Dataset Information

Differentially expressed genes in the head of the 2nd instar pre-molting larvae of the nm2 mutant of the silkworm, Bombyx mori.

ABSTRACT: Molting is an important physiological process in the larval stage of Bombyx mori and is controlled by various hormones and peptides. The silkworm mutant that exhibits the phenotype of non-molting in the 2nd instar (nm2) is incapable of molting in the 2nd instar and dies after seven or more days. The ecdysone titer in the nm2 mutant is lower than that in the wildtype, and the mutant can be rescued by feeding with 20E and cholesterol. The results of positional cloning indicated that structural alteration of BmCPG10 is responsible for the phenotype of the nm2 mutant. To explore the possible relationship between BmCPG10 and the ecdysone titer as well as the genes affected by BmCPG10, digital gene expression (DGE) profile analysis was conducted in the nm2 mutant, with the wildtype strain C603 serving as the control. The results revealed 1727 differentially expressed genes, among which 651 genes were upregulated and 1076 were downregulated in nm2. BLASTGO analysis showed that these differentially expressed genes were involved in various biological processes, cellular components and molecular functions. KEGG analysis indicated an enrichment of these differentially expressed genes in 240 pathways, including metabolic pathways, pancreatic secretion, protein digestion and absorption, fat digestion and absorption and glycerolipid metabolism. To verify the accuracy of the DGE results, quantitative reverse transcription PCR (qRT-PCR) was performed, focusing on key genes in several related pathways, and the results were highly consistent with the DGE results. Our findings indicated significant differences in cuticular protein genes, ecdysone biosynthesis genes and ecdysone-related nuclear receptors genes, but no significant difference in juvenile hormone and chitin biosynthesis genes was detected. Our research findings lay the foundation for further research on the formation mechanism of the nm2 mutant.

SUBMITTER: Wang P

PROVIDER: S-EPMC5519023 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Differentially expressed genes in the head of the 2nd instar pre-molting larvae of the nm2 mutant of the silkworm, Bombyx mori.

Wang Pingyang P Bi Simin S Wu Fan F Xu Pingzhen P Shen Xingjia X Zhao Qiaoling Q

PloS one 20170720 7

Molting is an important physiological process in the larval stage of Bombyx mori and is controlled by various hormones and peptides. The silkworm mutant that exhibits the phenotype of non-molting in the 2nd instar (nm2) is incapable of molting in the 2nd instar and dies after seven or more days. The ecdysone titer in the nm2 mutant is lower than that in the wildtype, and the mutant can be rescued by feeding with 20E and cholesterol. The results of positional cloning indicated that structural alt ...[more]

PMID: 28727825

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Project description:Molting is an important physiological process in the larval stage of Bombyx mori and is controlled by various hormones and peptides. The silkworm mutant that exhibits the phenotype of non-molting in the 2nd instar (nm2) is incapable of molting in the 2nd instar and dies after seven or more days. The ecdysone titer in the nm2 mutant is lower than that in the wildtype, and the mutant can be rescued by feeding with 20E and cholesterol. The results of positional cloning indicated that structural alteration of BmCPG10 is responsible for the phenotype of the nm2 mutant. To explore the possible relationship between BmCPG10 and the ecdysone titer as well as the genes affected by BmCPG10, digital gene expression (DGE) profile analysis was conducted in the nm2 mutant, with the wildtype strain C603 serving as the control. The results revealed 1727 differentially expressed genes, among which 651 genes were upregulated and 1076 were downregulated in nm2. BLASTGO analysis showed that these differentially expressed genes were involved in various biological processes, cellular components and molecular functions. KEGG analysis indicated an enrichment of these differentially expressed genes in 240 pathways, including metabolic pathways, pancreatic secretion, protein digestion and absorption, fat digestion and absorption and glycerolipid metabolism. To verify the accuracy of the DGE results, quantitative reverse transcription PCR (qRT-PCR) was performed, focusing on key genes in several related pathways, and the results were highly consistent with the DGE results. Our findings indicated significant differences in cuticular protein genes, ecdysone biosynthesis genes and ecdysone-related nuclear receptors genes, but no significant difference in juvenile hormone and chitin biosynthesis genes was detected. Considering the physiological characteristics of nm2 together with the results of DGE analysis, we speculate that there may be a relationship between BmCPG10 and ecdysone. In the mutant, the original function of BmCPG10 was lost due to the change in its protein structure. It appeared that BmCPG10 might influence the synthesis of ecdysone by controlling the production of substrate cholesterol. However, under a low titer of ecdysone, the expression of BmCPG10 was affected, as one of a series of effectors of the ecdysone signaling pathway. Blocking ecdysone biosynthesis resulted in non-molting silkworms and the formation of the nm2 mutant.

Project description:The ability to respond quickly and efficiently to transient extreme environmental conditions is an important property of all biota. However, the physiological basis of thermotolerance in different species is still unclear. Here, we found that the cot mutant showed a seizure phenotype including contraction of the body, rolling, vomiting gut juice and a momentary cessation of movement, and the heartbeat rhythm of the dorsal vessel significantly increases after hyperthermia. To comprehensively understand this process at the molecular level, the transcriptomic profile of cot mutant, which is a behavior mutant that exhibits a seizure phenotype, was investigated after hyperthermia (42°C) that was induced for 5 min. By digital gene expression profiling, we determined the gene expression profile of three strains (cot/cot ok/ok, +/+ ok/ok and +/+ +/+) under hyperthermia (42°C) and normal (25°C) conditions. A Venn diagram showed that the most common differentially expressed genes (DEGs, FDR<0.01 and log2 Ratio≥1) were up-regulated and annotated with the heat shock proteins (HSPs) in 3 strains after treatment with hyperthermia, suggesting that HSPs rapidly increased in response to high temperature; 110 unique DEGs, could be identified in the cot mutant after inducing hyperthermia when compared to the control strains. Of these 110 unique DEGs, 98.18% (108 genes) were up-regulated and 1.82% (two genes) were down-regulated in the cot mutant. KEGG pathways analysis of these unique DEGs suggested that the top three KEGG pathways were "Biotin metabolism," "Fatty acid biosynthesis" and "Purine metabolism," implying that diverse metabolic processes are active in cot mutant induced-hyperthermia. Unique DEGs of interest were mainly involved in the ubiquitin system, nicotinic acetylcholine receptor genes, cardiac excitation-contraction coupling or the Notch signaling pathway. Insights into hyperthermia-induced alterations in gene expression and related pathways could yield hints for understanding the relationship between behaviors and environmental stimuli (hyperthermia) in insects.

Project description:Amongst the various different insect groups, there is remarkable diversity in the number and size of wings. However the development of the basic body plan in insects is similar to a large extent. The genes of the hox complex regulate various pathways to bring about the development or modification of different organs. Ubx, a gene of the bithorax hox complex is expressed in the third thoracic segment of insects and is known to specify the fate of wing appendage in that segment.To understand the role of Ubx and how its regulatory mechanism has evolved through the course of evolution we have compared its genome wide targets in different insect orders. The identification of regulatory pathways and the key players Ubx regulates is crucial to understand how it has controlled wing development across insect orders. Our lab has previously identified direct targets of Ubx in Drosophila using ChIP-chip (Agrawal et al, 2011). To further our knowledge on the role of regulation in development and modification of hind wing appendage we have studied the targets in the hind wings of other insects (silk moth; Lepidoptera and honeybee; Hymenoptera) and performed a comparative analysis.To understand the differential development of wing appendages in insects we intend to compare the differential expression of fore and hind wing appendages in Diptera and Lepidoptera. We have employed RNA-seq using by illumina sequencing to identify the genes that are differentially expressed between fore and the hind wing bud of the Bombyx larvae. Only one replicate was performed as the intended study was to validate the differential targets in comparison to ChIP studies and other methods of select candidates. Total RNA was extarcted from wing buds of IV instar Bombyx larvae and sequenced on an illumina sequencer. Wing buds were collected directly in liquid nitrogen and 80 such buds each for fore and hind wing were collected from IV instar larvae. RNA was isolated from them and processed for Library preperation and illumina sequencing. One replicate was done as a supplement to ChIP studies.

Dataset Information

Differentially expressed genes in the head of the 2nd instar pre-molting larvae of the nm2 mutant of the silkworm, Bombyx mori.

Publications

Differentially expressed genes in the head of the 2nd instar pre-molting larvae of the nm2 mutant of the silkworm, Bombyx mori.

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