Natural rubber is synthesized in specialized articulated cells (laticifers) located in the inner liber of Hevea brasiliensis. Upon bark tapping, the laticifer cytoplasm (latex) is expelled due to liber tissue turgor pressure. In mature virgin (untapped) trees, shortterm kinetic studies confirmed that ethylene, the rubber yield stimulant used worldwide, increased latex yield, with a concomitant decrease in latex total solid content, probably through water influx in the laticifers. As the mature laticifers are devoid of plasmodesmata, the rapid water exchanges with surrounding liber cells probably occur via the aquaporin pathway. Two full-length aquaporin cDNAs (HbPIP2;1 and HbTIP1;1, for plasma membrane intrinsic protein and tonoplast intrinsic protein, respectively) were cloned and characterized. The higher efficiency of HbPIP2;1 than HbTIP1;1 in increasing plasmalemma water conductance was verified in Xenopus laevis oocytes. HbPIP2;1 was insensitive to HgCl 2 . In situ hybridization demonstrated that HbPIP2;1 was expressed in all liber tissues in the young stem, including the laticifers. HbPIP2;1 was upregulated in both liber tissues and laticifers, whereas HbTIP1;1 was down-regulated in liber tissues but up-regulated in laticifers in response to bark Ethrel treatment. Ethylene-induced HbPIP2;1 up-regulation was confirmed by western-blot analysis. The promoter sequences of both genes were cloned and found to harbor, among many others, ethylene-responsive and other chemical-responsive (auxin, copper, and sulfur) elements known to increase latex yield. Increase in latex yield in response to ethylene was emphasized to be linked with water circulation between the laticifers and their surrounding tissues as well as with the probable maintenance of liber tissue turgor, which together favor prolongation of latex flow.

MicroRNAs (miRNAs) are a newly discovered class of noncoding endogenous small RNAs involved in plant growth and development as well as response to environmental stresses. miRNAs have been extensively studied in various plant species, however, only few information are available in cassava, which serves as one of the staple food crops, a biofuel crop, animal feed and industrial raw materials. In this study, the 169 potential cassava miRNAs belonging to 34 miRNA families were identified by computational approach. Interestingly, mes-miR319b was represented as the first putative mirtron demonstrated in cassava. A total of 15 miRNA clusters involving 7 miRNA families, and 12 pairs of sense and antisense strand cassava miRNAs belonging to six different miRNA families were discovered. Prediction of potential miRNA target genes revealed their functions involved in various important plant biological processes. The cis-regulatory elements relevant to drought stress and plant hormone response were identified in the promoter regions of those miRNA genes. The results provided a foundation for further investigation of the functional role of known transcription factors in the regulation of cassava miRNAs. The better understandings of the complexity of miRNA-mediated genes network in cassava would unravel cassava complex biology in storage root development and in coping with environmental stresses, thus providing more insights for future exploitation in cassava improvement.

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The data and all previous information on sucrose transport show that HbSUT1A and HbSUT2A are related to the increase in sucrose import into laticifers, required for the stimulation of latex yield by ethylene in virgin trees.

Cassava (Manihot esculenta Crantz) is a root crop that accumulates large quantities of starch, and it is an important source of carbohydrate. Study on gene expressions during storage root development provides important information on storage root formation and starch accumulation as well as unlock new traits for improving of starch yield. cDNA-Amplified Fragment Length Polymorphism (AFLP) was used to compare gene expression profiles in fibrous and storage roots of cassava cultivar Kasetsart 50. Total of 155 differentially expressed transcript-derived fragments with undetectable or low expression in leaves were characterized and classified into 11 groups regarding to their functions. The four major groups were no similarity (20%), hypothetical or unknown proteins (17%), cellular metabolism and biosynthesis (17%) and cellular communication and signaling (14%). Interestingly, sulfite reductase (MeKD82), calcium-dependent protein kinase (CDPK) (MeKD83), ent-kaurene synthase (KS) (MeKD106) and hexose transporter (HT) (MeKD154) showed root-specific expression patterns. This finding is consistent with previously reported genes involved in the initiation of potato tuber. Semi-quantitative reverse transcription polymerase chain reaction of early-developed root samples confirmed that those four genes exhibited significant expression with similar pattern in the storage root initiation and early developmental stages. We proposed that KS and HT may involve in transient induction of CDPK expression, which may play an important role in the signaling pathway of storage root initiation. Sulfite reductase, on the other hand, may involve in storage root development by facilitating sulfur-containing protein biosynthesis or detoxifying the cyanogenic glucoside content through aspartate biosynthesis.

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MicroRNAs (miRNAs) are short RNAs with essential roles in gene regulation in various organisms including higher plants. In contrast to the vast information on miRNAs from many economically important plants, almost nothing has been reported on the identification or analysis of miRNAs from rubber tree (Hevea brasiliensis L.), the most important natural rubber-producing crop. To identify miRNAs and their target genes in rubber tree, high throughput sequencing combined with a computational approach was performed. Four small RNA libraries were constructed for deep sequencing from mature and young leaves of two rubber tree clones, PB 260 and PB 217, which provide high and low latex yield, respectively. 115 miRNAs belonging to 56 known miRNA families were identified, and northern hybridization validated miRNA expression and revealed developmental stage-dependent and clone-specific expression for some miRNAs. We took advantage of the newly released rubber tree genome assembly and predicted 20 novel miRNAs. Further computational analysis uncovered potential targets of the known and novel miRNAs. Predicted target genes included not only transcription factors but also genes involved in various biological processes including stress responses, primary and secondary metabolism, and signal transduction. In particular, genes with roles in rubber biosynthesis are predicted targets of miRNAs. This study provides a basic catalog of miRNAs and their targets in rubber tree to facilitate future improvement and exploitation of rubber tree.

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