Analysis of the promoter activity of late embryogenesis abundant protein genes in barley seedlings under conditions of water deficit

Promoters of the late embryogenesis abundant protein genes, HVA1s, Dhn8s and Dhn4s from barley and wsi18j and rab16Bj from rice, were analysed in barley seedlings to assess their strength and timing of induction under water deficit conditions using a transient expression system. Of the drought-inducible promoters, Dhn4s exhibited the highest activity, followed by HVA1s, wsi18j and rab16Bj. Dehydration-induced #-glucuronidase expression levels driven by the Dhn4s, HVA1s and wsi18j promoters were higher than or comparable to that induced by a strong constitutive rice Act1 promoter. The expression patterns of wsi18j and Dhn8s differed from those of their homologous genes (wsi18 and Dhn8): the wsi18j promoter was strongly induced by abscisic acid, while the Dhn8s promoter was highly active in hydrated seedlings. HVA1s- and wsi18j-driven green fluorescent protein expression was induced within 1 h of drying under water deficit stress, while Dhn4s promoter activity was not detectable until 3 h, thus showing a difference in the timing of induction.     

Food stress and predator-induced stress shape developmental performance in a damselfly

I studied effects of stress factors like food shortage, non-lethal predator presence and autotomy on survival and larval performance (growth rate, development rate and developmental stability) of larvae of the damselfly Lestes sponsa. In a laboratory experiment, larvae were raised during their last two instars at two food levels (high or low) crossed with two levels of autotomy (caudal lamellae present or absent). These treatments were nested within three levels of predation risk (Aeshna cyanea absent, Chironomus-fed caged Aeshna or Lestes-fed caged Aeshna). The diet of the predator had no effects. The low food level and the presence of Aeshna independently increased mortality rates of L. sponsa larvae. The low food level, presence of a caged Aeshna and autotomy all independently reduced growth rate (mass and body size at day 40) and wing size at emergence, and the first two stress factors also reduced development rate. Regardless of predator presence and autotomy, all damselfly larvae consumed the food available. This indicated that the predator-induced stress effects were not due to reduced food uptake, but probably reflected lowered assimilation efficiency and/or a higher metabolic rate. Besides a low food level, the presence of caged Aeshna predator larvae and autotomy also increased hind wing asymmetry. This result demonstrated that predator-induced stress may reduce developmental stability in the prey.     

Isolation and characterization of lipid transfer protein (LTP) genes from a wheat-rye translocation line

. Two genes (TaLTP1 and TaLTP2) encoding lipid transfer proteins (LTPs) were isolated from a cDNA library constructed from leaf tissue harvested from 4-week-old seedlings of a wheat-rye near-isogenic line (NIL) involving a translocation of rye chromosome 2RL with wheat 2BS. The spatial and temporal patterns of expression of TaLTP1 and TaLTP2 were examined by Northern blot analysis and in situ hybridization. Both TaLTP1 and TaLTP2 contained a 270-bp open reading frame and encoded a putative LTP precursor molecule of 90 amino acids. Expression of the two LTPs was detected in leaves, stems, and crowns of the NILs but not in the roots. The expression levels of TaLTP1 and TaLTP2 remained constant in response to cold and ABA treatments over a period of 24 h but increased 3 days after the initiation of drought stress. An in situ hybridization study indicated that TaLTP1 was expressed in the cells within the vascular bundles of leaves and in the tissue layers between the vascular bundles in the crowns of the control and drought-treated plants. Expression of TaLTP1 in the tissue layers between the vascular bundles was higher in the drought-treated plants than in the control plants. The results suggested that high levels of expression of TaLTPs in the tissue layers between the vascular bundles might play a role in the drought tolerance response of the wheat crown.     

Plant CDK inhibitors: studies of interactions with cell cycle regulators in the yeast two-hybrid system and functional comparisons in transgenic Arabidopsis plants

. The cyclin-dependent kinase (CDK) inhibitors ICK1 and ICK2 have been shown to inhibit plant CDK activity in vitro, and the expression of ICK1 was able to inhibit cell division in the plant and modify plant growth and morphology. In order to characterize other ICK1-related inhibitor genes and understand possible differences among plant CDK inhibitors, the interactions of plant CDK inhibitors with cell cycle regulators were analysed in the yeast two-hybrid system and their functions were compared in transgenic Arabidopsis plants. Yeast two-hybrid results indicate that there are likely two groups of plant CDK inhibitors. The A-group inhibitors ICK1, ICK2, ICK6 and ICK7 interact with Cdc2a and three D-type cyclins (D1, D2 and D3), while the B-group inhibitors ICK4, ICK5 and ICKCr interact with D-type cyclins but not with Arabidopsis Cdc2a. ICK1 (A-group), and ICK4 and ICKCr (B-group) were expressed separately in transgenic Arabidopsis plants. Overexpression of the three inhibitor genes resulted in plants of a smaller size with serrated leaves and modified flowers. These plants also had reduced nuclear DNA content (polyploidy), suggesting that expression of these inhibitors affected endoreduplication. Further, there were apparent differences in the strength of effect among the inhibitors. These results provide the first evidence on the CDK inhibitory function for ICK4 and ICKCr. They also suggest that these CDK inhibitors play important roles in cell division and plant growth.     

Identification of anther-specific genes in a cruciferous model plant, Arabidopsis thaliana, by using a combination of Arabidopsis macroarray and mRNA derived from Brassica oleracea

Arabidopsis thaliana, a member of the Brassicaceae, is a model plant whose genome was the first higher plant genome to be sequenced. Because of the small size of the flowers, it is difficult to dissect and separate reproductive organs (anthers and pistils) at different developmental stages in A. thaliana. In order to perform genome-wide identification of anther-specific genes in A. thaliana, an Arabidopsis cDNA macroarray was hybridized to cDNA derived from anthers and pistils of another crucifer, Brassica oleracea. After scanning the signal intensity for each clone, and cluster analysis, 52 anther-specific genes were identified. These clones contained several anther-specific genes that have already been characterized, as well as novel anther-specific genes. In RT-PCR analysis with mRNA of A. thaliana and B. oleracea, the expression pattern of one-third of the clones was similar to that determined by cDNA macroarray. This system of heterologous hybridization analysis (Arabidopsis cDNA macroarray vs Brassica tissue-specific mRNA) should be applicable to other model species and their close relatives.