Biology subjects

Singh, K.

Publications and source records attributed to Singh, K..

At least 55 records · Page 3Linked to original sources

bioRxiv2025

A 3D iPSC retina model reveals non-cell-autonomous and non-neuronal mechanism of photoreceptor degeneration in a lysosomal storage disorder.

Disruption of photoreceptor-retinal pigment epithelium (RPE) interface with loss of photoreceptor outer segments (POSs) in the retina is a pathological hallmark of several neurodegenerative and retinal diseases including lysosomal storage disorders like CLN3 disease. However, the retina is a functional composite in vivo; and in vitro stem cell models of retina that enable investigation of the photoreceptor-RPE interface in healthy and diseased retina are lacking. Here, we developed a 3D human pluripotent stem cell (hPSC)-derived retina model to investigate the photoreceptor-RPE interface in healthy and disease tissue. Using this 3D hPSC retina model, we demonstrated that the most common disease causing CLN3 mutation (CLN3{Delta}ex7-8) leads to reduced levels of acid ceramidase (AC) and consequently altered sphingolipid metabolism and signaling and POS loss in CLN3 disease. Consistent with the 3D hPSC retina model, altered sphingolipid metabolism and signaling coincided with POS loss in a large animal model of CLN3 disease, CLN3 miniswine. Therapeutically, recombinant human acid ceramidase (rhAC) targeted both altered sphingolipid metabolism and retina degeneration in the CLN3 hPSC retina model and the CLN3 miniswine eye. These findings demonstrate a proof-of-concept that rhAC can rescue disease phenotype in a large animal model of CLN3 disease and suggest that rhAC could be a therapeutic approach for CLN3 disease. One Sentence SummaryAcid ceramidase deficiency and consequently altered sphingolipid signaling promotes disease phenotype(s) in a lysosomal storage disorder, CLN3 disease.

neuroscience↗

bioRxiv2025

A Novel FNDC1-NAMPT-NAD axis is Implicated in Small and Large-vessel Arterial Disease and Drives Vascular Calcification

Vascular calcification represents a convergent pathological feature of diverse cardiovascular diseases, yet the upstream molecular programs orchestrating this process remain poorly defined. Here, we uncover fibronectin type III domain-containing 1 (FNDC1) as a previously unrecognized regulator of vascular calcification across both microvascular and macrovascular beds. Integrative transcriptomic profiling of human calciphylaxis lesions and atherosclerotic coronaries identified FNDC1 as one of the most significantly upregulated genes. In primary human vascular smooth muscle cells, FNDC1 drove osteogenic phenotype switch and vascular calcification through activation of PI3K/AKT signaling and metabolic reprogramming. Mechanistically, FNDC1 directly binds to nicotinamide phosphoribosyltransferase (NAMPT) resulting in elevated intracellular NAD levels, thus coupling vascular signaling to control of NAD biosynthesis. In murine models, genetic deletion of Fndc1 or pharmacologic inhibition of NAMPT suppressed arterial calcification and prolonged survival. Clinically, circulating FNDC1 levels were elevated in patients with both calciphylaxis and coronary artery disease and independently predicted cardiovascular risk in 42,687 UK Biobank participants. Together, these findings establish FNDC1 as a central mediator of vascular pathology and highlight the FNDC1- NAMPT-NAD+ axis as a promising target for therapeutic intervention.

cell biology↗

bioRxiv2025

Genomic analysis uncovers unique haplotype signatures from subspecies and agronomic types associated with blanchability in groundnut

Blanchability, defined as the ease of seed coat removal after roasting, is a vital trait for enhancing processing efficiency and product quality in groundnut (Arachis hypogaea L.). To enable a comprehensive haplotype-level genetic dissection of this trait, SNPs derived from whole-genome resequencing (WGRS) were used to perform genome-wide association study (GWAS) using multi-locus models (BLINK and FarmCPU) on a diverse groundnut mini-core collection phenotyped across two crop seasons. A total of 26 significant SNP-trait associations (STAs) were identified across multiple chromosomes, with major loci on chromosomes Ah05, Ah06, and Ah17, some of which were further validated using KASP (Kompetitive Allele-Specific PCR) markers. Candidate genes, such as those involved in cell wall biosynthesis (e.g., galactoside 2-alpha-L-fucosyltransferase-like protein, protein kinase superfamily members, and glycerophosphoryl diester phosphodiesterase 3), were found to be within linkage disequilibrium (LD) of the identified STAs, suggesting their plausible association with blanchability. Haplo-pheno analyses identified superior high-blanchability haplotypes; Ah05HapBL3, Ah06HapBL5, Ah06HapBL10, and Ah17HapBL6, which were predominantly found in the fastigiata subspecies (including Valencia and Spanish bunch agronomic types) from South Asia and South America, while the low-blanchability haplotypes were from hypogaea subspecies (including Virginia runner and Virginia bunch agronomic types) from Africa. Overall, this study provides valuable insights for customizing blanchability through haplotype-based breeding of processing-grade cultivars thereby improving groundnut value chains to meet diverse industrial demands.

genomics↗

bioRxiv2025

Genomic analysis reveals the interplay between ABA-GA in determining dormancy duration in groundnut

Groundnut is an important leguminous crop however; its productivity and seed quality are frequently reduced due to lack of fresh seed dormancy (FSD). To address this challenge, a mini-core collection of 184 accessions was phenotyped to identify donors in each agronomic type, in addition to analysing data on whole genome re-sequencing and multi-season phenotypic evaluations to identify stable marker-trait associations (MTAs) associated with FSD. Phenotypic analysis revealed substantial variability in dormancy durations, with days to 50% germination (DFG) ranging from 1 to 30 days. Multi-locus genome-wide association studies (ML-GWAS) identified 27 MTAs in individual seasons and 12 MTAs in pooled seasons data, respectively. Key candidate genes identified included Cytochrome P450 superfamily proteins, protein kinase superfamily proteins, and MYB transcription factors involved in the Abscisic acid (ABA) pathway, as well as F-box interaction domain proteins, ATP-binding ABC transporters, associated with the Gibberellic acid (GA) pathway. SNP-based KASP (Kompetitive Allele-Specific Polymerase chain reaction) markers for 12 SNPs were developed and validated, of these 6 markers (snpAH00577, snpAH00580, snpAH00582, snpAH00585, snpAH00586 and snpAH00588) showed polymorphism between dormant and non-dormant lines. Incorporating favourable dormant alleles into breeding strategies could enable the development of high-yielding cultivars with a dormancy period of 2-3 weeks.

genomics↗

bioRxiv2025

Heme drives cardiac endothelial senescence in sepsis via STING activation

Sepsis-induced cardiac dysfunction is a major contributor to sepsis-related mortality, and many patients continue to experience long-term cardiac complications after recovery. Here, we demonstrate that cardiac senescence is a key feature of sepsis-associated cardiac dysfunction, with endothelial cells identified as the predominant senescent population in septic cardiac tissue. Yet, the pathogenic drivers of endothelial senescence in sepsis remain poorly characterized. Among potential mediators, we found that elevated levels of heme, a byproduct of hemolysis, strongly correlate with increased endothelial senescence and impaired cardiac function. Mechanistic studies revealed that heme acts as a novel ligand for STING, exacerbating bacterial infection induced STING polymerization and activation, thereby promoting endothelial senescence. Notably, either STING inhibition or enhanced heme clearance via increased hemopexin expression significantly alleviated cardiac endothelial senescence and facilitated cardiac functional recovery in septic mice. These findings identify heme as a critical pathogenic driver of endothelial senescence and highlight heme clearance as a promising therapeutic strategy for mitigating sepsis induced cardiac dysfunction.

cell biology↗

bioRxiv2025

Identification of High Blanchability Donors, Candidate genes and Markers in Groundnut

Blanchability in groundnut, the ability of seeds to shed their seed coat (testa), is a trait of economic importance in the food processing industry, yet remains underexplored in breeding programs. This study aimed to assess blanchability in 184 accessions from the ICRISAT minicore collection and identify associated genomic regions, candidate genes, and molecular markers. Significant variability was observed over two seasons, with blanchability ranging from 3.98% to 70.08%. Ten genotypes, including ICG10890, ICG9507, ICG13982, and ICG297, exhibited good blanchability, with ICG297 emerging as a promising donor based on cluster analysis of blanchability and agronomic traits. Genome-wide association studies (GWAS) using the 58K Axiom_Arachis SNP array revealed 58 significant SNP-trait associations and important candidate genes such as isocitrate dehydrogenase and ubiquitin ligase, which influence seed coat structure and cell wall integrity. Nine SNPs were validated via allele mining, with four markers--on chromosomes A01 (snpAH00551), A06 (snpAH00554), B04 (snpAH00558), and B07 (snpAH00559), effectively distinguishing between high and low blanchability genotypes. These validated SNPs present valuable tools for genomics-assisted breeding. Overall, the findings is the first study contributing to a better understanding of the genomics and genetic basis of blanchability and offer resources for improving processing traits in groundnut.

genomics↗

bioRxiv2025

Effects of Natural Lithium and Lithium Isotopes on Voltage Gated Sodium Channel Activity in SH-SY5Y and IPSC Derived Cortical Neurons

Although lithium (Li) is a widely used treatment for bipolar disorder, its exact mechanisms of action remain elusive. Research has shown that the two stable Li isotopes, which differ in their mass and nuclear spin, can induce distinct effects in both in vivo and in vitro studies. Since sodium (Na+) channels are the primary pathway for Li+ entry into cells, we examined how Li+ affects the current of Na+ channels using whole-cell patch-clamp techniques on SH-SY5Y neuroblastoma cells and human iPSC-derived cortical neurons. Our findings indicate that mammalian Na+ channels in both neuronal models studied here display no selectivity between Na+ and Li+, unlike previously reported bacterial Na+ channels. We observed differences between the two neuronal models in three measured parameters (Vhalf, Gmax,z). We saw no statistically significant differences between any ions in SHSY-5Y cells, but small differences in the half-maximum activation potential (Vhalf) between Na+ and 6Li+ and between 7Li+ and 6Li+ were found in iPSC-derived cortical neurons. Although Na+ channels are widely expressed and important in neuronal function, the very small differences observed in this work suggest that Li+ regulation through Na+ channels is likely not the primary mechanism underlying Li+ isotope differentiation.

neuroscience↗

bioRxiv2025

Membrane remodeling by the Bacterial Dynamin-like Protein (BDLP) from Nostoc punctiforme

Membranes are fundamental to biological systems as they serve as barriers to compartmentalize the cell and its cytoplasm. They are constantly remodeled in shape and composition during the formation and maintenance of organelles. The large GTPase dynamins are well characterized for membrane shape remodeling in eukaryotes but their functions in prokaryotes are less characterized. Here, we determine the lipid-binding, enzymatic and membrane remodeling activities of the Bacterial Dynamin-Like Protein (BDLP) from the cyanobacterium Nostoc punctiforme. We find that BDLP binds phosphatidyl glycerol (PG)-containing membranes, but unlike other soluble dynamins, shows little stimulation in GTPase activity. This enzymatic feature is like that seen among the fusion dynamins to which BDLP shows structural similarities. FRET-based bulk vesicle fusion assays however reveal that BDLP is incapable of membrane fusion. To further understand BDLP functions, we turned to microscopic analysis of BDLP on membranes templates. We find that the GTP-bound BDLP forms a rigid scaffold that is capable of bending and imposing a defined curvature. In contrast, the GDP-bound state is loosely organized and unable to remodel membrane shape. Our results indicate that BDLP intrinsically functions as a protein scaffold whose organization is regulated by nucleotide binding.

biochemistry↗

bioRxiv2025

Kre6-Dependent β-1,6-glucan Biosynthesis Only Occurs in the Conidium of Aspergillus fumigatus

The structural role of {beta}-1,6-glucan has remained under-investigated in filamentous fungi compared to other fungal cell wall polymers, and previous studies have shown that the cell wall of the mycelium of A. fumigatus did not contain {beta}-1,6-glucans. In contrast, the current solid-state NMR investigations showed that the conidial cell wall contained a low amount of {beta}-1,6-glucan. ssNMR comparisons of the A. fumigatus and C. albicans {beta}-1,6-glucans showed they are structurally similar. Deletion of the KRE6 gene which is the only KRE gene in the A. fumigatus genome resulted in a mutant depleted of {beta}-1,6-glucan which has a growth phenotype similar to the parental strain. Even though it is not an essential polymer in A. fumigatus, {beta}-1,6-glucan play a role in cell wall organization since the kre6{Delta} mutant showed a higher sensitivity to Congo-red and Calcofluor white which are known to be general cell wall inhibitors. It is also another example of the significant structural differences seen between conidium and mycelium of filamentous fungi.

biochemistry↗

bioRxiv2025

Pathogenic variants in autophagy-tethering factor EPG5 drive neurodegeneration through mitochondrial dysfunction and innate immune activation

The autophagy-tethering factor, ectopic P-granule 5 autophagy protein (EPG5), plays a key role in autophagosome-lysosome fusion. Impaired autophagy associated with pathogenic variants in EPG5 cause a rare devastating multisystem disorder known as Vici syndrome, which includes neurodevelopmental defects, severe progressive neurodegeneration and immunodeficiency. The pathophysiological mechanisms driving disease presentation and progression are not understood. In patient-derived fibroblasts and iPS cells differentiated to cortical neurons, we found that impaired mitophagy leads to mitochondrial bioenergetic dysfunction. Physiological Ca2+ signals resulted in paradoxical mitochondrial Ca2+ overload attributed to downregulation of MICU1/3. Ca2+ signals caused mitochondrial depolarisation, mtDNA release and activation of the cGAS-STING pathway, reversed by pharmacological inhibition of the mitochondrial permeability transition pore (mPTP) or of the STING pathway. Thus, we have identified multiple potential therapeutic targets driving disease progression associated with pathogenic EPG5 mutations, including impaired mitochondrial bioenergetics, mitochondrial Ca2+ overload, vulnerability to mPTP opening and activation of innate immune signalling.

neuroscience↗

bioRxiv2025

Tracking Persistence and Dynamics of Active Soil Viruses with SIP-Viromics

Viruses are abundant and ecologically important in soils, yet the persistence and production dynamics of extracellular virions remain poorly understood. We applied a genome-resolved stable isotope probing viromics (SIP-viromics) approach, combining H 18O labeling with viral metagenomics, to track virion turnover in seasonally dry grassland soils following rewetting. We identified 354 viral populations (vOTUs) using individual-sample and combined metagenome assemblies. Only 22% of vOTUs exhibited significant 18O enrichment, indicating active replication and new virion production during the 1-week incubation; the majority (78%) persisted without detectable replication, consistent with a viral seed bank. Active vOTUs accounted for 4.76-5.15% of total virions per gram of soil, with viral loads ranging from 3.15 x 1010 to 6.59 x 1010 virions per gram. Probabilistic and deterministic sensitivity analyses spanning viral DNA fraction and genome length reinforced that persistent virions represented the majority of the extracellular viral pool post-wet-up, regardless of parameter assumptions. Host predictions linked both active and persistent vOTUs primarily to Actinomycetota and Pseudomonadota--bacterial groups known to rapidly resuscitate following rewetting--suggesting that some viruses exhibit rapid turnover while others persist over longer timescales, forming a stable viral pool capable of reinitiating infections during favorable conditions. These results demonstrate that SIP-viromics can distinguish newly produced from persistent virions and reveal host-associated patterns of lytic infection and virion production. Our findings advance understanding of soil virus-host interactions and highlight the ecological role of persistent virions as a genetic reservoir contributing to microbial turnover and biogeochemical cycling following environmental disturbance. ImportanceUnderstanding the persistence and production dynamics of soil viruses is critical for elucidating their roles in microbial community dynamics and nutrient cycling, yet these processes have remained largely uncharacterized due to methodological limitations. By integrating stable isotope probing with viromics, this study provides a robust framework for directly distinguishing newly produced from persistent virions in situ. Unlike conventional viromics, which only catalogs viral diversity, SIP-viromics enables quantification of active viral replication and persistence under natural soil conditions. Our results demonstrate that most virions in a seasonally dry soil persisted through a rewetting event, with active replication limited to a minority of viral populations. Persistent virions were primarily linked to dominant bacterial groups, indicating that host ecophysiology and environmental stability strongly influence lytic infection. Collectively, these findings highlight viruses as long-term reservoirs of genetic material, capable of shaping microbial dynamics and ecosystem processes over time. This work establishes SIP-viromics as a powerful approach for studying virus-host interactions and their ecological significance in terrestrial environments.

microbiology↗

bioRxiv2025

Translation Efficiency Impacts Phage Lysis Timing and Its Precision in Single Cells

The timing of host cell lysis is a fundamental life history parameter in bacteriophages as it presents an evolutionary trade-off between maximizing intracellular phage replication and optimizing transmission to new host cells, ultimately determining viral fitness in dynamic bacterial populations. In the bacteriophage {lambda}, lysis time is dependent on the expression of a timekeeper protein, holin, which accumulates in the Escherichia coli inner membrane. Cell lysis is triggered when the membrane concentration of holin crosses a critical threshold level. In the present study, we investigated the effects of the rate of holin translation on lysis timing and its precision. We show that modulating holin translation efficiency through genetic modifications and antibiotic treatment alters bacteriophage lysis timing and its precision, providing insight into how phages optimize the evolutionary trade-off between replication and transmission. Reducing ribosomal binding affinity decreased holin expression and delayed lysis, while optimizing the Shine-Dalgarno sequence enhanced translation and accelerated lysis. Surprisingly, very low tetracycline concentrations may have improved translation efficiency and hastened lysis, whereas higher doses predictably delayed it. In all cases, longer lysis times corresponded with decreased timing variability. A model that incorporates stochastic gene expression with additional stochasticity in the initiation of holin expression was sufficient to explain the data. Thus, we demonstrated that modifying the rate of holin translation can be used as an evolutionary strategy to achieve optimum lysis timing. ImportanceOur prior work demonstrated that phage lysis timing is calibrated to optimize fitness based on environmental conditions affecting host availability and phage replication. We identified an optimal threshold--the critical holin concentration--that minimizes lysis time variability. Genetic manipulation of the holin gene altered the lysis threshold and generated phages with non-optimal early and late lysis times. The current study reveals that adjusting holin translation rates offers another mechanism for modifying lysis timing. Phages therefore have multiple means to adjust the timing of host lysis in order to maximize fitness under different selective pressures.

microbiology↗

bioRxiv2025

Prognostically favorable immune responses to ovarian cancer are distinguished by self-reactive intra-epithelial plasma cells

Tumor-infiltrating B cells (TIL-Bs) are strongly associated with patient survival; however, the underlying mechanisms are poorly understood. Using integrated single-cell and spatial biology approaches, we defined at clonal resolution the molecular phenotypes, tumor reactivity patterns, and microenvironmental locations of TIL-Bs in high-grade serous ovarian cancer (HGSC). Prognostic benefit was associated with a TIL-B-rich tumor microenvironment with marked infiltration of malignant epithelium by plasma cells (PCs). PCs spanned five molecular phenotypes; exhibited high rates of somatic hypermutation and clonal expansion; and expressed predominantly IgG1 antibodies recognizing broadly expressed nuclear, cytoplasmic, and cell surface self-antigens. Many PC-derived antibodies were polyreactive. Self- and poly-reactive TIL-Bs penetrated tumor epithelium and stroma and expressed interferon-stimulated genes, indicating strong in situ activation. The self- and poly-reactive nature of TIL-B responses, reminiscent of autoimmune disease, may provide a means for the immune system to combat tumor heterogeneity and could potentially be harnessed for more effective immunotherapy.

cancer biology↗

bioRxiv2025

Widespread gray and white matter microstructural alterations in dual cognitive-motor impairment

INTRODUCTIONDual cognitive-motor impairment in aging is a strong predictor of dementia, yet its effects on vulnerable gray matter regions microstructure remain unexplored. METHODSThis study classified 582 individuals aged 36-90 into cognitive-motor impairment, isolated cognitive or motor impairment, and control groups. Microstructural differences in 27 temporal and motor-related gray matter regions and white matter tracts were assessed using DTI and mean apparent propagator (MAP-MRI), a technique well-suited for gray matter analysis. RESULTSWe found widespread microstructural alterations in gray and white matter among individuals with dual cognitive-motor impairment. These changes were not observed in isolated cognitive or motor impairment after multiple comparisons correction. DISCUSSIONDual cognitive-motor impairment is associated with reduced cellular density in temporal gray matter, decreased fiber coherence, and potential demyelination in white matter tracts, suggesting widespread microstructural disruption. These findings could help understand brain aging and facilitate interventions to slow neurodegeneration and delay dementia onset.

neuroscience↗

bioRxiv2025

Plasmodium Repetome: A mysterious space with a wealth of information

Eukaryotic proteomes harbour repetitive stretches of amino acids that may play critical roles in the biology of that organism. While several tandem repeats (TR) are known to contribute to protein structure and function, information about the vast majority of repeat regions remains obscure. In this article, we have analysed the repeat content of different Plasmodium species and found the leading human malaria-causing P. falciparum (Pf) and P. vivax to be exceptionally rich in TR regions (>40% TR containing proteins). Detailed analysis of Pf repetome showed this intracellular parasite to carry longer TRs, several of which were present in exported proteins important for parasite survival and immune evasion. The repeat regions of Pf were enriched in acidic amino acids and asparagine (Asn), where Asn was more abundant in short and intermediate TRs, suggesting an evolutionary bias influenced by replication slippage and positive selection. Gene ontology analysis of TR containing Pf proteins helped us to understand their cellular localization along with the molecular and biological processes they are involved in. The Pf variable surface antigen families with roles in important pathogenic processes like cytoadherence, immune evasion etc. had low repeat content present within seroreactive peptides. Three-dimensional structure predictions of TR regions revealed several repeats to adopt ordered super-secondary conformations that are known to facilitate intermolecular interactions. Overall, this is a comprehensive study attempting to gain insights on the importance of TRs in malaria parasite biology and suggests a novel route to understanding protein function through the characterization of repeat content.

bioinformatics↗

bioRxiv2025

A microbiome-derived transfer RNA modification underlies behavioral optimums and predator avoidance in mosquito larvae

The gut microbiome is a rich source of nutrients that are critical to the development and biology of eukaryotes. Transfer RNAs (tRNAs) are essential components of protein synthesis, and some chemical modifications to tRNA rely on the availability of microbiome-derived nutrients. In eukaryotes, the micronutrient queuosine (Q) is salvaged from the microbiome or diet and then incorporated into eukaryotic tRNA to influence the speed and efficiency of protein synthesis. Here, we examine the role of microbiome-derived Q in mosquito larval development and behavior. When mosquito larvae are grown with a microbiome incapable of synthesizing Q, there is a significant impact on tyrosine levels and processes, which correlate with defects in behavior and cuticle formation. Due to defects in movement and behavioral responses, Q-deficient larvae demonstrate impaired predator evasion, leading to higher instances of capture by predaceous beetle larvae. The broad effects of Q-deficiency in mosquito larvae highlight the importance of microbiome-derived nutrients for eukaryotic physiology and behavior.

physiology↗

bioRxiv2025

A Spatial Multi-Omic Framework Identifies Gliomas Permissive to TIL Expansion

Tumor-infiltrating lymphocyte (TIL) therapy is effective in several tumor types; however, its feasibility in immune subversive tumors like glioblastoma is unclear. We expanded TILs from glioblastoma specimens and observed marked variability in yield, composition, function, and TCR clonality. By interrogating TILs expanded ex vivo alongside their sourced glioma tissue, we sought to identify determinants of successful (TIL+) vs unsuccessful TIL expansion (TIL-). Expanded TILs were predominantly effector memory CD4 cells and exhibited oligoclonal TCR enrichment. Despite similar T cell abundance in TIL vs TIL- tumors, TIL tumors exhibited distinct spatial organization and cellular interactions, including increased endothelial-immune interactions/ proximity and enrichment of vascular-associated niches. CD4 T cells localized near CD68 macrophages in TIL tumors, while they were positioned near CD163 CD206 macrophages in TIL- tumors. Thus, TIL expansion and functionality are linked to spatial organization and myeloid context; these features may enable biomarker-driven stratification for future TIL therapy in gliomas.

cancer biology↗

bioRxiv2025

Rho2 regulates granulocyte-triggered stress adaptation and cell wall remodeling in Aspergillus fumigatus

The airborne opportunistic fungal pathogen Aspergillus fumigatus poses a deadly threat to immunocompromised patients. Neutrophil granulocytes play a key role in the defense against invasive infections caused by this pathogen. The mechanisms by which Aspergillus defends itself against attacks by the immune system are only partially understood. Here we show that human granulocytes activate the cell wall integrity (CWI) pathway of A. fumigatus and that key components of the CWI such as the cell wall stress sensor MidA and the Rho GTPases Rho2 and Rho4 are important for the survival of Aspergillus hyphae under granulocyte attacks. A more detailed investigation of the role of Rho2 revealed that a mutant lacking rho2 is less virulent in a Galleria mellonella infection model. Overexpression of Rho2 increases the resistance of A. fumigatus hyphae to killing by granulocytes. While a mutant lacking Rho2 has a normal cell wall composition, overexpression or constitutive activation of Rho2 leads to an altered cell wall composition and impairs growths of the pathogen. The fungicidal effect of constitutive activation of Rho2 signaling, which correlates with the formation of cell wall chitin bulges, depends on the CWI MAP kinase MpkA. However, Rho2 itself does not appear to be a direct activator of the CWI MAP kinase module. Our results support a model where Rho2 in A. fumigatus actively counteracts granulocyte attacks by upregulating cell wall biosynthesis, thereby strengthening the cell wall and aiding the fungus in surviving the stress condition.

microbiology↗