Understanding tick salivary secretions using RNA interference (RNAi).

摘要

RNAi is a recently discovered powerful tool to silence gene expression in a sequence specific manner at the mRNA level. Protein secretion from the female tick salivary glands occur by regulated exocytosis and requires the participation of proteins called SNAREs (Soluble N-ethylmaleimide sensitive factor Attachment protein Receptor). Members of this class of proteins are expressed in all eukaryotic cells and are designated as vesicle (v-SNARE) and target (t-SNARE) SNARE depending on the membranes they are associated with. Regulated exocytosis, in addition, requires the functions of several SNARE interacting proteins. Synaptobrevin is a key v-SNARE and nSec1 is a key t-SNARE interacting protein in many secreting cells studied till date. cDNA fragment homologues of synaptobrevin and nSec1 were cloned and sequenced from the salivary glands of partially fed female ticks. Synaptobrevin and nSec1 double stranded RNAs (dsRNA) were used to specifically reduce the amount of synaptobrevin and nSec1 mRNA and protein levels respectively. This decrease was accompanied by a decrease in anticoagulant protein release from the glands. In addition, nSec1 dsRNA was injected into unfed female ticks and allowed to feed on sheep. We were able to demonstrate differences in the transcript and protein levels with reduced protein secretion and slower growth patterns in ticks injected with dsRNA.