Targeting cyclin-dependent kinases in Drosophila with peptide aptamers

Abstract

Two-hybrid technology provides a simple way to isolate small peptide aptamers that specifically recognize and strongly bind to a protein of interest. These aptamers have the potential to dominantly interfere with specific activities of their target proteins and, therefore, could be used as in vivo inhibitors. Here we explore the ability to use peptide aptamers as in vivo inhibitors by expressing aptamers directed against cell cycle regulators in Drosophila. We expressed two peptide aptamers, each of which specifically recognizes one of the two essential cyclin-dependent kinases (Cdks), DmCdk1 and DmCdk2, in Drosophila. Expression of each Cdk aptamer during organogenesis caused adult eye defects typical of those caused by cell cycle inhibition. Co-overexpression of DmCdk1 or DmCdk2 resulted in suppression of the eye phenotypes, indicating that each aptamer interacts with a Cdk target in vivo and suggesting that these peptides disrupt normal eye development by inhibiting Cdk function. Moreover, the specificity of each aptamer for one of the two Cdks as determined in two-hybrid assays was retained in Drosophila. Combined, our results demonstrate that peptide aptamers generated by yeast two-hybrid methods can serve as inhibitory reagents to target specific proteins in vivo.

Peptide-mediated broad-spectrum plant resistance to tospoviruses

“…18) have successfully been used in yeast cells, E. coli, Drosophila, and cultured human cells. They are a useful tool for basic research, but they also represent a new approach in drug target validation and drug discovery (18,19,(31)(32)(33)(34). The application of dominant negative peptide aptamers to inhibit protein functions in vivo in a living multicellular organism has been reported so far only for Drosophila (19,35).…”

“…The strategy exemplifies a promising new concept that uses target-specific peptides (''aptamers''), selected with the yeast two-hybrid system, to modulate͞inhibit protein functions in vivo (18,19).…”

Sequence-specific Peptide Aptamers, Interacting with the Intracellular Domain of the Epidermal Growth Factor Receptor, Interfere with Stat3 Activation and Inhibit the Growth of Tumor Cells

“…The interaction between a specific peptide sequence inserted into the active loop of the scaffold protein thioredoxin A and the target protein domain of interest restores the activity of a Gal4 transcription factor and allows the growth of yeast cells under selection. Several examples have been described in which aptamers were used to interfere with specific protein interactions in vivo and thus elicit distinct cellular phenotypes (22)(23)(24)(25)(26)(27)(28).…”

Isolation of peptide aptamers that inhibit intracellular processes

“…For example, overexpression of an aptamer's target might be predicted to reverse the phenotype attributed to the aptamer (20). To demonstrate the feasibility of this approach, and to verify that ThyA is the target of the aptamer encoded by pBMT101Q, we set up a system for inducing expression of both ThyA and the aptamer in the same cells under independent control.…”