sal1 determines the number of aleurone cell layers in maize endosperm and encodes a class E vacuolar sorting protein

A microscopy-based screen of a large collection of maize Mutator (Mu) transposon lines identified the supernumerary aleurone layers 1-1 (sal1-1) mutant line carrying up to seven layers of aleurone cells in defective kernel endosperm compared with only a single layer in wild-type grains. Normal, well filled endosperm that is homozygous for the sal1-1 mutant allele contains two to three layers of aleurone cells. Cloning of the sal1 gene was accomplished by using Mu tagging, and the identity of the cloned gene was confirmed by isolating an independent sal1-2 allele by reverse genetics. Homozygous sal1-2 endosperm has two to three layers of aleurone cells in normal, well filled grains. In situ hybridization experiments reveal that the sal1 gene is ubiquitously expressed in vegetative as well as zygotic grain tissues, with no difference being detected between aleurone cells and starchy endosperm cells. Northern blot analysis failed to detect the sal1-2 transcript in leaves of homozygous plants, suggesting that the allele is a true sal1 knockout allele. The sal1 gene encodes a homologue of the human Chmp1 gene, a member of the conserved family of the class E vacuolar protein sorting genes implicated in membrane vesicle trafficking. In mammals, CHMP1 functions in the pathway targeting plasma membrane receptors and ligands to lysosomes for proteolytic degradation. Possible roles for the function of the sal1 gene in aleurone signaling, including a defect in endosome trafficking, are discussed.