Arabidopsis WRKY proteins are plant-specific transcription factors, encoded by a large gene family, which contain the highly conserved amino acid sequence WRKYGQK and the zinc-finger-like motifs, Cys 2 His 2 or Cys 2 HisCys. They can recognize and bind the TTGAC(C/T) W-box ciselements found in the promoters of target genes, and are involved in the regulation of gene expression during pathogen defense, wounding, trichome development, and senescence. Here we investigated the physiological function of the Arabidopsis WRKY22 transcription factor during dark-induced senescence. WRKY22 transcription was suppressed by light and promoted by darkness. In addition, AtWRKY22 expression was markedly induced by H 2 O 2 . These results indicated that AtWRKY22 was involved in signal pathways in response to abiotic stress. Darktreated AtWRKY22 over-expression and knockout lines showed accelerated and delayed senescence phenotypes, respectively, and senescence-associated genes exhibited increased and decreased expression levels. Mutual regulation existed between AtWRKY22 and AtWRKY6, AtWR-KY53, and AtWRKY70, respectively. Moreover, AtWRKY22 could influence their relative expression levels by feedback regulation or by other, as yet unknown mechanisms in response to dark. These results prove that AtWRKY22 participates in the dark-induced senescence signal transduction pathway.

Arabidopsis thaliana WRKY39, a transcription factor that is induced by heat stress, is a member of the group II WRKY proteins and responds to both abiotic and biotic stress. Heat-treated seeds and plants of WRKY39 knock-down mutants had increased susceptibility to heat stress, showing reduced germination, decreased survival, and elevated electrolyte leakage compared with wild-type plants. In contrast, WRKY39 over-expressing plants exhibited enhanced thermotolerance compared with wild-type plants. RT-PCR and qRT-PCR analysis of wrky39 mutants and WRKY39 over-expressing plants identified putative genes regulated by WRKY39. Consistent with a role for WRKY39 in heat tolerance, the expression levels of salicylic acid (SA)-regulated PR1 and SA-related MBF1c genes were downregulated in wrky39 mutants. In contrast, over-expression of WRKY39 increased the expression of PR1 and MBF1c. The WRKY39 transcript was induced in response to treatment with SA or methyljasmonate. Analysis of heat stressinduced WRKY39 in defense signaling mutants, including coi1, ein2, and sid2, further indicated that WRKY39 was positively co-regulated by the SA and jasmonate (JA) signaling pathways. Together, these findings reveal that heat stress-induced WRKY39 positively regulates the cooperation between the SA-and JA-activated signaling pathways that mediate responses to heat stress.

WRKY proteins are a superfamily of plant transcriptional factors with potential regulatory roles pertaining to a variety of biotic and abiotic stress responses. In this study, we investigated the expression profiles of Os-WRKY23 under different developmental stages, pathogen infection, continuous-dark and hormone treatments. Under normal growth conditions, OsWRKY23 expressed exclusively in roots and senescing leaves. Under biotic and abiotic stresses treatments, OsWRKY23 was markedly induced by continuous-dark-induced leaf senescence and infection by rice pathogen Pyricularia oryzae Cav as well as salicylic acid (SA). Further analysis of 35S:OsWRK23 Arabidopsis plants showed that over-expression of OsWRKY23 resulted in enhanced expression of the pathogenesis-related (PR) genes and increased resistance to the bacterial pathogen Pseudomanas syringae. Furthermore, over-expression of OsWRKY23 accelerated leaf senescence in darkness. The senescence-associated marker genes SAG12 and SEN1 were altered in darkness in 35S:WRKY23 Arabidopsis plants. In conclusion, these results suggest that OsWRKY23 is a novel modulator of pathogen responses as well as darkinduced leaf senescence.

A. rigida flowers during the rainy season, when pollinators are scarce, and ombrophily functions to provide reproductive assurance without compromising opportunities for outcrossing.

Reproductive biology of saprophytic plants has been poorly studied. Epipogium roseum (D.Don) Lindl. is a small saprophytic orchid that is widely distributed in tropical and subtropical Asia, Australia and Africa. The floral biology and insect visitation of E. roseum were studied in Xishuangbanna, south Yunnan Province, China. E. roseum possesses an obligate self-pollination system, in which the degenerative rostellum has lost its function as a physical barrier separating the stigma and stamens (pollinia), allowing contact between the stigmatic secretions and the pollinia during bud development. Flowers of E. roseum usually open and successfully attract insect visitors. The Asian honey bee (Apis cerana cerana) was the only visitor observed, and regularly visited flowers of E. roseum for nectar. However, these bees did not carry pollinia away after visiting the flowers due to the absence of a viscid disk in E. roseum; the results of experiments also indicated that the Asian honey bee does not contribute to fruit set in E. roseum. The visiting frequency of Asian honey bees to flowers of E. roseum varied both spatially and temporally. E. roseum does not undergo outcrossing mediated by insects and is adapted to obligate self-pollination. We suggest that this may have evolved because of the uncertainty of pollinator services associated with its saprophytic lifestyle. Our current studies do not support the hypothesis that obligate autogamy is favoured by myco-heterotrophic plants due to resource limitations.