Molecular Characterization, mRNA Expression and Alternative Splicing of Ryanodine Receptor Gene in the Brown Citrus Aphid, Toxoptera citricida (Kirkaldy).

Ke-Yi Wang,Xuan-Zhao Jiang,Feng Shang,Guo-Rui Yuan,Jin-Jun Wang

doi:10.3390/ijms160715220

Ke-Yi Wang, Xuan-Zhao Jiang + Show 3 more

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https://doi.org/10.3390/ijms160715220

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Abstract

Ryanodine receptors (RyRs) play a critical role in regulating the release of intracellular calcium, which enables them to be effectively targeted by the two novel classes of insecticides, phthalic acid diamides and anthranilic diamides. However, less information is available about this target site in insects, although the sequence and structure information of target molecules are essential for designing new control agents of high selectivity and efficiency, as well as low non-target toxicity. Here, we provided sufficient information about the coding sequence and molecular structures of RyR in T. citricida (TciRyR), an economically important pest. The full-length TciRyR cDNA was characterized with an open reading frame of 15,306 nucleotides, encoding 5101 amino acid residues. TciRyR was predicted to embrace all the hallmarks of ryanodine receptor, typically as the conserved C-terminal domain with consensus calcium-biding EF-hands (calcium-binding motif) and six transmembrane domains, as well as a large N-terminal domain. qPCR analysis revealed that the highest mRNA expression levels of TciRyR were observed in the adults, especially in the heads. Alternative splicing in TciRyR was evidenced by an alternatively spliced exon, resulting from intron retention, which was different from the case of RyR in Myzus persicae characterized with no alternative splicing events. Diagnostic PCR analysis indicated that the splicing of this exon was not only regulated in a body-specific manner but also in a stage-dependent manner. Taken together, these results provide useful information for new insecticide design and further insights into the molecular basis of insecticide action.

Highlights

Intracellular calcium (Ca2+) homeostasis is crucial for normal life activities because Ca2+ is a key second messenger that regulates a number of cellular processes, such as muscle contraction, hormone secretion, synaptic transmission, and gene transcription [1]
Three Ryanodine receptors (RyRs) isoforms have been reported: RyR1, which is predominant in skeletal muscles, RyR2 is largely present in cardiac muscles, and RyR3, which is primarily present in the brain and diaphragm [6,7]
A sequence of 15,639 bp was assembled from ten overlapping cDNA products (Figure 1), which covered the complete open reading frame (ORF) of 15,315 bp

Summary

Introduction

Intracellular calcium (Ca2+) homeostasis is crucial for normal life activities because Ca2+ is a key second messenger that regulates a number of cellular processes, such as muscle contraction, hormone secretion, synaptic transmission, and gene transcription [1]. Ryanodine receptors (RyRs), known as Ca2+ release channels, play an critical role in intracellular Ca2+ signaling by inducing the release of. RyRs are homomeric tetramers with a total molecular mass higher than 2 MDa [4], composed of a hydrophilic domain at the N-terminus and a membrane-spanning domain at the. The N-terminal cytoplasmic domain, known as the “foot structure”, is large and spans the junction between the transverse tubules and SR membranes. The C-terminal hydrophobic domain contains 4–12 transmembrane segments and forms the putative pore of the Ca2+ release channel [4,6]. Three RyR isoforms have been reported: RyR1, which is predominant in skeletal muscles, RyR2 is largely present in cardiac muscles, and RyR3, which is primarily present in the brain and diaphragm [6,7]. In birds, amphibians, and fish only two RyR isoforms (RyRA and RyRB), which are closely related to the mammalian RyR1 and RyR3, have been identified [8,9]

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Discussion

Conclusion