Non-additive gene regulation in a citrus allotetraploid somatic hybrid between C. reticulata Blanco and C. limon (L.) Burm.

Polyploid plants often produce new phenotypes, exceeding the range of variability existing in the diploid gene pool. Several hundred citrus allotetraploid hybrids have been created by somatic hybridization. These genotypes are interesting models to study the immediate effects of allopolyploidization on the regulation of gene expression. Here, we report genome-wide gene expression analysis in fruit pulp of a Citrus interspecific somatic allotetraploid between C. reticulata cv 'Willowleaf mandarin'+C. limon cv 'Eureka lemon', using a Citrus 20K cDNA microarray. Around 4% transcriptome divergence was observed between the two parental species, and 212 and 160 genes were more highly expressed in C. reticulata and C. limon, respectively. Differential expression of certain genes was confirmed by quantitative real-time RT-PCR. A global downregulation of the allotetraploid hybrid transcriptome was observed, as compared with a theoretical mid parent, for the genes displaying interspecific expression divergence between C. reticulata and C. limon. The genes underexpressed in mandarin, as compared with lemon, were also systematically repressed in the allotetraploid. When genes were overexpressed in C. reticulata compared with C. limon, the distribution of allotetraploid gene expression was far more balanced. Cluster analysis on the basis of gene expression clearly indicated the hybrid was much closer to C. reticulata than to C. limon. These results suggest there is a global dominance of the mandarin transcriptome, in consistence with our previous studies on aromatic compounds and proteomics. Interspecific differentiation of gene expression and non-additive gene regulation involved various biological pathways and different cellular components.

Differential gene expression analysis provides new insights into the molecular basis of iron deficiency stress response in the citrus rootstock Poncirus trifoliata (L.) Raf.

Iron chlorosis is one of the major abiotic stresses affecting fruit trees and other crops in calcareous soils and leads to a reduction in growth and yield. Usual remediation strategies consist of amending iron to soil, which is an expensive practice, or using tolerant cultivars, which are difficult to develop when not available. To understand the mechanisms underlying the associated physiopathy better, and thus develop new strategies to overcome the problems resulting from iron deficiency, the differential gene expression induced by iron deficiency in the susceptible citrus rootstock Poncirus trifoliata (L.) Raf. have been examined. The genes identified are putatively involved in cell wall modification, in determining photosynthesis rate and chlorophyll content, and reducing oxidative stress. Additional studies on cell wall morphology, photosynthesis, and chlorophyll content, as well as peroxidase and catalase activities, support their possible functions in the response to iron deficiency in a susceptible genotype, and the results are discussed.

Class prediction of closely related plant varieties using gene expression profiling.

In recent years, class prediction experiments have been largely developed in cancer research with the aim of classifying unknown samples by examining their expression signature. In natural populations, a significant component of gene expression variability is also heritable. Citrus species are an ideal model to accomplish the study of these questions in plants, due to the existence of varieties derived from somatic mutations that are likely to differ from each other by one or a few point mutations but are phenotypically indistinguishable at early vegetative stages. The small genetic variability existing among these varieties makes molecular markers ineffective in distinguishing genotypes within a particular species. Gene expression profiles have been used to predict mandarin clementine varieties (Citrus clementina Hort. ex Tan.) by means of two independent supervised learning algorithms: Support Vector Machines and Prediction Analysis of Microarrays. The results show that transcriptional variation is variety-dependent in citrus, and supervised clustering methods may correctly assign blind samples to varieties when both training and test samples are under the same experimental conditions.

A distinct member of the basic (class I) chitinase gene family in potato is specifically expressed in epidermal cells.

We have isolated cDNA clones encoding class I chitinase (ChtC) from potato leaves which share a high degree of nucleotide and amino acid sequence similarity to other, previously described basic (class I) chitinases (ChtB) from potato. Despite this similarity, characteristic features distinguish ChtC from ChtB, including an extended proline-rich linker region between the hevein and catalytic domains and presence of a potential glycosylation site (NDT) in the deduced protein. These differences are in accordance with the properties of purified chitinase C which is glycosylated and hence has a higher molecular mass in comparison to chitinase B. In contrast to the coding sequences, the 3'-untranslated regions of ChtC and ChtB exhibited a low degree of similarity, which allowed us to generate gene-specific probes to study the genomic organization and expression of both types of gene. Genomic DNA blots suggest that ChtC and ChtB are each encoded by one or two genes per haploid genome. RNA blot analysis showed that in healthy potato plants ChtC mRNA is most abundant in young leaves, the organs which also contain high levels of chitinase C. By contrast, ChtB mRNA abundance is highest in old leaves, which accumulate chitinase B. By in situ RNA hybridization with gene-specific probes we could demonstrate that ChtC mRNA in leaves is restricted to epidermal cells, whereas ChtB mRNA showed no distinct pattern of cell-type-specific localization. Infection of potato leaves with Phytophthora infestans, or treatment with fungal elicitor, ethylene, or wounding resulted in accumulation of both ChtC and ChtB mRNAs; however, for ChtC, in contrast to ChtB, no corresponding accumulation of the encoded protein could be detected, suggesting a post-transcriptional mechanism of regulation. Salicylic acid treatment did not induce accumulation of either mRNA. The possible functional implications of these findings for pathogen defence and developmental processes are discussed.