Identification and functional analysis of C-type lectin from mosquito Aedes albopictus in response to dengue virus infection.

BACKGROUND: C-type lectins (CTLs) are a large family of proteins with sugar-binding activity. CTLs contain an evolutionarily conserved C-type lectin domain (CTLD) that binds microbial carbohydrates in a calcium-dependent manner, thereby playing a key role in both microbial pathogenesis and innate immune responses. Aedes albopictus is an important vector for transmitting dengue virus (DENV) worldwide. Currently, the molecular characteristics and functions of CTLs in Ae. albopictus are largely unknown. METHODS: Transcripts encoding CTL proteins in the Ae. albopictus genome assembly were analyzed via sequence blast. Phylogenetic analysis and molecular characterization were performed to identify the functional domains of the CTLs. Quantitative analysis was performed to determine the gene expression features of CTLs during mosquito development and in different tissues of female adults after blood feeding. In addition, the functional role of CTLs in response to DENV infection was investigated in Ae. albopictus mosquito cells. RESULTS: We identified 39 transcripts encoding CTL proteins in the Ae. albopictus transcriptome. Aedes albopictus CTLs are classified into three groups based on the number of CTLDs and the domain architecture. These included 29 CTL-Ss (single-CTLDs), 1 immulectins (dual-CTLD) and 9 CTL-Xs (CTLDs with other domains). Phylogenetic analysis and structural modeling indicated that CTLs in Ae. albopictus are highly conserved with the homologous CTLs in Aedes aegypti. The expression profile assay revealed differential expression patterns of CTLs in both developmental stages and in adult female tissues. Knockdown and overexpression of three CTLs (CTL-S12, S17 and S19) confirmed that they can promote dengue virus infection in Ae. albopictus cells. CONCLUSIONS: The CTL genes in Ae. albopictus mosquito and other mosquito species are evolutionarily conserved and exhibit different developmental and tissue expression features. The functional assay indicated that three CTLs in Ae. albopictus mosquitoes are involved in promoting dengue virus infection. Our study revealed that CTLs play important roles in both the physiological processes and viral infection in mosquito vectors.

Hax1 regulate focal adhesion dynamics through IQGAP1.

Cell migration is a highly orchestrated process requiring the coordination between the cytoskeleton, cell membrane and extracellular matrix adhesions. Our previous study demonstrated that Hax1 interacts with EB2, a microtubule end-binding protein, and this interaction regulate cell migration in keratinocytes. However, little is known about the underlying regulatory mechanism. Here, we show that Hax1 links dynamic focal adhesions to regulate cell migration via interacting with IQGAP1, a multidomain scaffolding protein, which was identified by affinity purification coupled with LC-MS/MS. Biochemical characterizations revealed that C-terminal region of Hax1 and RGCT domain of IQGAP1 are the most critical binding determinants for its interaction. IQGAP1/Hax1 interaction is essential for cell migration in MCF7 cells. Knockdown of HAX1 not only stabilizes focal adhesions, but also impairs the accumulation of IQGAP in focal adhesions. Further study indicates that this interaction is critical for maintaining efficient focal adhesion turnover. Perturbation of the IQGAP1/Hax1 interaction in vivo using a membrane-permeable TAT-RGCT peptide results in impaired focal adhesion turnover, thus leading to inhibition of directional cell migration. Together, our findings unravel a novel interaction between IQGAP1 and Hax1, suggesting that IQGAP1 association with Hax1 plays a significant role in focal adhesion turnover and directional cell migration. Video Abstract.

Molecular and expression characterization of insulin-like signaling in development and metabolism of Aedes albopictus.

BACKGROUND: Insulin-like signaling (IS) in insects is a conserved pathway that regulates development, reproduction and longevity. Insulin-like peptides (ILPs) activate the IS pathway by binding to the insulin receptor (InR) and trigger the ERK and AKT cascades. A varying number of ILPs were identified in Aedes aegypti mosquito and other insects. Aedes albopictus is an invasive mosquito which transmits dengue and Zika viruses worldwide. Until now, the molecular and expression characteristics of IS pathway in Ae. albopictus have not been investigated. METHODS: The orthologues of ILP in Ae. albopictus genome assembly was analyzed by using sequence blast. Phylogenetic analysis and molecular characterization were performed to identify the functional domains of ILPs. Quantitative analysis was performed to determine the expression characteristics of ILPs, InR as well as ERK and AKT in mosquito development and different tissues of female adults after blood-feeding. In addition, the knockdown of InR was achieved by feeding larvae with Escherichia coli-producing dsRNA to investigate the impact of IS pathway on mosquito development. RESULTS: We identified seven putative ILP genes in Ae. albopictus genome assembly, based on nucleotide similarity to the ILPs of Ae. aegypti and other insects. Bioinformatics and molecular analyses suggested that the ILPs contain the structural motif which is conserved in the insulin superfamily. Expression levels of ILPs, InR as well as ERK and AKT varied in Ae. albopictus development stages and between male and female adults. Quantitative analyses revealed that expression of ILP6, the putative orthologue of the insulin growth factor peptides, was highest in the midgut of female adults after blood-feeding. Knockdown of Ae. albopictus InR induces a significant decrease in the phosphorylation levels of ERK and AKT proteins and results in developmental delays and smaller body sizes. CONCLUSIONS: The IS pathway of Ae. albopictus mosquito contains ILP1-7, InR and ERK/AKT cascades, which exhibited different developmental and tissue expression characteristics. Feeding Ae. albopictus larvae with E. coli-producing InR dsRNA blocks the ERK and AKT cascades and interferes with the development of mosquito. Our data suggest that IS pathway plays an important role in the metabolism and developmental process and could represent a potential target for controlling mosquito-borne diseases.

Frog Skin Derived Peptides With Potential Protective Effects on Ultraviolet B-Induced Cutaneous Photodamage.

Hyla annectans is a tree frog living in the southwestern plateau area of China where there is strong ultraviolet radiation and long duration of sunshine. So their naked skin may possess chemical defense components that protect it from acute photo-damage. However, no such peptide or components has been identified till to date. In the current work, two novel peptides (FW-1, FWPLI-NH2 and FW-2, FWPMI-NH2) were identified from the skin of the tree frog. Five copies of FW-1 and four copies of FW-2 are encoded by an identical gene and released from the same protein precursor, which possess 167 amino acid residues. FW-1 and -2 can exert significant anti-inflammatory functions by directly inhibiting Ultraviolet B irradiation (UVB)-induced secretion of inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). They may achieve this function by modulating the UV-induced stress signaling pathways such as Mitogen-activated protein kinases (MAPK) and Nuclear Factor Kappa B (NF-κB). Besides, FW-1 and -2 showed potential antioxidant effects on epidermis by attenuating the UVB-induced reactive oxygen species (ROS) production through an unknown mechanism. Considering small peptides' easy production, storage, and potential photo-protective activity, FW-1/2 might be exciting leading compounds or templates for the development of novel pharmacological agents for the suppression of UVB-induced skin inflammation. Moreover, this study might expand our knowledge on skin defensive mechanism of tree frog upon UVB irradiation.