Biology subjects

Arshad, H.

Publications and source records attributed to Arshad, H..

7 recordsLinked to original sources

bioRxiv2026

PRNP mutations initially generate an alternatively misfolded PrP species that dissuades prion replication

Mutations in the PRNP gene, which encodes the prion protein (PrP), cause genetic prion disease. However, how PRNP mutations lead to spontaneous prion formation remains poorly understood. Building off the observation that expression of mutant bank vole PrP (BVPrP) in mice causes spontaneous prion disease, we sought to identify early events in prion formation by expressing mutant BVPrP in cultured cells lacking endogenous PrP. Expression of D178N- and E200K-mutant, but not wild-type BVPrP, in CAD5-PrP-/- and N2a-PrP-/- cells generates an alternatively misfolded PrP species, which we term PrPAM, that is detergent-insoluble and resistant to digestion with the protease thermolysin. PrPAM is also present in the brains of young, asymptomatic knock-in mice expressing mutant BVPrP. However, PrPAM does not appear to be a direct precursor of prions since 1) the prion disease-protective G127V substitution enhances PrPAM levels when coupled with the D178N or E200K mutations; 2) small molecule anti-prion drugs fail to reduce PrPAM accumulation in cells; 3) unlike cells expressing wild-type BVPrP, cells expressing D178N- or E200K-mutant BVPrP are largely resistant to infection with several prion strains; and 4) PrPAM-containing cell homogenate does not contain seeds that induce misfolding of wild-type BVPrP in either cells or the RT-QuIC assay. These results raise the possibility that prion disease-causing PRNP mutations initially generate a protective misfolded PrP species that is refractory to prion formation, which provides a potential explanation for why genetic prion diseases manifest later in life despite the presence of a mutation from birth.

neuroscience↗

bioRxiv2026

A self-complementary recombinant adeno-associated virus vector coding for an anchorless prion protein carrying the G127V mutation extends survival in a rodent prion disease model

The replacement of a single codon in the human prion gene, causing the substitution of glycine with valine at position 127 (G127V) of the prion protein (PrP), prevents development of prion disease. We set out to explore if prion disease survival extension manifests in mice if the V127 mutant is delivered through a recombinant adeno-associated virus (rAAV) packaged as a self-complementary DNA. The notorious delivery limitations of rAAVs were overcome using a cross-correction approach that relied on the expression of the mutation in the context of glycosylphosphatidylinositoI-anchorless ({Delta}GPI) PrP. In this proof-of-concept study, we inoculated Rocky Mountain Laboratory (RML) prions into knock-in mice, in which the endogenous murine prion protein gene (Prnp) was replaced with the bank vole prion protein gene (BvPrnp). Prion-inoculated mice that were retro-orbitally transduced with a protective rAAV vector encoding BvPrnpV127{Delta}GPI survived [~]50 days longer than control mice that were unprotected. A deep proteomic analysis revealed that BvPrnpV127{Delta}GPI was protective by slowing perturbations to the proteome observed in late-stage RML prion disease. In addition to capturing details of synaptic decay and depletion of proteins in proximity to PrP, the proteomic dataset revealed the identity of proteins of potential diagnostic value that may be central to the brains attempt to fight prion disease by contributing to astrocytosis or microgliosis, by coping with calcium influx, or by enhancing the endoplasmic reticulum processing of essential proteins. Taken together, our results demonstrate that a gene therapy based on a GPI-anchorless PrP containing the G127V mutation can delay the onset of prion disease in mice, providing a framework for development of a corresponding therapy in humans. AUTHOR SUMMARYA rare change in the human prion protein, involving a single building block, has been linked to strong protection against prion diseases--fatal neurodegenerative disorders. This study tested whether that protective effect could be reproduced using gene therapy in mice. To this end, we exposed the animals to infectious prions and then delivered the protective version of the protein into mice using a viral carrier. Treated mice survived about seven weeks longer than untreated animals, showing that the approach can meaningfully slow disease progression. To understand why, we examined changes in brain proteins during disease and found that treatment helped preserve the normal protein levels of cellular proteins, particularly those involved in communication between nerve cells. The analysis also identified proteins altered in the disease that are linked to the brains defense responses, including inflammation, stress handling, and protein processing, some of which may serve as future disease markers. Importantly, the limited protection observed was not due to poor delivery of the therapy but likely reflects biological limits of the model used. Overall, the findings support the idea that gene therapies based on naturally protective human variants could help slow prion diseases and improve understanding of how the brain responds to them.

neuroscience↗

bioRxiv2025

The Protein Disulfide Isomerase P4HB/PDIA1 Modulates PrPC Levels and Prion Replication

Prions are misfolded, self-propagating versions of the cellular prion protein (PrPC) that cause invariably fatal transmissible neurodegenerative diseases in humans and animals. Little is known about how prions replicate in the brain, including whether other proteins participate in prion replication in vivo. Several members of the protein disulfide isomerase family have been shown to reside in close spatial proximity to PrPC in cells and mice, implying that they could be involved in prion biogenesis. Here, we show that stable knock-down of the protein disulfide isomerase P4HB (also called PDIA1) in prion-susceptible CAD5 cells reduces PrPC levels and results in lower levels of protease-resistant PrP (PrPres), a marker of infectious prions, following infection with two different prion strains. Partial reduction of P4HB activity using the P4HB-selective inhibitor KSC-34 also decreases PrPC levels in uninfected CAD5 cells whereas treatment of prion-infected CAD5 cells with KSC-34 results in higher levels of PrPres. A proportion of P4HB reaches the cell surface where PrPC is located, and a secreted P4HB variant increases PrPres levels in cells. Collectively, these results suggest that P4HB facilitates prion replication in cells by stabilizing PrPC and potentially acting as a chaperone that directly modulates prion conversion. Thus, targeting P4HB during prion disease may have therapeutic benefit.

neuroscience↗

bioRxiv2024

The Brain Interactome of a Permissive Prion Replication Substrate

Bank voles are susceptible to prion strains from many different species, yet the molecular mechanisms underlying the ability of bank vole prion protein (BVPrP) to function as a universal prion acceptor remain unclear. Potential differences in molecular environments and protein interaction networks on the cell surface of brain cells may contribute to BVPrPs unusual behavior. To test this hypothesis, we generated knock-in mice that express physiological levels of BVPrP (M109 isoform) and employed mass spectrometry to compare the interactomes of mouse (Mo) PrP and BVPrP following mild in vivo crosslinking of brain tissue. Substantial overlap was observed between the top interactors for BVPrP and MoPrP, with established PrP-interactors such as neural cell adhesion molecules, subunits of Na+/K+-ATPases, and contactin-1 being equally present in the two interactomes. We conclude that the molecular environments of BVPrP and MoPrP in the brains of mice are very similar. This suggests that the unorthodox properties of BVPrP are unlikely to be mediated by differential interactions with other proteins.

neuroscience↗

bioRxiv2024

The Molecular Determinants of a Universal Prion Acceptor

In prion diseases, the species barrier limits the transmission of prions from one species to another. However, cross-species prion transmission is remarkably efficient in bank voles, and this phenomenon can be recapitulated in mice by expression of the bank vole prion protein (BVPrP). The molecular determinants of BVPrPs ability to function as a universal or near-universal acceptor for prions remain incompletely defined. Building on our finding that cultured cells expressing BVPrP can replicate both mouse and hamster prion strains, we conducted a systematic analysis to identify key residues in BVPrP that permit cross-species prion replication. Consistent with previous findings, we demonstrate that residues N155 and N170 of BVPrP, which are absent in mouse PrP but present in hamster PrP, are critical for cross-species prion replication. Additionally, BVPrP residues V112, I139, and M205, which are absent in hamster PrP but present in mouse PrP, are also required to enable replication of both mouse and hamster prions. Unexpectedly, we found that residues E227 and S230 near the C-terminus of BVPrP severely restrict the accumulation of prions following cross-species prion challenge, suggesting that they may have evolved to counteract the inherent propensity of BVPrP to misfold. PrP variants with an enhanced ability to replicate both mouse and hamster prions displayed accelerated spontaneous aggregation kinetics in vitro. These findings suggest that BVPrPs unusual properties are governed by a key set of amino acids and that the enhanced misfolding propensity of BVPrP may enable cross-species prion replication.

neuroscience↗

bioRxiv2023

Convergent generation of atypical prions in knock-in mouse models of genetic prion disease

Most cases of human prion disease arise due to spontaneous misfolding of wild-type or mutant prion protein. Though recapitulating spontaneous prion conversion in animal models has proven challenging, transgenic mice expressing the misfolding-prone bank vole prion protein (BVPrP) recreate certain key aspects of sporadic and genetic prion disease. However, it remains unclear whether spontaneous prion generation can occur in the absence of protein over-expression and how disease-causing mutations affect prion strain properties. To address these issues, we generated knock-in mice expressing physiological levels of either wild-type or mutant BVPrP with isoleucine at codon 109. While mice expressing wild-type BVPrP remained free from neurological disease, a subset of knock-in mice expressing BVPrP with mutations that cause either fatal familial insomnia (D178N) or familial Creutzfeldt-Jakob disease (E200K) developed progressive neurological illness. Brains from spontaneously ill knock-in mice contained prion disease-specific neuropathological changes as well as atypical protease-resistant prion protein. Moreover, brain extracts from spontaneously ill D178N- or E200K-mutant BVPrP knock-in mice transmitted disease to mice expressing wild-type BVPrP. Surprisingly, the properties of the D178N- and E200K-mutant prions appeared identical both pre- and post-transmission, suggesting that both mutations guide the formation of a highly similar atypical prion strain. These findings imply that knock-in mice expressing mutant BVPrP spontaneously develop a bona fide prion disease and that mutations causing prion diseases may share a uniform initial mechanism of action. Therefore, these mice represent useful tools for studying the early stages of genetic prion diseases.

neuroscience↗

bioRxiv2020

Comprehensive studies of Head Maralla, Punjab, Pakistan vegetation for ethnopharmacological and ethnobotanical uses and their elaboration through quantitative indices

Head Maralla is a non-recognized wetland with diversified flora that becomes focus of current studies. Whole plant or their parts are being used for treating various maladies and they are the integral part of livelihood in the area. Unique species including Osmunda regalis is used for blood and renal diseases purifier. Wild plant resources are key to traditional ethnobotanical knowledge based practices and industrial applications. Current study reports Head Marala inhabitants interaction with these resources and identify priorities at species and habitat level for conservation. Four sites viz., River Tavi, Upstream Chenab, River Manawarwala Tavi and Bhalolpur were designated to record traditional knowledge through questionnaire and interviews during field trips. One hundred nineteen (119) plant species were identified belonging to 54 families, of which 87 species were of dicot, 12monocots, 05ferns, and 4 species of bryophytes. Fifty percent of the plant species were utilized as a whole for therapeutic purposes, followed by leaves which had more than 20% usage of total consumption. Ailments viz., urination (14%) followed by cough (8%), cold (7%), stomach (6%), asthma (6%), constipation (5%), laxative (5%), diarrhea (4%) etc., were associated with vegetation. Bronchial disorders, pneumonia, dyspepsia, anthelmintic and kidney stones, etc., were also among other diseases commonly cured by traditional knowledge. Fifteen percent of vegetation contributes as fodder species consumed by local community for livestock while almost 17% of local plants were utilized for industrial purposes like timber, fuel, furniture, wooden pots and sports goods. In conclusion the ecosystem of Head Maralla is a complex of aquatic, terrestrial and agricultural land that is located on climatic and geographical divides, which further add to botanical interest as included many wetland habitats with unique diversity of plants. It is suggested to devise comprehensive conservation strategies to safe indigenous knowledge in systematic way for comprehending ecological services.

plant biology↗