No abstract

Signaling pathways, specifically calcium and calcium-dependent protein kinase (CDPK), have been implicated in the regulation of stress and developmental signals in plants. Here, we reported the isolation and characterization of an orchid, Phalaenopsis amabilis, CDPK gene, PaCDPK1, by using the rapid amplification of cDNA ends (RACE)-PCR technique. The full length cDNA of 2,310 bp contained an open reading frame for PaCDPK1 consisting of 593 amino acid residues. Sequence alignment indicated that PaCDPK1 shared similarities with other plant CDPKs. PaCDPK1 transcripts were expressed strongly in labellum but not in leaves and roots. In addition, the PaCDPK1 gene was transcriptionally activated in response to low temperature, wounding, and pathogen infection. To identify the regulatory role of the PaCDPK1 promoter, a construct containing the PaCDPK1 promoter fused to a beta-glucuronidase (GUS) gene was transferred into Arabidopsis by Agrobacterium-mediated transformation. GUS staining revealed that PaCDPK1/GUS expression was induced by cold, wounding, and pathogen challenge in leaves and stems of transgenic Arabidopsis. These results suggested that this PaCDPK1 gene promoter could be used as an endogenous promoter for biotechnological purposes in orchids.

Heterozygosity-fitness correlations (HFCs) provide insights into the genetic bases of individual fitness variation in natural populations. However, despite decades of study, the biological significance of HFCs is still under debate. In this study, we investigated HFCs in a large population of the sexually dimorphic lizard Takydromus viridipunctatus (Lacertidae). Because of the high prevalence of parasitism from trombiculid mites in this lizard, we expect individual fitness (i.e., survival) to decrease with increasing parasite load. Furthermore, because morphological asymmetry is likely to influence individuals' mobility (i.e., limb asymmetry) and male biting ability during copulation (i.e., head asymmetry) in this species, we also hypothesize that individual fitness should decrease with increasing morphological asymmetry. Although we did not formally test the relationship between morphological asymmetry and fitness in this lizard, we demonstrated that survival decreased with increasing parasite load using a capture-mark-recapture data set. We used a separate sample of 140 lizards to test the correlations between individual heterozygosity (i.e., standardized mean d2 and HL based on 10 microsatellite loci) and the two fitness traits (i.e., parasite load and morphological asymmetry). We also evaluated and excluded the possibility that single-locus effects produced spurious HFCs. Our results suggest male-only, negative correlations between individual heterozygosity and parasite load and between individual heterozygosity and asymmetry, suggesting sex-specific, positive HFCs. Male T. viridipunctatus with higher heterozygosity tend to have lower parasite loads (i.e., higher survival) and lower asymmetry, providing a rare example of HFC in reptiles.

Erwinia chrysanthemi is a devastating bacterial pathogen in Phalaenopsis amabilis and causes soft-rotting disease by secretion of cell wall-degrading enzymes. However, the molecular mechanisms underlying the interaction of P. amabilis with E. chrysanthemi remain elusive. In this study, early molecular events of the plant in response to the pathogen attack were investigated. The alteration in reactive oxygen species accumulation and peroxidase activity occurred at the site of infection. Subsequently, a systematic sequencing of expressed sequence tags (ESTs) using suppression subtractive hybridization (SSH) was performed to obtain the first global picture of the assembly of genes involved in the pathogenesis. The majority of the SSH clones showed a high identity with genes coding for proteins that have known roles in redox homeostasis, responses to pathogens and metabolism. A notable number of the SSH clones were those encoding WRKY, MYB and basic leucine zipper transcription factors, indicating the stimulation of intracellular signal transduction. An orchid gene encoding trans-2-enoyl-CoA reductase (ECR) was the most abundant transcripts in the EST library. ECR is an enzyme catalyzing the very long chain fatty acids (VLCFAs) biosynthesis, and the full-length cDNA of the ECR gene (PaECR1) was obtained. Functional analysis of PaECR1 was conducted by virus-induced gene silencing to knock down the gene expression in P. amabilis. The PaECR1-silenced plants were more susceptible to E. chrysanthemi infection, implying potential roles for VLCFAs in the pathogenesis. In summary, the pathogen-responsive gene expression profiles facilitated a more comprehensive view of the molecular events that underlie this economically important plant-pathogen interaction.