Biology subjects

Howell, S. A.

Publications and source records attributed to Howell, S. A..

5 recordsLinked to original sources

bioRxiv2025

Efficient delivery of mRNA-LNPs in primary and secondary liver cancer

Primary liver cancer is the sixth most prevalent cancer globally and is often diagnosed late, when treatment options are limited. Secondary liver cancer, arising from metastasis of other cancers to the liver, is a common complication of advanced solid cancers and a significant cause of cancer-related morbidity and mortality. Existing treatment options for advanced primary and secondary liver tumours have limited efficacy and new treatment modalities have the potential to improve patient outcomes. mRNA therapeutics are readily delivered to the healthy liver after systemic administration, but their uptake and expression within liver tumours is unclear. Here we show that intravenous delivery of mRNA-LNPs efficiently transfects virtually all hepatocytes in healthy, fibrotic, and cirrhotic liver, and also many cells of spontaneous hepatocellular carcinomas in situ. Delivery of mRNA is also possible to xenograft models of both primary and secondary liver cancer, albeit with attenuated protein expression relative to the normal liver. These findings demonstrate the potential for systemically delivered mRNA-LNP therapies for liver disease and cancer.

molecular biology↗

bioRxiv2023

A malaria parasite phospholipase facilitates efficient asexual blood stage egress

Malaria parasite release (egress) from host red blood cells involves parasite-mediated membrane poration and rupture, thought to involve membrane-lytic effector molecules such as perforin-like proteins and/or phospholipases. With the aim of identifying these effectors, we disrupted the expression of two Plasmodium falciparum perforin-like proteins simultaneously and showed that they have no essential roles during blood stage egress. Proteomic profiling of parasite proteins discharged into the parasitophorous vacuole (PV) just prior to egress detected the presence in the PV of a lecithin:cholesterol acyltransferase (LCAT; PF3D7_0629300). Conditional ablation of LCAT resulted in abnormal egress and a reduced replication rate. Lipidomic profiles showed drastic changes in several phosphatidylserine and acylphosphatidylglycerol species during egress. We thus show that, in addition to its previously demonstrated role in liver stage merozoite egress, LCAT is required to facilitate efficient egress in asexual blood stage malaria parasites. Author SummaryMalaria kills over half a million people every year worldwide. It is caused by a single-celled parasite called Plasmodium falciparum that grows and multiplies within a bounding vacuole, inside red blood cells of the infected individuals. Following each round of multiplication, the infected cell is ruptured in a process known as egress to release a new generation of parasites. Egress is required for the disease to progress and is orchestrated by the parasite. The parasite sends out various molecules to puncture and destroy the membranes of the vacuole and the red blood cell. However, little is known about these molecules. In this work, we set out to identify these molecules by using genetic and proteomics approaches. We screened the molecules the parasite sends out during egress and identified a parasite enzyme called LCAT present in the vacuole. Our experiments found that mutant parasites that were unable to make LCAT clumped together and could not escape the infected cell properly. As a result, we saw a reduction in the rate at which these parasites spread through the red blood cells. Taken together, our findings suggest that P. falciparum needs LCAT to efficiently break out of red blood cells.

microbiology↗

bioRxiv2021

The Aspartyl Protease Ddi1 Is Essential for Erythrocyte Invasion by the Malaria Parasite

Malaria pathology is caused by the exponential replication of Plasmodium parasites in the blood stream. The bottleneck of the parasite life cycle is the invasion of erythrocytes immediately after parasites egress from infected red blood cells. DNA damage-inducible protein 1 (Ddi1) is a conserved eukaryotic proteasome shuttle protein containing an internal retroviral-like protease domain. Using conditional genetics, we now show that the proteolytic activity of the P. falciparum homologue, PfDdi1, is critically required for invasion of red blood cells. Furthermore, PfDdi1 disruption results in the accumulation of highly polyubiquitinated proteins that can be processed by purified PfDdi1 or distant eukaryotic homologues. We also show that PfDdi1 interacts with multiple components of the ubiquitin-proteasome system and that parasites lacking PfDdi1 are more sensitive to proteasome inhibition. Overall, this study establishes PfDdi1 as a key component of the eukaryotic ubiquitin-proteasome system and as a promising antimalarial target.

cell biology↗

bioRxiv2020

A multipass membrane protein interacts with the cGMP-dependent protein kinase to regulate critical calcium signals in malaria parasites

In malaria parasites, all cGMP-dependent signalling is mediated through a single cGMP-dependent protein kinase (PKG), a major function of which is to control essential calcium signals. However, how PKG transmits these signals in the absence of known second messenger-dependent calcium channels or scaffolding proteins is unknown. Here we identify a polytopic membrane protein, ICM1, with homology to transporters and calcium channels that is tightly-associated with PKG in both Plasmodium falciparum asexual blood stages and P. berghei gametocytes. Phosphoproteomic analyses in both Plasmodium species reveal multiple ICM1 phosphorylation events dependent upon PKG activity. Stage-specific depletion of P. berghei ICM1 blocks gametogenesis due to the inability of mutant parasites to mobilise intracellular calcium upon PKG activation, whilst conditional loss of P. falciparum ICM1 results in reduced calcium mobilisation, defective egress and lack of invasion. Our findings provide new insights into atypical calcium homeostasis in malaria parasites essential for pathology and disease transmission.

microbiology↗

bioRxiv2020

The malaria parasite sheddase SUB2 governs host red blood cell membrane sealing at invasion

Red blood cell (RBC) invasion by malaria merozoites involves formation of a parasitophorous vacuole into which the parasite moves. The vacuole membrane seals and pinches off behind the parasite through an unknown mechanism, enclosing the parasite within the RBC. During invasion, several parasite surface proteins are shed by a membrane-bound protease called SUB2. Here we show that genetic depletion of SUB2 abolishes shedding of a range of parasite proteins, identifying previously unrecognized SUB2 substrates. Interaction of SUB2-null merozoites with RBCs leads to either abortive invasion with rapid RBC lysis, or successful entry but developmental arrest. Selective failure to shed the most abundant SUB2 substrate, MSP1, reduces intracellular replication, whilst conditional ablation of the substrate AMA1 produces host RBC lysis. We conclude that SUB2 activity is critical for host RBC membrane sealing following parasite internalisation and for correct functioning of merozoite surface proteins. Key highlightsO_LIMany malaria parasite surface proteins are shed by SUB2 during RBC invasion C_LIO_LISUB2-null merozoites either induce rapid host RBC lysis, or invade then die C_LIO_LIMerozoite surface protein shedding is crucial for host RBC membrane sealing C_LI

microbiology↗