Biology subjects

Cyrklaff, M.

Publications and source records attributed to Cyrklaff, M..

6 recordsLinked to original sources

bioRxiv2023

Microtubule inner proteins of Plasmodium are essential for transmission of malaria parasites

Microtubule inner proteins, MIPs, are microtubule associated proteins that bind to tubulin from the luminal side. MIPs can be found in axonemes to stabilize flagellar beat or within cytoplasmic microtubules. Plasmodium spp. are the causative agents of malaria that feature different forms across a complex life cycle with both unique and divergent microtubule-based arrays. Here we investigate the role of four MIPs in a rodent malaria parasite for their role in transmission to and from the mosquito. We show by single and double gene deletions that SPM1 and TrxL1, MIPs associated with the subpellicular microtubules are dispensable for transmission from the vertebrate host to the mosquito and back. In contrast, FAP20 and FAP52, MIPs associated with the axonemes of gametes, are essential for transmission to mosquitoes but only if both genes are deleted. In the absence of both, FAP20 and FAP52 the B-tubule of the axoneme partly detaches from the A-tubule resulting in the deficiency of axonemal beating and hence gamete formation and egress. Our data suggest that a high level of redundancy ensures microtubule stability in the transmissive stages of Plasmodium, which is important for parasite transmission.

microbiology↗

bioRxiv2021

Reversible brain edema in experimental cerebral malaria is associated with transcellular blood-brain barrier disruption and delayed microhemorrhages

Brain swelling occurs in cerebral malaria (CM) and may either reverse or result in fatal outcome. It is currently unknown how brain swelling in CM reverses, as investigations have been hampered by inadequate animal models. In this study, we show that reversible brain swelling in experimental murine cerebral malaria (ECM) can be induced reliably after single vaccination with radiation-attenuated sporozoites as revealed by in vivo high-field (9.4T) magnetic resonance imaging. Our results provide evidence that parenchymal fluid increase and consecutive brain swelling results from transcellular blood-brain barrier disruption (BBBD), as revealed by electron microscopy. This mechanism enables reversal of brain swelling but does not prevent persistent focal brain damage, evidenced by microhemorrhages, in areas of most severe BBBD. In a cohort of 27 pediatric and adult CM patients (n=4 fatal, n=23 non-fatal) two out of four fatal CM patients (50%) and 8 out of 23 non-fatal CM patients (35%) showed microhemorrhages on MRI at clinical field strength of 1.5T, emphasizing the translational potential of the experimental model. Our data suggest that targeting transcellular BBBD may represent a promising adjunct therapeutic approach in cerebral brain swelling to reduce edema and may ultimately lead to a reduced permanent brain damage and a better longtime neurological outcome. Author summaryBrain swelling, which occurs in diseases such as cerebral malaria, is not necessarily fatal, and may reverse. Even upon reversal of brain swelling, neurological sequelae can still occur. The factors contributing to the reversibility of brain edema are not known, and treatment options remain therefore limited. Identifying the mechanisms leading to such reversibility could inform clinical management aimed at decreasing brain swelling and consecutive brain injury. Here we introduce a reproducible and simple animal model that allows comprehensive in vivo studies of reversible brain swelling in cerebral malaria at the peak of disease and upon recovery. We identify a specific type of blood-brain barrier disruption (BBBD) as a mechanism that occurs in brain swelling. We show that BBBD can reverse, but also highlight remaining brain damage in areas of most severe BBBD. As the ECM model introduced here bares crucial similarities to the CM in humans, our findings open strategies to study new therapeutic avenues and point to compounds that specifically target transcellular BBBD to reduce brain edema, and increase survival rates.

neuroscience↗

bioRxiv2021

KAHRP dynamically relocalizes to remodeled actin junctions and associates with knob spirals in P. falciparum-infected erythrocytes

The knob-associated histidine-rich protein (KAHRP) plays a pivotal role in the pathophysiology of Plasmodium falciparum malaria by forming membrane protrusions in infected erythrocytes, which anchor parasite-encoded adhesins to the membrane skeleton. The resulting sequestration of parasitized erythrocytes in the microvasculature leads to severe disease. Despite KAHRP being an important virulence factor, its physical location within the membrane skeleton is still debated, as is its function in knob formation. Here, we show by super-resolution microscopy that KAHRP initially associates with various skeletal components, including ankyrin bridges, but eventually co-localizes with remnant actin junctions. We further present a 35[A] map of the spiral scaffold underlying knobs and show that a KAHRP-targeting nanoprobe binds close to the spiral scaffold. Single-molecule localization microscopy detected [~]60 KAHRP molecules per knob. We propose a dynamic model of KAHRP organization and a function of KAHRP in attaching other factors to the spiral scaffold.

microbiology↗

bioRxiv2021

Asynchronous nuclear cycles in multinucleated Plasmodium falciparum enable rapid proliferation

Malaria-causing parasites proliferate within erythrocytes through schizogony, forming multinucleated stages before cellularization. Nuclear multiplication does not follow a strict geometric 2n progression and each proliferative cycle produces a heterogeneous number of progeny. Here, by tracking nuclei and DNA replication, we show that individual nuclei replicate their DNA at different times, despite residing in a shared cytoplasm. Extrapolating from experimental data using mathematical modeling, we demonstrate that a limiting factor must exist that slows down the nuclear multiplication rate. Indeed, our data show that temporally overlapping DNA replication events were significantly slower than partially or non-overlapping events. Our findings suggest an evolutionary pressure that selects for asynchronous DNA replication, balancing available resources with rapid pathogen proliferation.

microbiology↗

bioRxiv2021

An extended DNA-free intranuclear compartment organizes centrosomal microtubules in Plasmodium falciparum

Rapid proliferation of Plasmodium falciparum parasites in human red blood cells is the cause of malaria and is underpinned by an unconventional cell division mode, called schizogony. Contrary to model organisms, P. falciparum replicates by multiple rounds of closed and asynchronous nuclear divisions that are not interrupted by cytokinesis. Organization and dynamics of the critical nuclear division factors are, however, poorly understood. Centriolar plaques, the centrosomes of P. falciparum, are important regulators of division and serve as microtubule organizing centers. Early microscopy studies reveal an acentriolar, amorphous structure although its detailed organization remains elusive. Intranuclear microtubules mediate chromosome segregation, but the small size of parasite nuclei has precluded detailed analysis of their arrangement by classical fluorescence microscopy. We apply recently developed STED, expansion microscopy, and live cell imaging protocols to describe the reconfiguration of microtubules during schizogony. Analysis of centrin, nuclear pore, and microtubule positioning reveals a bipartite organization of the centriolar plaque. While centrin is extranuclear, we confirm by correlative light and electron tomography that microtubules are nucleated in a previously unknown and extended intranuclear compartment, which is devoid of chromatin. This study enables us to build a working model of the organization of an unconventional centrosome and better understand the diversity of eukaryotic cell division modes.

cell biology↗

bioRxiv2021

Co-chaperone involvement in knob biogenesis implicates host-derived chaperones in malaria virulence

The pathology associated with malaria infection is largely due to the ability of infected human erythrocytes to adhere to a number of receptors on endothelial cells within tissues and organs. This phenomenon is driven by the export of parasite-encoded proteins to the host cell, the exact function of many of which is still unknown. Here we inactivate the function of one of these exported proteins, PFA66, a member of the J-domain protein family. Although parasites lacking this protein were still able to grow in cell culture, we observed severe defects in normal host cell modification, including aberrant morphology of surface knobs, disrupted presentation of the cytoadherence molecule PfEMP1, and a total lack of cytoadherence, despite the presence of the knob associated protein KAHRP. Complementation assays demonstrate that an intact J-domain is required for recovery to a wild-type phenotype and suggest that PFA66 functions in concert with a HSP70 to carry out host cell modification. Strikingly, this HSP70 is likely to be of host origin. Taken together, our data reveal a role for PFA66 in host cell modification, implicate human HSP70 as also being essential in this process, and uncover a KAHRP-independent mechanism for correct knob biogenesis. Our observations open up exciting new avenues for the development of new anti-malarials.

cell biology↗