Pollen germination, along with pollen tube growth, is an essential process for the reproduction of flowering plants. The germinating pollen with tip-growth characteristics provides an ideal model system for the study of cell growth and morphogenesis. As an essential step toward a detailed understanding of this important process, the objective of this study was to comprehensively analyze the transcriptome changes during pollen germination and pollen tube growth. Using Affymetrix Arabidopsis (Arabidopsis thaliana) ATH1 Genome Arrays, this study is, to our knowledge, the first to show the changes in the transcriptome from desiccated mature pollen grains to hydrated pollen grains and then to pollen tubes of Arabidopsis. The number of expressed genes, either for total expressed genes or for specifically expressed genes, increased significantly from desiccated mature pollen to hydrated pollen and again to growing pollen tubes, which is consistent with the finding that pollen germination and tube growth were significantly inhibited in vitro by a transcriptional inhibitor. The results of Gene Ontology analyses showed that expression of genes related to cell rescue, transcription, signal transduction, and cellular transport was significantly changed, especially for up-regulation, during pollen germination and tube growth. In particular, genes of the calmodulin/calmodulin-like protein, cation/hydrogen exchanger, and heat shock protein families showed the most significant changes during pollen germination and tube growth. These results demonstrate that the overall transcription of genes, both in the number of expressed genes and in the levels of transcription, was increased. Furthermore, the appearance of many novel transcripts during pollen germination as well as tube growth indicates that these newly expressed genes may function in this complex process.

Drought is a major threat to plant growth and crop productivity. Calcium-dependent protein kinases (CDPKs, CPKs) are believed to play important roles in plant responses to drought stress. Here, we report that Arabidopsis thaliana CPK8 functions in abscisic acid (ABA)-and Ca 2+ -mediated plant responses to drought stress. The cpk8 mutant was more sensitive to drought stress than wild-type plants, while the transgenic plants overexpressing CPK8 showed enhanced tolerance to drought stress compared with wild-type plants. ABA-, H 2 O 2 -, and Ca 2+ -induced stomatal closing were impaired in cpk8 mutants. Arabidopsis CATALASE3 (CAT3) was identified as a CPK8-interacting protein, confirmed by yeast two-hybrid, coimmunoprecipitation, and bimolecular fluorescence complementation assays. CPK8 can phosphorylate CAT3 at Ser-261 and regulate its activity. Both cpk8 and cat3 plants showed lower catalase activity and higher accumulation of H 2 O 2 compared with wild-type plants. The cat3 mutant displayed a similar drought stress-sensitive phenotype as cpk8 mutant. Moreover, ABA and Ca 2+ inhibition of inward K + currents were diminished in guard cells of cpk8 and cat3 mutants. Together, these results demonstrated that CPK8 functions in ABA-mediated stomatal regulation in responses to drought stress through regulation of CAT3 activity.

The Arabidopsis Di19 (Drought-induced) gene family encodes seven Cys2/His2-type zinc-finger proteins, most with unknown functions. Here, we report that Di19 functioned as a transcriptional regulator and was involved in Arabidopsis responses to drought stress through up-regulation of pathogenesis-related PR1, PR2, and PR5 gene expressions. The Di19 T-DNA insertion mutant di19 was much more sensitive to drought stress, whereas the Di19-overexpressing lines were much more tolerant to drought stress compared with wild-type plants. Di19 exhibited transactivation activity in our yeast assay, and its transactivation activity was further confirmed in vivo. DNA-binding analysis revealed that Di19 could bind to the TACA(A/G)T element and chromatin immunoprecipitation (ChIP) assays demonstrated that Di19 could bind to the TACA(A/G)T element within the PR1, PR2, and PR5 promoters. qRT-PCR results showed that Di19 promoted the expressions of PR1, PR2, and PR5, and these heightened expressions were enhanced by CPK11, which interacted with Di19 in the nucleus. Similarly to the Di19-overexpressing line, PR1-, PR2-, and PR5-overexpressing lines also showed the drought-tolerant phenotype. The pre-treatment with salicylic acid analogs INA can enhance plants' drought tolerance. Taken together, these data demonstrate that Di19, a new type of transcription factor, directly up-regulates the expressions of PR1, PR2, and PR5 in response to drought stress, and its transactivation activity is enhanced by CPK11.

Triacylglycerol (TAG) is the major seed storage lipid and is important for biofuel and other renewable chemical uses. Acylcoenzyme A:diacylglycerol acyltransferase1 (DGAT1) is the rate-limiting enzyme in the TAG biosynthesis pathway, but the mechanism of its regulation is unknown. Here, we show that TAG accumulation in Arabidopsis (Arabidopsis thaliana) seedlings increased significantly during nitrogen deprivation (0.1 mM nitrogen) with concomitant induction of genes involved in TAG biosynthesis and accumulation, such as DGAT1 and OLEOSIN1. Nitrogen-deficient seedlings were used to determine the key factors contributing to ectopic TAG accumulation in vegetative tissues. Under low-nitrogen conditions, the phytohormone abscisic acid plays a crucial role in promoting TAG accumulation in Arabidopsis seedlings. Yeast one-hybrid and electrophoretic mobility shift assays demonstrated that ABSCISIC ACID INSENSITIVE4 (ABI4), an important transcriptional factor in the abscisic acid signaling pathway, bound directly to the CE1-like elements (CACCG) present in DGAT1 promoters. Genetic studies also revealed that TAG accumulation and DGAT1 expression were reduced in the abi4 mutant. Taken together, our results indicate that abscisic acid signaling is part of the regulatory machinery governing TAG ectopic accumulation and that ABI4 is essential for the activation of DGAT1 in Arabidopsis seedlings during nitrogen deficiency.

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Pollen germination (PG) and pollen tube growth (PTG) play crucial roles in sexual reproduction of flowering plants by sending sperm cells to the ovule. These two processes are regarded as ideal model system for the study of cell signaling and cell polarized growth. It has been considered for a long time that ion transports across the pollen tube membranes are essential for pollen tube navigation and growth. Previous transcriptome analyses for Arabidopsis have shown that the transcripts related to cellular transport are correspondingly overrepresented during the process of pollen tube growth. Here, we showed that 459 transporter genes expressed during PG and PTG in Arabidopsis. In addition, the gene expression profiles of ion (including Ca(2+), H(+), K(+), Cl(-)) channels and transporters were further analyzed. This analysis provides novel information for the potential candidate genes involving in ion fluxes across the pollen tube membranes and in regulation of pollen tube tip growth.

Triacylglycerol (TAG) is the major storage component accumulated in seed. However the regulatory mechanism of TAG synthesis and accumulation in non-seed tissues remains unknown. Recently, we found that nitrogen (N) deficiency (0.1mM N) caused an inducement of TAG biosynthesis in Arabidopsis seedlings. ABSCISIC ACID INSENSITIVE 4 (ABI4) was essential for the activation of Acyl-CoA:diacylglycerol acyltransferase1(DGAT1) expression during N deficiency in Arabidopsis seedlings. In this addendum, we further discussed the approaches to provide a net increase in total oil production in higher plants by using the low N platform. First, the N-deficient seedlings can be used to determine the key factors that regulate the ectopic expression of key genes in TAG metabolism. Second, the research on the relationship between TAG homeostasis and cell division will be helpful to find the key factors that specifically regulate TAG accumulation under the nutrient-limited condition.