Minisymposium 23: Protein targeting

Abs # 44005: mRNA targeting to endoplasmic reticulum subdomains regulates protein localization in rice endosperm cells

Presenter:	Washida, Haruhiko , was@wsu.edu
Authors	Washida, Haruhiko  (A)   Sugino, Aya  (A)   katsube-Tanaka, Tomoyuki  (A)   Sakulsingharoj, Chotipa  (A)   Kaneko, Sachiyo  (A)   Wang, Changlin  (A)   Okita, Thomas W (A)
Affiliations:	(A): Institute of Biological Chemistry, Washington State University

The rice storage proteins are stored in different endomembrane compartments; prolamines assemble within the endoplasmic reticulum (ER) to form ER-bounded spherical protein bodies (PB-I), whereas glutelins accumulate within storage vacuoles (PB-II). The packaging of these proteins into different intracellular compartments may be facilitated by their site of synthesis as their RNAs are segregated on different ER subdomains, protein body-ER (PB-ER, prolamines) and cisternal ER (cis-ER, glutelins). Developing maize seeds also synthesize and accumulate prolamine- (zein) and 11S globulin- (legumin-1) type proteins, which are sequestered into separate endomembrane compartments. in situ RT-PCR analysis of maize endosperm sections showed that the zein and legumin-1 RNAs display a distribution pattern on the ER similar to that seen in rice. Zein RNAs are associated with ER-bounded, zein-containing PBs, whereas the 51 kD legumin-1 RNA is localized to cis-ER. Transgenic rice expressing sunflower 2S albumin (2SA), maize 10 kD zein and hybrid storage protein genes were studied to explore the relationship between RNA and protein localization. 10 kD zein- nos 3'UTR RNAs are distributed on the PB-ER and the proteins localized within PB-I. Replacement of the nos 3'UTR with corresponding sequences from glutelin RNA, which contains a cis-ER targeting zip code, results in re-directing the localization of 10kD zein-3'glutelin RNA from PB-ER to cis-ER and the coded zein protein from PB-I to PB-II. Similar to glutelins, 2SA RNAs are targeted to cis-ER and the protein localized to PB-II. However, 2SA hybrid RNAs containing the 10kD zein signal peptide coding sequence, which contains a PB-ER targeting zip code, results in localization to both cis and PB-ER with corresponding accumulation of 2SA protein in both PB-I and II. These results indicate that seed storage protein localization within the endomembrane compartments is correlated closely with the distribution of the RNAs to specific ER subdomains. Transgenic rice expressing this hybrid zein-2SA accumulated significantly larger quantities of 2SA protein suggesting that RNA transport may be a limiting process in gene expression.