Selective knockdowns in maize by sequence-specific protein aggregation

Protein aggregation is determined by 5-15 amino acids peptides of the target protein sequence, so-called aggregation-prone regions (APRs) that specifically self-associate to form β-structured inclusions. The presence of APRs in a target protein can be predicted by a dedicated algorithm, such as TANGO. Synthetic aggregation-prone proteins are designed by expressing specific APRs fused to a fluorescent carrier for stability and visualization. Previously, the stable expression of these proteins in Zea mays (maize) has been demonstrated to induce aggregation of target proteins with specific localization, such as the starch-degrading enzyme α-glucan water dikinase, giving rise to plants displaying knockdown phenotypes. Here, we describe how to design synthetic aggregation-prone proteins to harness the sequence specificity of APRs to generate aggregation-associated phenotypes in a targeted manner and in different subcellular compartments. This method points toward the application of induced targeted aggregation as a useful tool to knockdown protein functions in maize and to generate crops with improved traits.