Biology subjects

Alpaugh, M.

Publications and source records attributed to Alpaugh, M..

5 recordsLinked to original sources

bioRxiv2026

A structure-guided pipeline yields peptide inhibitors that disarm fungal peptidase-driven virulence and resistance

Fungal infections are a major global health challenge, with current antifungal therapies limited by toxicity, cost, and resistance. For Cryptococcus neoformans, key virulence factors that initiate and sustain infection are regulated by fungal peptidases to produce a polysaccharide capsule, promote immune evasion, and support antifungal resistance. These peptidases represent promising targets for antivirulent therapeutic strategies. Here, we developed a computational pipeline to predict and design peptide- and protein-based inhibitors against cryptococcal peptidases. Specifically, we targeted three virulence-associated peptidases: Rim13 (cysteine), May1 (aspartic), and CnMpr1 (metallo). Cysteine peptidase inhibition decreased capsule/cell size ratios without impeding fungal growth and reduced fungal survival within macrophages. Similarly, aspartic peptidase inhibition enhanced fungal clearance within alveolar macrophages and disrupted biofilm formation with additive effects towards fluconazole susceptibility in resistant strains. Additionally, metallopeptidase inhibition through catalytic zinc chelation and blocked substrate binding led to enhanced enzymatic inhibition and reduced in vitro blood-brain barrier crossing. Moreover, an in vivo larval model assessing inhibitor efficacy produced additive effects with fluconazole and lacked host cell cytotoxicity and fungicidal properties, reinforcing anti-virulence mechanisms and therapeutic potential while limiting the evolution of resistance. Further, global proteome profiling of inhibitor treated cells defined a mechanism of cell wall disruption, impeding fungal virulence. Taken together, the designed peptidase inhibitors exhibited potent antifungal activity without harming mammalian cells, establishing a predictive framework for rational scaffold design of next-generation antifungals that disarm the pathogen enabling immune-mediated clearance.

microbiology↗

bioRxiv2025

Direct Measure of DNA Damage in Huntington Disease Reveals Elevated Oxidative Genotoxic Stress and Dysfunctional DNA Repair

DNA damage repair (DDR) pathway proteins are genetic modifiers of Huntington disease (HD) age of onset and severity. Deficient DDR in HD at the first steps of poly ADP-ribosylation makes DNA damage quantification by traditional downstream DNA damage response markers inaccurate. Repair Assisted Damage Detection (RADD) allows for an accurate direct assessment of oxidative DNA damage, a DDR pathway in which huntingtin protein directly participates. Using RADD, we show that human HD-derived cells have elevated oxidative DNA damage and assess the effect of relevant HD therapeutic targets in rescuing this phenotype. Using huntingtin protein level lowering, we define a dysfunctional role of mutant huntingtin in oxidative DDR. We show that ataxia-telangiectasia mutated (ATM) signaling is deficient and that ATM inhibition rescues elevated oxidative DNA damage in HD cells. In contrast, we show that N6-furfuryladenine (N6FFA) treatment, to increase huntingtin phosphorylation within the amino terminal N17 domain (p-N17), is not effective at restoring HD DDR but reveals dysfunctional N6FFA mediated DDR signaling. We propose a model in which elevated DNA damage arises from both aberrant mutant huntingtin involvement in oxidative DDR and the impairment of oxidative DDR pathways, such as ATM kinase activity, N6FFA processing, and poly ADP-ribose signaling in HD. This highlights the importance of using direct measures of DNA damage such as RADD, rather than measures of a DNA damage response pathway.

cell biology↗

bioRxiv2025

Vascular dysfunction in Huntington's disease is located at the blood-CSF barrier and is rescued by sphingosine-1-phosphate receptor agonist

Brain vascular barriers are dysfunctional in many neurological disorders, including Huntingtons disease (HD), but which vessels are affected and how remains unclear. Using in vivo two-photon microscopy in R6/2 HD-model mice, we reveal a striking divergence in barrier dysfunction. Surface vessels of the blood-cerebrospinal fluid barrier (BCSFB) drive pathology by paracellular leakage, while the blood-brain barrier (BBB) of parenchymal arterioles and capillaries exhibited increased vesicular transport and adsorptive-mediated transcytosis (AMT), with arterioles being the most susceptible segment. The pathology of both barriers was congruent with astrocyte activation localized to affected vessels in R6/2 mice, as well as in post-mortem HD human brains. Sphingosine-1-phosphate receptor 1 (S1PR1) agonist treatment rescued BCSFB leakage but only partially restored BBB function, selectively reducing AMT in arterioles, with no effect in capillaries and venules. These findings redefine HD vascular pathology, demonstrating heterogeneous, vessel-type-specific barrier failure beyond generalized "disruption," uncovering mechanistic vulnerabilities and the therapeutic potential of S1PR1 modulation.

neuroscience↗

bioRxiv2024

Cas9 nickase-mediated contraction of CAG/CTG repeatsat multiple disease loci

Expanded CAG/CTG repeats cause over 15 different diseases that all remain without a disease-modifying treatment. Because repeat length accounts for most of the variation in disease severity, contracting them presents an attractive therapeutic avenue. Here, we show that the CRISPR-Cas9 nickase targeted to CAG/CTG repeats leads to efficient contractions in Huntingtons disease patient-derived neurons and astrocytes, and in myotonic dystrophy type 1 patient-derived neurons. The approach is allele-selective and free of detectable off-target mutations. Striatal injection of the Cas9 nickase in a mouse model for Huntingtons disease using adeno-associated viral vectors led to contractions in over half the infected cells. Upon injection, we observed a reduction in the number of inclusion bodies, improved transcriptome, and ameliorated locomotion. The effects were greater than expected from the contractions induced and suggest that non-cell autonomous mechanisms may be involved. Our results provide the proof-of-concept that correction of CAG/CTG repeats can improve Huntingtons disease phenotypes in vivo. One sentence summaryThe Cas9 nickase contracts CAG/CTG repeats at multiple disease loci in patient-derived cells and improves molecular and behavioral phenotypes in HD.

neuroscience↗

bioRxiv2019

Male sex exacerbates the plasticity and pathogenicity of Th17 cells in a chronic model of CNS autoimmunity.

Sex differences in the incidence and severity of multiple sclerosis (MS) have long been recognized. However, the underlying cellular and molecular mechanisms for why male sex is associated with more aggressive and debilitating disease remain poorly defined. Using an T cell adoptive transfer model of chronic EAE, we find that male Th17 cells induced disease of increased severity relative to female Th17 cells, irrespective of whether transferred to male or female recipients. Throughout the disease course, a greater frequency of male Th17 cells produced the heterodox cytokine IFN{gamma}, a hallmark of pathogenic Th17 responses. Intriguingly, sex chromosomal complement, and not hormones, were responsible for the increased pathogenicity of male Th17 cells and an X-linked immune regulator, Jarid1c, was downregulated in both pathogenic male Th17 and CD4+ T cells from men with MS. Together, our data indicate that male sex critical regulates Th17 cell plasticity and pathogenicity via sex chromosomal complement.

immunology↗