Maize <i>defective kernel</i> mutant generated by insertion of a <i>Ds</i> element in a gene encoding a highly conserved TTI2 cochaperone.

Abstract

We have used the newly engineered transposable element <i>Dsg</i> to tag a gene that gives rise to a defective kernel (<i>dek</i>) phenotype. <i>Dsg</i> requires the autonomous element <i>Ac</i> for transposition. Upon excision, it leaves a short DNA footprint that can create in-frame and frameshift insertions in coding sequences. Therefore, we could create alleles of the tagged gene that confirmed causation of the <i>dek</i> phenotype by the <i>Dsg</i> insertion. The mutation, designated <i>dek38-Dsg</i>, is embryonic lethal, has a defective basal endosperm transfer (BETL) layer, and results in a smaller seed with highly underdeveloped endosperm. The maize <i>dek38</i> gene encodes a TTI2 (Tel2-interacting protein 2) molecular cochaperone. In yeast and mammals, TTI2 associates with two other cochaperones, TEL2 (Telomere maintenance 2) and TTI1 (Tel2-interacting protein 1), to form the triple T complex that regulates DNA damage response. Therefore, we cloned the maize <i>Tel2</i> and <i>Tti1</i> homologs and showed that TEL2 can interact with both TTI1 and TTI2 in yeast two-hybrid assays. The three proteins regulate the cellular levels of phosphatidylinositol 3-kinase-related kinases (PIKKs) and localize to the cytoplasm and the nucleus, consistent with known subcellular locations of PIKKs. <i>dek38-Dsg</i> displays reduced pollen transmission, indicating TTI2's importance in male reproductive cell development.