Intracellular development and impact of a eukaryotic parasite on its zombified microalgal host in the marine plankton

Parasites are widespread and diverse in the oceanic plankton, and many of them infect single-celled algae for survival. How these parasites develop and scavenge energy within the host and whether the cellular organization and metabolism of the host is altered remain open questions. Combining quantitative structural and chemical imaging with time-resolved transcriptomics, we unveil dramatic morphological and metabolic changes of the parasite Amoebophrya (Syndiniales) during intracellular infection (e.g. 200-fold increase of mitochondrion volume), particularly following digestion of nutrient-rich host chromosomes. Some of these changes are also found in the apicomplexan parasites (e.g. sequential acristate and cristate mitochondrion, switch from glycolysis to TCA), thus underlining key evolutionary-conserved mechanisms. In the algal host, energy-producing organelles (chloroplast) remain intact during most of the infection, but sugar reserves diminish while lipid droplets increase. Thus, rapid infection of the host nucleus could be a zombifying strategy to digest nutrient-rich chromosomes and escape cytoplasmic defense while benefiting from the maintained C-energy production of the host cell.

Fates of Sec, Tat, and YidC Translocases in Mitochondria and Other Eukaryotic Compartments

Formation of mitochondria by the conversion of a bacterial endosymbiont was a key moment in the evolution of eukaryotes. It was made possible by outsourcing the endosymbiont’s genetic control to the host nucleus, while developing the import machinery for proteins synthesized on cytosolic ribosomes. The original protein export machines of the nascent organelle remained to be repurposed or were completely abandoned. This review follows the evolutionary fates of three prokaryotic inner membrane translocases Sec, Tat, and YidC. Homologs of all three translocases can still be found in current mitochondria, but with different importance for mitochondrial function. Although the mitochondrial YidC homolog, Oxa1, became an omnipresent independent insertase, the other two remained only sporadically present in mitochondria. Only a single substrate is known for the mitochondrial Tat and no function has yet been assigned for the mitochondrial Sec. Finally, this review compares these ancestral mitochondrial proteins with their paralogs operating in the plastids and the endomembrane system.

The HSV1 Tail-Anchored Membrane Protein pUL34 Contains a Basic Motif That Supports Active Transport to the Inner Nuclear Membrane Prior to Formation of the Nuclear Egress Complex

Herpes simplex virus type 1 nucleocapsids are released from the host nucleus by a budding process through the nuclear envelope called nuclear egress. Two viral proteins, the integral membrane proteins pUL34 and pUL31, form the nuclear egress complex at the inner nuclear membrane, which is critical for this process. The nuclear import of both proteins ensues separately from each other: pUL31 is actively imported through the central pore channel, while pUL34 is transported along the peripheral pore membrane. With this study, we identified a functional bipartite NLS between residues 178 and 194 of pUL34. pUL34 lacking its NLS is mislocalized to the TGN but retargeted to the ER upon insertion of the authentic NLS or a mimic NLS, independent of the insertion site. If co-expressed with pUL31, either of the pUL34-NLS variants is efficiently, although not completely, targeted to the nuclear rim where co-localization with pUL31 and membrane budding seem to occur, comparable to the wild-type. The viral mutant HSV1(17+)Lox-UL34-NLS mt is modestly attenuated but viable and associated with localization of pUL34-NLS mt to both the nuclear periphery and cytoplasm. We propose that targeting of pUL34 to the INM is facilitated by, but not dependent on, the presence of an NLS, thereby supporting NEC formation and viral replication.

Participation of Multifunctional RNA in Replication, Recombination and Regulation of Endogenous Plant Pararetroviruses (EPRVs)

Pararetroviruses, taxon Caulimoviridae, are typical of retroelements with reverse transcriptase and share a common origin with retroviruses and LTR retrotransposons, presumably dating back 1.6 billion years and illustrating the transition from an RNA to a DNA world. After transcription of the viral genome in the host nucleus, viral DNA synthesis occurs in the cytoplasm on the generated terminally redundant RNA including inter- and intra-molecule recombination steps rather than relying on nuclear DNA replication. RNA recombination events between an ancestral genomic retroelement with exogenous RNA viruses were seminal in pararetrovirus evolution resulting in horizontal transmission and episomal replication. Instead of active integration, pararetroviruses use the host DNA repair machinery to prevail in genomes of angiosperms, gymnosperms and ferns. Pararetrovirus integration – leading to Endogenous ParaRetroViruses, EPRVs – by illegitimate recombination can happen if their sequences instead of homologous host genomic sequences on the sister chromatid (during mitosis) or homologous chromosome (during meiosis) are used as template. Multiple layers of RNA interference exist regulating episomal and chromosomal forms of the pararetrovirus. Pararetroviruses have evolved suppressors against this plant defense in the arms race during co-evolution which can result in deregulation of plant genes. Small RNAs serve as signaling molecules for Transcriptional and Post-Transcriptional Gene Silencing (TGS, PTGS) pathways. Different populations of small RNAs comprising 21–24 nt and 18–30 nt in length have been reported for Citrus, Fritillaria, Musa, Petunia, Solanum and Beta. Recombination and RNA interference are driving forces for evolution and regulation of EPRVs.

How SARS-CoV-2 and Other Viruses Build an Invasion Route to Hijack the Host Nucleocytoplasmic Trafficking System

The host nucleocytoplasmic trafficking system is often hijacked by viruses to accomplish their replication and to suppress the host immune response. Viruses encode many factors that interact with the host nuclear transport receptors (NTRs) and the nucleoporins of the nuclear pore complex (NPC) to access the host nucleus. In this review, we discuss the viral factors and the host factors involved in the nuclear import and export of viral components. As nucleocytoplasmic shuttling is vital for the replication of many viruses, we also review several drugs that target the host nuclear transport machinery and discuss their feasibility for use in antiviral treatment.

The Microsporidian Encephalitozoon Hellem Secretes A Host Nucleus-Targeting Protein (Ehhntp1) to Upregulate Endoplasmic Reticulum-Associated Degradation and Promote Protein Ubiquitination

Background: Microsporidia, a group of obligate intracellular parasites that can infect humans and nearly all animals, have lost the pathways for de novo amino acid, lipid and nucleotide synthesis and instead evolved strategies to manipulate host metabolism and immunity. The endoplasmic reticulum (ER) is a vital organelle for producing and processing proteins and lipids and is often hijacked by intracellular pathogens. However, little is known about how microsporidia modulate host ER pathways. Herein, we identified a secreted protein of Encephalitozoon hellem, EhHNTP1, and characterized its subcellular localization and functions in host cells.Methods: A polyclonal antibody against EhHNTP1 was produced to verify the protein subcellular localization in E. hellem-infected cells using indirect immunofluorescence assay (IFA) and Western blotting. HEK293 cells were transfected with wild-type or mutant EhHNTP1 fused with HA-EGFP, and the impacts on pathogen proliferation, protein subcellular localization and sequence functions were assessed. RNA sequencing of EhHNTP1-transfected cells was conducted to identify differentially expressed genes (DEGs) and pathway responses by bioinformatics analysis mainly with R packages. The DEGs in the transfected cells were experimentally confirmed with RT-qPCR and Western blotting. The regulatory effects of candidate DEGs were analyzed via RNA interference and cell transfection, and the effects were determined with RT-qPCR and Western blotting.Results: EhHNTP1 is secreted into the host nucleus, and its translocation depends on a nuclear localization signal sequence (NLS) at the C-terminus from amino acids 239 to 250. Transfection and overexpression of EhHNTP1 in HEK293 cells significantly promoted pathogen proliferation. RNA-seq of the transfected cells showed that genes involved in ER-associated degradation (ERAD), a quality control mechanism that allows for the targeted degradation of proteins in the ER, were prominently upregulated. Upregulation of the ERAD genes PDIA4, HERP, HSPA5 and Derlin3 determined by RNA-seq data was verified using RT-qPCR and Western blotting. Protein ubiquitination in the transfected cells was then assayed and found to be markedly increased, confirming the activation of ERAD. PDIA4 knockdown with RNAi significantly suppressed the expression of HERP, indicating that PDIA4 is a vital ERAD component exploited by EhHNTP1. Moreover, EhHNTP1ΔHRD, a deletion mutant lacking the histidine-rich domain (HRD) in the C-terminus, predominantly suppressed the upregulation of ERAD genes, indicating that the HRD is essential for EhHNTP1 functions.Conclusion: This study is the first report on a microsporidian secretory protein that targets the host nucleus to upregulate the ERAD pathway and subsequently promote protein ubiquitination. Our work provides new insights into microsporidia-host interactions.

Dinoflagellate Host Chloroplasts and Mitochondria Remain Functional During Amoebophrya Infection

Dinoflagellates are major components of phytoplankton that play critical roles in many microbial food webs, many of them being hosts of countless intracellular parasites. The phototrophic dinoflagellate Scrippsiella acuminata (Dinophyceae) can be infected by the microeukaryotic parasitoids Amoebophrya spp. (Syndiniales), some of which primarily target and digest the host nucleus. Early digestion of the nucleus at the beginning of the infection is expected to greatly impact the host metabolism, inducing the knockout of the organellar machineries that highly depend upon nuclear gene expression, such as the mitochondrial OXPHOS pathway and the plastid photosynthetic carbon fixation. However, previous studies have reported that chloroplasts remain functional in swimming host cells infected by Amoebophrya. We report here a multi-approach monitoring study of S. acuminata organelles over a complete infection cycle by nucleus-targeting Amoebophrya sp. strain A120. Our results show sustained and efficient photosystem II activity as a hallmark of functional chloroplast throughout the infection period despite the complete digestion of the host nucleus. We also report the importance played by light on parasite production, i.e., the amount of host biomass converted to parasite infective propagules. Using a differential gene expression analysis, we observed an apparent increase of all 3 mitochondrial and 9 out of the 11 plastidial genes involved in the electron transport chains (ETC) of the respiration pathways during the first stages of the infection. The longer resilience of organellar genes compared to those encoded by the nucleus suggests that both mitochondria and chloroplasts remain functional throughout most of the infection. This extended organelle functionality, along with higher parasite production under light conditions, suggests that host bioenergetic organelles likely benefit the parasite Amoebophrya sp. A120 and improve its fitness during the intracellular infective stage.

A conserved oomycete CRN effector targets tomato TCP14-2 to enhance virulence

Phytophthora spp. secrete vast arrays of effector molecules during infection to aid in host colonization. The CRN protein family forms an extensive repertoire of candidate effectors that accumulate in the host nucleus to perturb processes required for immunity. Here, we show that CRN12_997 from P. capsici binds a TCP transcription factor, SlTCP14-2, to inhibit its immunity-associated activity against Phytophthora. Co-immuno-precipitation and split-YFP studies confirm a specific CRN12_997-SlTCP14-2 interaction in vivo. Co-expression of CRN12_997 specifically counteracts the TCP14-enhanced immunity phenotype, suggesting CRN mediated perturbation of SlTCP14-2 function. We show that SlTCP14-2 associates with nuclear chromatin and that CRN12_997 diminishes SlTCP14-2 DNA-binding. Collectively, our data support a model in which SlTCP14-2 associates with chromatin to enhance immunity. The interaction between CRN12_997 and SlTCP14-2 reduces DNA-binding of the immune regulator. We propose that the modulation of SlTCP14-2 chromatin affinity, caused by CRN12-997, enhances susceptibility to Phytophthora capsici.

Agrobacterium-delivered VirE2 interacts with host nucleoporin CG1 to facilitate the nuclear import of VirE2-coated T complex

Agrobacterium tumefaciensis the causal agent of crown gall disease. The bacterium is capable of transferring a segment of single-stranded DNA (ssDNA) into recipient cells during the transformation process, and it has been widely used as a genetic modification tool for plants and nonplant organisms. Transferred DNA (T-DNA) has been proposed to be escorted by two virulence proteins, VirD2 and VirE2, as a nucleoprotein complex (T-complex) that targets the host nucleus. However, it is not clear how such a proposed large DNA–protein complex is delivered through the host nuclear pore in a natural setting. Here, we studied the natural nuclear import of theAgrobacterium-delivered ssDNA-binding protein VirE2 inside plant cells by using a split-GFP approach with a newly constructed T-DNA–free strain. Our results demonstrate that VirE2 is targeted into the host nucleus in a VirD2- and T-DNA–dependent manner. In contrast with VirD2 that binds to plant importin α for nuclear import, VirE2 directly interacts with the host nuclear pore complex component nucleoporin CG1 to facilitate its nuclear uptake and the transformation process. Our data suggest a cooperative nuclear import model in which T-DNA is guided to the host nuclear pore by VirD2 and passes through the pore with the assistance of interactions between VirE2 and host nucleoporin CG1. We hypothesize that this large linear nucleoprotein complex (T-complex) is targeted to the nucleus by a “head” guide from the VirD2–importin interaction and into the nucleus by a lateral assistance from the VirE2–nucleoporin interaction.

Cyberloma acerinae. [Descriptions of Fungi and Bacteria].

A description is provided for Cyberloma acerinae, which infects fish of the families Atherinidae, Gobiidae and Percidae in Europe; infections developing in the host cytoplasm (but not in nerve tissues), without diplokarya, and resulting in tumours (also known as 'xenomas') in which the host nucleus is centrally located. Some information on its dispersal and transmission, habitats and conservation status is given, along with details of its geographical distribution (Asia (Kazakhstan (Aktobe), Russia (Novosibirsk Oblast)) and Europe (Czech Republic, Finland, France, former Soviet Union, Ukraine)). No evaluation has been made of any possible positive economic impact of this species (e.g. as a source of useful products, as a provider of checks and balances within its ecosystem, etc.), although Kvach et al. (2014) observed that it parasitizes some invasive fish, and so may have potential as part of an integrated biological control programme.