Preliminary Analyses Of ERECTA Family Gene Expression And Functions In Irce

Arabidopsis ERECTA gene family, which encodes a transmembrane signal receptorsubfamily, is composed of three members known as AtER, AtER-LIKE1(AtERL1) andAtERL2. Arabidopsis ERECTA family genes are widely expressed in many tissues and encodea serine/threonine protein kinase. ERECTA genes have been reported to regulate thedifferenation and morphogenesis of aerial organs including leaf, stomata and flower bycoordinating the cell division and cell expansion. In addition, ERECTA genes are alsoinvolved in many physiological processes such as the Rubisco carboxylation rate, electrontransport capacity, plant transpiration, as well as responses to biotic and abiotic stress such asdrought and bacteria infection. These findings demonstrate that ERECTA genes play importantroles in plant growth and development. To our knowledgr, however the information about riceERECTA genes has not been widely reported. To explore the roles of ERECTA genes in rice,we predicted three members of ER family in rice genome, and analyzed its expression patternand preliminary function in rice growth and development. The major results we obtained areas followed:1Expression patterns of rice ERECTA family genes(1) We surveyed the rice genome and found that the ERRECTA family has three members:ERECTA (ER)(Os06g0203800), ERECTA-LIKE1(ERL1)(JN991005) and ERL2(Os06g0130100). Real-time PCR analysis showed that ERECTA family genes express higherin meristem or reproductive organ, such as young panicle, but relatively lower in matureabove-ground organs, such as stem, leaf and flag leaf. ERECTA family genes express lowestin root.(2) Real-time PCR performed with RNA from WT and phyB mutant seedlings grownunder red light and dark revealed that ERECTA family genes in WT grown in the darkexpressed higher than they did under red light. This result suggests that red light can inhibitERECTA family genes expression. But this inhibitory effect was reduced in phyB mutant bycomparison with WT, which indicate that phyB play major roles in red light-triggeredinhibition of ERECTA family genes expression. However, red light did not inhibit the ERECTA family genes expression in phyAphB mutants, suggesting that both phyA and phyBcan perceive red light to inhibit ERECTA family genes expression in rice.2Introns are essential for ERECTA expressionWe produced two types of transgenic rice plants either with ERL1gene with intronlessERL1cDNA fragment (cERL1) or with intron-containing DNA fragment (gERL1). RT-PCRanalyses revealed that no significant difference in ERL1s transcript levels was observedbetween cERL1transgenic plant and WT. However, the expression of ERL1was significantlyincreased in gERL1transgenic plant relative to that in WT. These results suggest that intronsare essential for ERL1gene expression.3The function of ERECTA in rice growth and development(1) To examine the role of ERL1in rice drought tolerance, we compared the phenotypes ofgERL1transgenic plant and WT treated with15%PEG4000. WT leaves turned yellow andwithered at8th day after PEG treatment, whereas the leaves of transgenic plants still exhibitedturgor and green. Almost all of WT plants wilted after being treated with PEG for16days andthen recovered for8days. However, most of transgenic plants were recovered and grow well.These results suggest that over-expression of ERL1enhance drought tolerance in rice.(2) Seeds of gERL1transgenic lines were longer than WT, indicating that ERL1isprobably involved in controling the organ size.(3) The erl2mutant, which is single-site insertion mutant, was screened from rice TOS17mutant library. The erl2mutant displayed normal vegetative growth whereas the seed-settingrate is obviously decreased. The anatomical analyses of pistil and stamen revealed that pistilin erl2mutant appears indistinguishable from WT, but the length of anther was about half ofthat in WT.4Rice ERECTA gene encode a serine/threonine protein kinaseMal-ER fusion protein was constructed by cloning of ER cDNA into pMal-c2x vector.This construct was transformed into E. coli stain for expression. Cells were harvested bycentrifugation and the maltose binding protein (MBP)-ER fusion protein was purified. Thephosphoaminoacid analysis of recombinant protein showed that MBP-ER wasautophosphorylates on serine and threonine. The result demonstrated that ER encodes anactive serine/threonine protein kinase.