An abundant TIP expressed in mature highly vacuolated cells

Aquaporins are water channel proteins found in vacuolar membranes and plasm a membranes, and belong to the major intrinsic protein (MIP) family of prot eins. In the present study, we purified a 75 kDa MIP protein from a crude f raction of spinach leaf intracellular membranes. Upon urea/SDS-PAGE, the 75 kDa protein appeared as a 21 kDa polypeptide, and the 75 kDa species there fore probably represents a tetramer. The corresponding cDNA was obtained by PCR cloning and had an open reading frame encoding a 25.1 kDa protein. The protein, So-delta TIP, was most homologous to the tonoplast intrinsic prot ein (TIP) subfamily of plant MIPs. Using affinity-purified So-delta TIP-spe cific peptide antibodies, we investigated the subcellular and tissue distri bution of So-delta TIP. So-delta TIP was specifically located in the vacuol ar membrane. It was abundant in most vacuolated cells in all vegetative org ans, but was excluded from the leaf epidermis as well as from the root phlo em parenchyma and meristem. In spite of the high sequence homology between delta-TIPs of spinach, Arabidopsis, sunflower and radish, their expression patterns were totally different. However, a comparison of the expression pa ttern of So-delta TIP with that of more distantly related TIPs showed simil arities with Arabidopsis gamma-TIP, which is expressed in zones of cell elo ngation/differentiation but excluded from meristematic tissues. Meristemati c cells are characterized by many small vacuoles as opposed to elongating a nd mature cells, which generally harbour a single, large vacuole. Our resul ts indicate that the expression of So-delta TIP may be induced when the lar ge vacuole is formed.