Poster: Organelle Biogenesis

Abs # 1142: Tissue specific expression of rice tonoplast intrinsic protein isoforms

Presenter:	Takahashi, Hideyuki , takahashi@kab.seika.kyoto.jp
Authors	Takahashi, Hideyuki  (A)   Rai, Mika  (A)   Kitagawa, Tomoya  (A)   Morita, Shigeto  (A) (B)  Masumura, Takehiro  (A) (B)  Tanaka, Kunisuke  (A) (B)
Affiliations:	(A): Laboratory of Genetic Engineering, Kyoto Prefectural University (B): Kyoto Prefectural Insutitute of Agricultural Biotecnology

Protein storage vacuoles (PSVs), containing seed-type storage proteins, and lytic vacuoles (LVs), marked by the presence of active proteases, are considered as discrete organelles. PSVs are marked by the presence of a-tonoplast intrinsic protein (TIP), whereas LVs are marked by g-TIP. To investigate the relation between existence of specific TIP isoforms and the different vacuole functions, we isolated three novel cDNAs from rice that are homologous to rice g-TIP. Every amino acid sequence, deduced from the corresponding cDNA, contained a typical structure of the aquaporin family identified so far. To determine the expression patterns of them, total RNA fractions were prepared from different tissues and various developmental stages. Northern blot hybridization analyses revealed that g-TIP was expressed in all plant organs. In seeds, the transcripts for g-TIP were detected predominantly in embryo. However, OsTIP1 was specifically expressed in seed, and the transcripts were detected in both embryo and endosperm. OsTIP2 was expressed in mature seeds and during seed germination. OsTIP3 was expressed exclusively in roots. These suggest that each TIP isoform is classified into a distinct group of the TIP family and might have specific function. To examine the subcellular localization of g-TIP and OsTIP1, we constructed fusion genes for both TIPs with sGFP (S65T) and introduced in onion epidermal cells using a particle bombardment method. Fluorescence of gTIP-GFP and TIP1-GFP was observed as a single fluorescent line surrounding the vacuoles. These results suggest that rice g-TIP is located in the tonoplast of LVs similar to g-TIP of other plants. On the other hand, OsTIP1 is located in the tonoplast of PSVs similar to a-TIP.