Study On Differentially Expressed Genes Associated With Response To UVA Radiation In Drosophila Melanogaster (Diptera:Drosophilidae)

Drosophila melanogaster Meigen (Diptera:Drosophilidae) is one kind of diurnal insects and display a conspicuous positive photo tactic behavior to light stimuli. It shows spectral sensitivities to light in the UVA spectrum (400-315 nm). In the present study, the transcriptional response to UVA radiation in the model insect D. melanogaster which were irradiated at different UVA-exposure intensities (350 μW/cm2 and 1,000 μW/cm2) and times (0 h,0.5 h, and 1.0 h) was investigated, as well as systematically analyzed the expression patterns and biological functions of UVA-induced genes. Main results were summarized as follows:1. Contruction of subtracted cDNA libraries using SSH in D. melanogasterTo better understand the molecular mechanisms of insect response to UVA radiation, suppression subtractive hybridization (SSH) and real-time quantitative PCR (qRT-PCR) approaches were combined to reveal differential transcript expression in D. melanogaster. In this study, two subtractive cDNA libraries were constructed and sequenced, obtaining 131 high-quality unique expressed sequence tags (ESTs) that were up or down-regulated in D. melanogaster exposed to UVA radiation at 350 μW/cm2 for 0.5 h. Of the 131 ESTs, 102 unique ESTs were differentially expressed and classified into 10 functional categories. The results showed that UVA radiation induces expression of genes related to stress and defense response and metabolism. Potential transcription factor binding motifs upstream of these genes were associated with multiple signaling pathways that may help the insect cope with the stress of UVA radiation. Our results reveal that UVA radiation can influence the expression profiles of stress-responsive genes which involved in many aspects of functions and pathways and provide further insights into the mechanisms of adaptive response to UVA radiation stress.2. RNA-Seq libriaries in the Heads of D.melanogaster under UVA radiationWe used the model insect D. melanogaster to test its transcriptional response to UVA radiation (at 1,000 μW/cm2 for 0 h,0.5 h, and 1.0 h) via high-throughput sequencing and systematically analyzed the head gene-expression patterns between differential time points. A great number of expressed genes were detected and KEGG/pathways analysis predicted and confirmed several significant UVA-associated signaling pathways, such as the MAPK, ErbB, chemokine, Calcium, and Wnt signaling pathways. Furthermore, we analyzed the temporal expression patterns of UVA-responsive genes at three time-points using Short Time-series Expression Miner analysis. Of the 13,362 UVA-responsive genes, 5506 were clustered into 15 model profiles with 6 significant model profiles (profile 0,2, 3,4,7, and 10). Then, based on their expression pattern,6 significant model profiles were classified into three clusters. Subsequently, Gene Ontology (GO) assignment programs were employed to combine with Short Time-series Expression Miner (STEM) analysis to research on biological functions of DE genes in our experimental conditions,3 clusters enriched significantly 36,33, and 31 terms compared to the whole genome background, respectively. Among the biological processes, GO terms involved in the stress response were overrepresented and included "response to stimulus (GO:0050896)" and "immune system process (GO:0002376)". Prominently, "metabolic process (GO:0008152)" and "cellular process (GO:0009987)" were more highly proportion in each cluster. Our findings suggest that D. melanogaster can perceive UVA-stress signals and respond with various strategy mechanisms to counteract UVA-induced stress and adapt to their environmental conditions. Our systems analysis in this study will be beneficial for a better understanding of the mechanisms of the insect responses to UVA radiation.3. Effects of UVA radiation on the expression levels of CG7708 and ACh content in D. melanogasterWe analyzed the effects of UVA radiation on the transcripts of CG7708 and ACh content in D. melanogaster and the difference of the transcripts of CG7708 and ACh content between adults (which are phototacic) and larves (which are lucifugous). Compared with the corresponding controls, the expression levels of CG7708 and ACh content were also significantly influenced in D. melanogaster which were irradiated at different UVA-exposure intensities (P<0.01) and treatment patterns (P<0.001) in our experimental conditions. Besides, the effects of UVA radiation on the expression levels of CG7708 and ACh content were significantly different (P<0.001) between adults and larves. Forthermore, the expression levels of CG7708 and ACh content in D. melanogaster larva induced by the UVA intensity of 1,000 μW/cm2, were significantly negatively correlated (P<0.05) and the regression equation was Y=-0.0023X+3.3764. This result suggests that CG7708 may be involved in the process of ACh synthesis under the certain condition. In short, our results found that CG7708 and ACh play a role in regulating neural signal transduction induced by UVA radiation in D. melanogaster.