Molecular characterization and immunohistochemical localization of a novel troponin C during silkworm development

Abstract

We have cloned and sequenced a novel Bombyx mori gene that encodes a protein having a high degree of homology with other known troponin C (TnC) proteins. The amino acid sequence, DX[DN]X[DSG]X 6 E, a highly conserved putative Ca 2+ -binding motif found in loops within the globular domains of previously identified TnC proteins, is also present in BmTnC. We have expressed and purified to homogeneity a His-tagged BmTnC fusion protein having a molecular weight of approximately 21.6 kDa. We have used this purified fusion protein to produce polyclonal antibodies against BmTnC for Western blot analyses. These analyses have revealed that BmTnC is expressed in the larval head, the Malpighian tubule, the epidermis, the testis, and the gut, as has been confirmed by immunohistochemistry. In addition, real-time reverse transcription/polymerase chain reaction has shown that BmTnC mRNA levels differ substantially among these tissues. Our findings indicate that BmTnC is selectively expressed in the muscular tissues of the silkworm, including portions of the head, the Malpighian tubule, the body wall, and the gut.

RNA interference‐mediated knockdown and virus‐induced suppression of Troponin C gene adversely affect the behavior or fitness of the green rice leafhopper, Nephotettix cincticeps

“…Compared with the thorax and the abdomen, transcript level of Nc‐TnC was relative higher in the intestinal canal, head, and the foot (Figure b). The expression pattern was consistent with that reported recently for other invertebrate insects (Chen et al., ; Feng et al., ; Herranz, Mateos, & Marco, ; Wu, Xu, & Lin, ; ), suggesting the similar functions of TnC protein in modulating the behavior of these insects.…”

“…Each EF‐hand consists of two α‐helixes connected by a flexible loop, forming what is known as a helix‐loop‐helix. The loop, which contains a potential Ca 2+ ‐binding site, has a characteristic sequence of 12 amino acid residues, D‐{W}‐[DNS]‐{ILVFYW}‐[DENSTG]‐[DNQGHRK]‐{GP}‐[LIVMC]‐[DENQSTAGC]‐X2‐[DE] (Chen et al., ; Herzberg & James, ). Two of these motifs were also present in the deduced amino acid sequence of the Nc‐TnC protein (Figures.…”

Bombyx moriTranscription Factors: Genome-Wide Identification, Expression Profiles and Response to Pathogens by Microarray Analysis

“…This may be related to species-specific gene expansion in some families. For example, in the largest family C2H2-type zinc finger, 55.6% of the silkworm genes were species-specific; in Drosophila , the proportion was lower at 43.9% (Duan et al 2008). In another example, in the second largest family homeobox domain, a new group of 12 genes ( Bmshx ) in silkworm had a relatively independent cluster relative to other insects according to phylogenetic analysis, indicating that shx subfamily was species-specific in silkworm (Chai et al 2008).…”

Molecular Characterization, Tissue Distribution, Subcellular Localization and Actin-Sequestering Function of a Thymosin Protein from Silkworm

“…Polyclonal antibody was prepared by immunizing New Zealand White rabbits using purified rBmTHY as antigen [25]. Subsequently, 100 mg of rBmTHY (equal to about 200 ml of the antigen/adjuvant mix) was injected into each of 8–10 subcutaneous sites at the back of the rabbit.…”