Qin Yao

The basic helix-loop-helix transcription factor family in Bombyx mori

The basic helix-loop-helix (bHLH) proteins are a superfamily of transcription factors that play important roles in a wide range of developmental processes in higher organisms. bHLH family members have been identified in a dozen of organisms including fruit fly, mouse and human. We identified 52 bHLH members in silico in the silkworm genome. Phylogenetic analyses revealed that they belong to 39 bHLH families with 21, 10, 12, 1, 7 and 1 members in groups A, B, C, D, E and F, respectively. Genes that encode ASCb, NeuroD, Oligo, MyoRb, Figα and Mad were not found in the silkworm genome. The present study provides important background information for future studies using the silkworm as a model system for insect development. Besides, the in-group phylogenetic analysis was demonstrated to be effective in classifying identified bHLH sequences into corresponding families, which can be helpful in the classification of bHLH members of other organisms.

Phylogenetic Analysis of Zebrafish Basic Helix-Loop-Helix Transcription Factors

The basic helix-loop-helix (bHLH) proteins play important regulatory roles in eukaryotic developmental processes including neurogenesis, myogenesis, hematopoiesis, sex determination, and gut development. Zebrafish is a good model organism for developmental biology. In this study, we identified 139 bHLH genes encoded in the zebrafish genome. Phylogenetic analyses revealed that zebrafish has 58, 29, 21, 5, 19, and 5 bHLH members in groups A, B, C, D, E, and F, respectively, while 2 members were classified as “orphan.” A comparison between zebrafish and human bHLH repertoires suggested that both organisms have a certain number of specific bHLH members. Eight zebrafish bHLH genes were found to have multiple coding regions in the genome. Two of these, Bmal1 and MITF, are good anchor genes for identification of fish-specific whole-genome duplication events in comparison with mouse and chicken genomes. The present study provides useful information for future studies on gene family evolution and vertebrate development.

A genome-wide survey on basic helix-loop-helix transcription factors in rat and mouse

The basic helix-loop-helix (bHLH) proteins play essential roles in a wide range of developmental processes in higher organisms. bHLH family members have been identified in over 20 organisms, including nematode, fruit fly, and human. Our study identified 114 rat and 14 additional mouse bHLH members in rat and mouse genomes, respectively. Phylogenetic analyses revealed that both rat and mouse had 49, 26, 15, 4, 12, and 4 bHLH members in groups A, B, C, D, E, and F, respectively. Only the rat Mxi1 gene has two copies in the genome. All other rat bHLH genes and all mouse bHLH genes are single-copy genes. The chromosomal distribution pattern of mouse, rat, and human bHLH genes suggests the emergence of some bHLH genes through gene duplication, which probably happened at least before the divergence of vertebrates from invertebrates. The present study provides useful information for future studies using rat as a model animal for mammalian development.

Phylogenetic analysis of Nosema antheraeae (Microsporidia) isolated from Chinese Oak Silkworm, Antheraea pernyi

ABSTRACT. The microsporidian Nosema antheraeae is a pathogen that infects the Chinese oak silkworm, Antheraea pernyi. We sequenced the complete small subunit (SSU) rRNA gene and the internal transcribed spacer (ITS) of N. antheraeae, and compared the SSU rRNA sequences in other microsporidia. The results indicated that Nosema species, including N. antheraeae, formed two distinct clades, consistent with previous observations. Furthermore, N. antheraeae is clustered with N. bombycis with high bootstrap support. The organization of the rRNA gene of N. antheraeae is LSU–ITS1–SSU–ITS2‐5S, also following a pattern similar to the Nosema type species, N. bombycis. Thus, N. antheraeae is a Nosema species and has a close relationship to N. bombycis.

Determination of Protein Composition and Host-Derived Proteins of Bombyx mori Nucleopolyhedrovirus by 2-Dimensional Electrophoresis and Mass Spectrometry

Objective: The occlusion-derived form of baculovirus is specially adapted for primary infection of host midgut epithelial cells. The virion contains proteins essential for host-range determination and initiation of infection. Determination of protein composition of the occlusion-derived virus (ODV) will help us to understand this virion’s infection mechanism. Methods: We obtained the ODV virion of Bombyx mori nucleopolyhedrovirus (BmNPV) by sucrose density gradient centrifugation, and used SDS-PAGE, 2-dimensional electrophoresis and mass spectrometry to identify – for the first time – proteins present within or associated with the ODV virion. Results: 20 proteins, including 16 major proteins of the virion encoded by BmNPV and 4 proteins of the silkworm, were identified with confidence. Conclusion: Three BmNPV proteins (orf40, P95 and protein tyrosine phosphatase), which had not been detected as components of ODV previously, were identified in this study. We propose that the host-derived proteins may be important for the assembly and maturation of the ODV capsid.

Comparative proteomic analysis reveals that caspase-1 and serine protease may be involved in silkworm resistance to Bombyx mori nuclear polyhedrosis virus.

The silkworm Bombyx mori is of great economic value. The B. mori nuclear polyhedrosis virus (BmNPV) is one of the most common and severe pathogens for silkworm. Although certain immune mechanisms exist in silkworms, most silkworms are still susceptible to BmNPV infection. Interestingly, BmNPV infection resistance in some silkworm strains is varied and naturally existing. We have previously established a silkworm strain NB by genetic cross, which is highly resistant to BmNPV invasion. To investigate the molecular mechanism of silkworm resistance to BmNPV infection, we employed proteomic approach and genetic cross to globally identify proteins differentially expressed in parental silkworms NB and 306, a BmNPV-susceptible strain, and their F(1) hybrids. In all, 53 different proteins were found in direct cross group (NB♀, 306♂, F(1) hybrid) and 21 in reciprocal cross group (306♀, NB♂, F(1) hybrid). Gene ontology and KEGG pathway analyses showed that most of these different proteins are located in cytoplasm and are involved in many important metabolisms. Caspase-1 and serine protease expressed only in BmNPV-resistant silkworms, but not in BmNPV-susceptible silkworms, which was further confirmed by Western blot. Taken together, our data suggests that both caspase-1 and serine protease play a critical role in silkworm resistance against BmNPV infection.

The Basic Helix-Loop-Helix Transcription Factor Family in the Honey Bee, Apis mellifera

Abstract The basic helix-loop-helix (bHLH) transcription factors play important roles in a wide range of developmental processes in higher organisms. bHLH family members have been identified in a dozen of organisms including fruit fly, mouse and human. In this study, we identified 51 bHLH sequences in silico in the honey bee, Apis mellifera L. (Hymenoptera: Apidae), genome. Phylogenetic analyses revealed that they belong to 38 bHLH families with 21, 11, 9, 1, 8 and 1 members in high-order groups A, B, C, D, E and F, respectively. Using phylogenetic analyses, all of the 51 bHLH sequences were assigned to their corresponding families. Genes that encode ASCb, NeuroD, Oligo, Delilah, MyoRb, Figa and Mad were not found in the honey bee genome. The present study provides useful background information for future studies using the honey bee as a model system for insect development.

A genome-wide identification and analysis of the basic helix-loop-helix transcription factors in the ponerine ant, Harpegnathos saltator

BackgroundThe basic helix-loop-helix (bHLH) transcription factors and their homologs form a superfamily that plays essential roles in transcriptional networks of multiple developmental processes. bHLH family members have been identified in over 20 organisms, including fruit fly, zebrafish, human and mouse.ResultIn this study, we conducted a genome-wide survey for bHLH sequences, and identified 57 bHLH sequences encoded in complete genome sequence of the ponerine ant, Harpegnathos saltator. Phylogenetic analysis of the bHLH domain sequences classified these genes into 38 bHLH families with 23, 14, 10, 1, 8 and 1 members in group A, B, C, D, E and F, respectively. The number of PabHLHs (ponerine ant bHLHs) with introns is higher than many other insect species, and they are found to have introns with average lengths only inferior to those of pea aphid. In addition, two H. saltator bHLHs named PaCrp1 and PaSide locate on two separate contigs in the genome.ConclusionsA putative full set of PabHLH genes is comparable with other insect species and genes encoding Oligo, MyoRb and Figα were not found in genomes of all insect species of which bHLH family members have been identified. Moreover, in-family phylogenetic analyses indicate that the PabHLH genes are more closely related with Apis mellifera than others. The present study will serve as a solid foundation for further investigations into the structure and function of bHLH proteins in the regulation of H. saltator development.

Display of Bombyx mori alcohol dehydrogenases on the Bacillus subtilis spore surface to enhance enzymatic activity under adverse conditions.

Alcohol dehydrogenases (ADHs) are oxidoreductases catalyzing the reversible oxidation of alcohols to corresponding aldehydes or ketones accompanied by nicotinamide adenine dinucleotide (NAD) or nicotinamide adenine dinucleotide phosphate (NADP) as coenzyme. ADHs attract major scientific and industrial interest for the evolutionary perspectives, afforded by their wide occurrence in nature, and for their use in industrial synthesis. However, the low activity of ADHs under extremes of pH and temperature often limits their application. To obtain ADH with high activity, in this study, we used Bombyx mori alcohol dehydrogenases (BmADH) as foreign gene and constructed a recombinant integrative plasmid pJS700-BmADH. This pJS700-BmADH was transformed into Bacillus subtilis by double cross-over and produced an amylase inactivated mutant. The fusion protein containing BmADH was expressed on the spore surface and recognized by BmADH-specific antibody. We also assayed the alcohol dehydrogenase activity of the fusion protein together with the native BmADH at different pH and temperature levels, which indicated the recombinant enzyme exhibits activity over wider ranges of temperature and pH than its native form, perhaps due to the resistance properties of B. subtilis spores against adverse conditions.

Arginine kinase is highly expressed in a resistant strain of silkworm (Bombyx mori, Lepidoptera): Implication of its role in resistance to Bombyx mori nucleopolyhedrovirus

A gene encoding Bombyx mori arginine kinase (BmAK) has been indentified differentially expressed in the midguts of Bombyx mori strain NB which is resistant to nucleopolyhedrovirus (BmNPV), strain 306 which is susceptible to NPV and a near isogenic line BC(8) with similar genetic background to 306 but resistant to NPV by two-dimensional gel electrophoresis (2-DE). In this study, we characterized the expression profiles of BmAK using RT-PCR and real-time quantitative PCR. The expression level of BmAK fluctuated in various developing stage and various tissue. Remarkably, the expression level of BmAK increased more than 10-fold 24 hours post inoculation (h p.i.) of NPV in strain NB and BC(8), while such increment was abraded in strain 306 although the basal expression level of BmAK in strain 306 was higher than that of strain NB and BC(8). Western blotting analysis using polyclonal antibody against BmAK verified such observation, and immunofluoresence analysis indicated for the first time that BmAK was mainly located to the cytoplasm or some structures in cytoplasm. These findings suggest that arginine kinase is involved in the antiviral process of Bombyx mori larvae against NPV infection.