ABSTRACT. Several receptor proteins of Cry toxin have been previously identified, including cadherin-like, aminopeptidase N, and alkaline phosphatase. In the present work, a novel binding protein, V-ATPase subunit A (HpVAA), was identified in Holotricia parallela larvae and characterized. We performed reverse transcriptionpolymerase chain reaction and rapid amplification of cDNA ends technology to obtain the cDNA of the full-length hpvaa. Sequencing analysis showed that the open reading frame of hpvaa (GenBank accession No. KU497557) is 1845 bp long, encoding 614 amino acid residues. The predicted molecular weight and isoelectric point of HpVAA were 67.85 kDa and 4.9, respectively. The HpVAA protein, which includes two putative conserved domains, ATP-synt_ab_N and ATP-synt_ab_C, and a Walker A (GAFGCGKT) motif and a Walker B (SMMAD) motif, possesses the same structural characteristics as V-ATPase subunit A from other insects. The protein was successfully 2 W. Wei et al. Genetics and Molecular Research 15 (3): gmr.15038994 expressed in Escherichia coli, and a ligand blot assay showed binding of the protein with Cry8Ea3 toxin. Transcriptional analysis of hpvaa in different tissues of H. parallela larvae was performed by qRT-PCR, which showed that the relative expression of hpvaa in the Malpighian tubules is higher than that in other tissues.

No abstract

Chitin deacetylases (CDAs) are critical enzymes required for chitin degradation, which play an important role in modifying the physical properties of insect cuticles and peritrophic matrices. A novel chitin deacetylase gene (hccda5) was identified from Hyphantria cunea. The recombinant expressed HcCDA5 protein, containing a chitin deacetylase domain (CDAD), did not show chitin deacetylase activity, but instead showed a strong chitin‐binding activity. Transcriptional analysis using qRT‐PCR indicated that hccda5 was mainly expressed in the midgut of H. cunea larvae. Feeding H. cunea larvae with Bt (Bacillus thuringiensis) Cry1A protein induced its downregulation. The gene of hccda5 was successfully silenced by different dsRNAs in insect cell line (Spodoptera frugiperda line, Sf9), which provided a tool to study the function of an expressed gene in insect cell line and laid a foundation for RNAi of hccda5 in H. cunea larvae.

No abstract