Dongliang Yu

Transcriptome comparison between honey bee queen- and worker-destined larvae

Caste differentiation in the female honey bee is one of the most intriguing polyphenism phenomena. This developmental switch depends on the differential expression of entire suites of the genes involved in the larval fate between the queens and workers. In this study, we compared the transcriptome differences between full-sister queen- (QL) and worker-destined larvae (WL) using high-throughput RNA-Seq. QL and WL at fourth (L4) and fifth instar (L5) were used to prepare four libraries and to generate 50,191,699 (QL4), 57,628,541 (WL4), 56,613,619 (QL5), and 58,626,829 (WL5) usable reads, which were assembled into groups of 7,952, 7,993, 7,971, and 8,023 genes, respectively. The transcriptome changes were investigated using the DEGs Package (DEGseq), which resulted in more than 4,500 differentially expressed genes (DEGs) between the castes. Eight of the DEGs were verified by quantitative real-time RT-PCR (qRT-PCR), and the results supported our sequencing data. All of the DEGs were analysed using Web Gene Ontology Annotation Plot (WEGO) and then mapped using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. These results suggest that over 70% of the DEGs in each instar were more highly expressed in QL than in WL, possibly suggesting that the QL genes had higher transcriptional activity than the WL genes during differentiation. The same gene set is active (but differentially expressed) in both castes, which in turn result in dimorphic females. The L4 stage is a very active gene expression period for both QL and WL before their pupal stage. The activity of the mTOR (a target of rapamycin) encoding gene in the mTOR signalling pathway is higher in QL4 than in WL4, and this difference was no longer present by the L5 feeding stage. The genes down-stream of mTOR maintained this change at the L5 stage. These results could contribute to an in-depth study of the candidate genes during honey bee caste differentiation and improve our current understanding of the polyphenism phenomenon in insects.

PdbrlA, PdabaA and PdwetA control distinct stages of conidiogenesis in Penicillium digitatum

Penicillium digitatum is one of the most important citrus postharvest pathogens worldwide. Reproduction of massive asexual spores is the primary factor contributing to the epidemic of citrus green mold. To understand the molecular mechanisms underlying conidiogenesis in P. digitatum, we functionally characterized the Aspergillus nidulans orthologs of brlA, abaA and wetA. We showed that deletion of PdbrlA completely blocked formation of conidiophores, whereas deletion of PdabaA led to the formation of aberrant and non-functional phialides. The PdwetA mutant showed various defective phenotypes, such as abnormal conidia with loose cell walls, delayed germination and reduced tolerance to osmotic, detergent, heat shock and menadione stresses, but elevated resistance to H2O2. PdbrlA-influenced genes were identified by comparing global gene expression profiles between the wild-type and the PdbrlA deletion mutant during conidiation. Gene ontology analysis of these differentially expressed genes (DEGs) revealed the diverse roles of PdbrlA in metabolism, transportation and cell structure. Moreover, out of 39 genes previously reported to be involved in conidiogenesis in Aspergillus, mRNA levels of 14 genes were changed in ΔPdbrlA. Our results confirm the roles of brlA, abaA and wetA in P. digitatum conidiogenesis and provide new insights into the genetics of conidiation in filamentous fungi.

Complete mitochondrial genome sequence of the phytopathogenic fungus Penicillium digitatum and comparative analysis of closely related species

The complete mitochondrial genome of Penicillium digitatum (Pers.:Fr) Sacc is reported, the first time in a phytopathogenic Penicillium species. Comparative analysis revealed its close relationship to mitochondrial genomes of other Penicillium and Aspergillus species, both in gene content and in arrangement. The intron content of protein coding genes revealed several differences. The different exon-intron organization of Cytochrome Oxidase Subunit 1 genes indicated their common origin before the divergence of Penicillium and Aspergillus, and that, largely, their introns were transmitted vertically.

A Genomics Based Discovery of Secondary Metabolite Biosynthetic Gene Clusters in Aspergillus ustus

Secondary metabolites (SMs) produced by Aspergillus have been extensively studied for their crucial roles in human health, medicine and industrial production. However, the resulting information is almost exclusively derived from a few model organisms, including A. nidulans and A. fumigatus, but little is known about rare pathogens. In this study, we performed a genomics based discovery of SM biosynthetic gene clusters in Aspergillus ustus, a rare human pathogen. A total of 52 gene clusters were identified in the draft genome of A. ustus 3.3904, such as the sterigmatocystin biosynthesis pathway that was commonly found in Aspergillus species. In addition, several SM biosynthetic gene clusters were firstly identified in Aspergillus that were possibly acquired by horizontal gene transfer, including the vrt cluster that is responsible for viridicatumtoxin production. Comparative genomics revealed that A. ustus shared the largest number of SM biosynthetic gene clusters with A. nidulans, but much fewer with other Aspergilli like A. niger and A. oryzae. These findings would help to understand the diversity and evolution of SM biosynthesis pathways in genus Aspergillus, and we hope they will also promote the development of fungal identification methodology in clinic.

Genomic and transcriptomic analyses of the tangerine pathotype of Alternaria alternata in response to oxidative stress.

The tangerine pathotype of Alternaria alternata produces the A. citri toxin (ACT) and is the causal agent of citrus brown spot that results in significant yield losses worldwide. Both the production of ACT and the ability to detoxify reactive oxygen species (ROS) are required for A. alternata pathogenicity in citrus. In this study, we report the 34.41 Mb genome sequence of strain Z7 of the tangerine pathotype of A. alternata. The host selective ACT gene cluster in strain Z7 was identified, which included 25 genes with 19 of them not reported previously. Of these, 10 genes were present only in the tangerine pathotype, representing the most likely candidate genes for this pathotype specialization. A transcriptome analysis of the global effects of H2O2 on gene expression revealed 1108 up-regulated and 498 down-regulated genes. Expressions of those genes encoding catalase, peroxiredoxin, thioredoxin and glutathione were highly induced. Genes encoding several protein families including kinases, transcription factors, transporters, cytochrome P450, ubiquitin and heat shock proteins were found associated with adaptation to oxidative stress. Our data not only revealed the molecular basis of ACT biosynthesis but also provided new insights into the potential pathways that the phytopathogen A. alternata copes with oxidative stress.

Acquisition of hydrogenosomal presequences: examples from Trichomonas vaginalis

Presequences play an important role in protein import into mitochondria-like organelles. Acquisition pathways have been revealed for some mitochondrial presequences, but little is known about hydrogenosomal presequences. Here we investigated the hydrogenosomal proteins of Trichomonas vaginalis and suggest that several hydrogenosomal presequences probably evolved from pre-existing sequences that were thereafter modified.

Disruption of LRRK2 in Zebrafish leads to hyperactivity and weakened antibacterial response

As a protein with complex domain structure and roles in kinase, GTPase and scaffolding, LRRK2 is believed to be an important orchestration node leading to several cascades of signal transduction rather than one specific pathway. LRRK2 variants were found to be associated with Parkinsons disease, Crohns disease and leprosy. Here we disrupt LRRK2 in zebrafish and found hyperactivity rather than hypoactivity in adult zebrafish mutants. By RNA-seq we found genes involved in infectious disease and immunological disease were notably affected. Functional studies also revealed a weakened antibacterial response in LRRK2 mutant. This mutant can be further explored for revealing molecular mechanisms and modeling of LRRK2 related diseases.

Genomic and transcriptome analysis of triclosan response of a multidrug-resistant Acinetobacter baumannii strain, MDR-ZJ06

During the last decade, an increasing amount of attention has focused on the potential threat of triclosan to both the human body and environmental ecology. However, the role of triclosan in the development of drug resistance and cross resistance is still in dispute ascribed to largely unknown of triclosan resistance mechanism. In this work, Acinetobacter baumannii MDR-ZJ06, a multidrug-resistant strain, was induced by triclosan, and the genomic variation and transcriptional levels were investigated, respectively. The comparative transcriptomic analysis found that several general protective mechanisms were enhanced under the triclosan condition, including responses to reactive oxygen species and cell membrane damage. Meanwhile, all of the detected fifteen single nucleotide polymorphisms were not directly associated triclosan tolerance. In summary, this work revealed the crucial role of the general stress response in A. baumannii under a triclosan stress condition, which informs a more comprehensive understanding of the role of triclosan in the spread of drug-resistant bacteria.

The complete mitochondrial genome of a rare human pathogen, Aspergillus ustus

Abstract Aspergillus ustus is among the most ubiquitous soil species and has been implicated in human infections. The complete mitochondrial genome of A. ustus has been determined by high-throughput sequencing technology in this work. Our study revealed that the mitochondrial genome of A. ustus is 33,007 bp long, with AT content of 74.84%, which consists of a usual set of mitochondrial proteins and RNA genes, including large and small ribosomal RNAs, 15 proteins and 20 tRNA genes and contains two introns. Notably, it also contains three hypothetical proteins without obvious homology to any known proteins. All structural genes are located on one strand and are apparently transcribed in one direction. The complete mitochondrial genomes of A. ustus would be useful for future investigation of the genetics, evolution and clinical identification of Aspergillus species.

Pou4f2‐GFP knock‐in mouse line: A model for studying retinal ganglion cell development

Pou4f2 acts as a key node in the comprehensive and step‐wise gene regulatory network (GRN) and regulates the development of retinal ganglion cells (RGCs). Accordingly, deletion of Pou4f2 results in RGC axon defects and apoptosis. To investigate the GRN involved in RGC regeneration, we generated a mouse line with a POU4F2‐green fluorescent protein (GFP) fusion protein expressed in RGCs. Co‐localization of POU4F2 and GFP in the retina and brain of Pou4f2‐GFP/+ heterozygote mice was confirmed using immunofluorescence analysis. Compared with those in wild‐type mice, the expression patterns of POU4F2 and POU4F1 and the co‐expression patterns of ISL1 and POU4F2 were unaffected in Pou4f2‐GFP/GFP homozygote mice. Moreover, the quantification of RGCs showed no significant difference between Pou4f2‐GFP/GFP homozygote and wild‐type mice. These results demonstrated that the development of RGCs in Pou4f2‐GFP/GFP homozygote mice was the same as in wild‐type mice. Thus, the present Pou4f2‐GFP knock‐in mouse line is a useful tool for further studies on the differentiation and regeneration of RGCs.