi2a in unisexual reproduction are not fully understood. To elucidate the functions of Sxi1α and Sxi2a, we first applied AlphaFold3 prediction, which identified potential heterodimeric and homodimeric complexes. Formation of a Sxi2a homodimer was then experimentally validated through yeast two-hybrid assays. We subsequently deleted SXI1 α and SXI2 a in the hyper-filamentous self-fertile C. deneoformans strains XL280α and XL280a. Disruption of these genes did not result in noticeable defects in vegetative growth, virulence-associated traits, colony morphology, sporulation, or competitive fitness during unisexual crosses. Interestingly, both bilateral (mutant x mutant) and unilateral (mutant x wild type) crosses involving the sxi1 αΔ mutant showed significantly increased α-α cell fusion efficiency, suggesting a previously unrecognized inhibitory role for Sxi1α in regulating same-sex cell fusion. Consistently, genes encoding mating pheromones and the α-pheromone receptor Ste3 were upregulated in the sxi1 αΔ fusion assays. Transcriptomic analysis of sxi1 αΔ and sxi2 aΔ mutants led to the identification of unique subsets of genes negatively regulated by each transcription factor during unisexual reproduction. Additionally, α x a crosses between sxi1 αΔ and sxi2 aΔ mutants revealed differential regulation of mating-type ( MAT ) loci genes dependent only on Sxi1α or Sxi2a. Together, our findings reveal a novel role for Sxi1α in governing cell fusion and demonstrate that Sxi1α and Sxi2a have distinct transcriptional control during unisexual and α x a sexual reproduction, potentially exerting opposing regulation of sex-specific MAT genes.
DOI: 10.1371/journal.pgen.10120842026-03-09
Suman Maharjan, Ryan Sloan, Jada Lusk, Rose Bevienguevarr, Jacob Surber, Randy M. Morgenstein
by Suman Maharjan, Ryan Sloan, Jada Lusk, Rose Bevienguevarr, Jacob Surber, Randy M. Morgenstein The bacterial actin-homolog MreB is a crucial component of the Rod-system (elongasome) that maintains rod shape in many bacteria. It is localized beneath the cytoplasmic membrane, where it organizes the elongasome complex. Depletion or deletion of mreB results in loss of rod shape and cell death; however, the mechanism of how MreB operates is not known. Past studies have reported that mutations in mreB cause varying degrees of cell shape and size alterations based on the type and position of the substitution. To better understand the role of MreB in rod shape formation we have taken the first truly systematic approach by replacing the native copy of mreB with an alanine-scanning mutagenesis library. Surprisingly, we observed stably growing spherical mutants that have lost MreB’s function(s) for shape regulation without losing viability. Hence, MreB has vital functions related to growth in addition to shape maintenance that can be separated. In support of this, rod shape suppressor analysis of these spherical mutants only revealed reversions or intragenic mreB mutations, suggesting that MreB is indispensable for rod shape. Additionally, our results imply the elongasome is no longer active in these strains, suggesting a novel way for rod shaped bacteria to synthesize cell wall.
DOI: 10.1371/journal.pgen.1012070