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[EN] Laticifers are specialized plant cells capable of indefinite elongation that ramify extensively and are responsible for latex biosynthesis and accumulation. However, the mechanisms underlying laticifer cell differentiation, growth and production of latex remain largely unknown. In a search for mutants showing enhanced accumulation of latex we identified two LOT OF LATEX (LOL) loci in Euphorbia lathyris. lol2 and lol5 mutants show enhanced production of latex contained within laticifer cells. The recessive lol2 mutant carries increased biosynthesis of the plant hormone jasmonoyl-isoleucine (JA-Ile) and therefore establishes a genetic link between jasmonic acid (JA) signaling and latex production in laticifers. Instead, heightened production of latex in lol5 plants obeys to enhanced proliferation of laticifer cells. Phylogenetic analysis of laticifer-expressed genes in E. lathyris and in two other latex-bearing species, Euphorbia corallioides and Euphorbia palustris, allowed the identification of canonical JA responsive elements present in the gene promoter regions of laticifer marker genes. Moreover, we identified that the hormone JA functions not as a morphogen for laticifer differentiation but as a trigger for the fill out of laticifers with latex and the associated triterpenoids. The identification of LOL loci represents a further step towards the understanding of mechanisms controlling latex production in laticifer cells., This work was supported by Spanish MINECO (BFU2015 -68199 -R to P.V.) and Generalitat Valenciana (Prometeo 2014/024 to P.V.).

The secondary laticifer in the secondary phloem of rubber tree are a specific tissue differentiating from vascular cambia. The number of the secondary laticifers is closely related to the rubber productivity of Hevea. Factors involved in the mechanical wounding-induced laticifer differentiation were analyzed by using paraffin section, gas chromatography-mass spectrometry (GC-MS), and Northern-blot techniques. Dehydration of the wounded bark tissues triggered a burst of hydrogen peroxide, abscisic acid, and jasmonates and up-regulated the expression of HbAOSa, which was associated with the secondary laticifer differentiation strictly limited to the wounded area. Application of exogenous hydrogen peroxide, methyl jasmonate, and polyethylene glycol 6000 (PEG6000) could induce the secondary laticifer differentiation, respectively. Moreover, 6-Benzylaminopurine, a synthetic cytokinin, enhanced the methyl jasmonate-induced secondary laticifer differentiation. However, the dehydration-induced secondary laticifer differentiation was inhibited by exogenous abscisic acid. Diphenyleneiodonium chloride (DPI), a specific inhibitor of NADPH oxidase, was effective in inhibiting the accumulation of hydrogen peroxide as well as of jasmonates upon dehydration. It blocked the dehydration-induced but not the methyl jasmonate-induced secondary laticifer differentiation. The results suggested a stress signal pathway mediating the wound-induced secondary laticifer differentiation in rubber tree.

The latex dilution reaction during the tapping flow course has been well documented and associated with the facilitation of tapping latex flow. However, its underlying mechanism has not experimentally examined. The latex total solid content, osmotic potential and phloem turgor pressure change during the tapping flow course were simultaneously measured to investigate the cause of water movement during the tapping flow course. It was found that there are three different stages for the laticifer water equilibrium during the tapping flow course. The tapping-induced rapid turgor pressure drop is the cause of the first stage water influx into laticifers, while osmoregulation prevails during water exchange in the second and third stages of tapping flow. Meanwhile, aquaporin expressions were, for the first time, investigated during the tapping flow course. The rapid transcript up-regulation of HbPIP1, HbPIP2;1 and HbPIP2;3 contributes to the latex dilution reaction. However, their activity gating cannot be ruled out. Ethrel stimulation can significantly dilute the corresponding latex fractions during the tapping flow course due to its up-regulations of HbPIP1, HbPIP2;1 and HbPIP2;3. Nevertheless, the latex dilution reaction pattern for the Ethrel treated trees did not change, except for a lower degree of dilution compared with the un-treated trees. All these results suggest that both phloem turgor pressure and aquaporins are involved in the latex dilution reaction during the tapping flow course.

MYB transcription factors hold vital roles in the regulation of plant secondary metabolic pathways. Laticifers in rubber trees (Hevea brasiliensis) are of primary importance in natural rubber production because natural rubber is formed and stored within these structures. To understand the role of MYB transcription factors in the specialized cells, we identified 44 MYB genes (named HblMYB1 to HblMYB44) by using our previously obtained transcriptome database of rubber tree laticifer cells and the public rubber tree genome database. Expression profiles showed that five MYB genes were highly expressed in the laticifers. HblMYB19 and HblMYB44 were selected for further study. HblMYB19 and HblMYB44 bound the promoters of HbFDPS1, HbSRPP, and HRT1 in yeast. Furthermore, the transient overexpression of HblMYB19 and HblMYB44 in tobacco plants significantly increased the activity of the promoters of HbFDPS1, HbSRPP, and HRT1. Basing on this information, we proposed that HblMYB19 and HblMYB44 are the regulators of HbFDPS1, HbSRPP, and HRT1, which are involved in the biosynthesis pathway of natural rubber.

Autophagy is involved in cytoplasmic degradation through directly engulfing cytosol and organelles by autophagosomes and then fusing with lysosome-like vesicles during the development of nonarticulated laticifers in Euphorbia kansui Liou. Autophagy has been reported to play an important role in a wide range of eukaryotic organisms during responses to various abiotic and biotic stresses. However, until recently, the functions of autophagy in normal plant differentiation and development were still in their infancy. Nonarticulated laticifers, a type of secretory tissue in plants, undergo the degradation of cytosol and organelles during their development. However, little evidence of autophagy in laticifer differentiation has been provided. In the present study, using anti-ATG8 antibody-Alexa Fluor 488, Lyso-Tracker Red (LTR) and monodansylcadaverine (MDC) as markers for detecting autophagosomes, as well as autophagy-related structures, we observed that the green fluorescence of ATG8a largely colocalized with the red fluorescence of LTR and purple fluorescence of MDC and the quantity of autophagosomes experienced a trend from less to more to less during laticifer development. Additionally, we described the autophagy process during the development of nonarticulated laticifers in Euphorbia kansui Liou at the ultrastructural level in detail. In addition, further immunogold TEM studies also verified the presence of autophagosomes, autolysosomes and lysosome-like structures in laticifers. Taken together, these results suggest that autophagy contributes to the development of the nonarticulated laticifers in E. kansui Liou and that autophagosomes fuse with lysosome-like structures for degradation. These results will lay an important foundation for further studies on laticifer regulation.

Laticifers are highly specialized cells that synthesize and store natural rubber. Rubber trees (Hevea brasiliensis Muell. Arg.) contain both primary and secondary laticifers. Morphological and functional differences between the two types of laticifers are largely unknown, but such information is important for breeding and cultivation practices. Morphological comparison using paraffin sections revealed only distribution differences: the primary laticifers were distributed randomly, while the secondary laticifers were distributed in concentric rings. Using isolated laticifer networks, the primary laticifers were shown to develop via intrusive “budding” and formed necklace-like morphology, while the secondary laticifers developed straight and smooth cell walls. Comparative transcriptome analysis indicated that genes involved in cell wall modification, such as pectin esterase, lignin metabolic enzymes, and expansins, were highly up-regulated in the primary laticifers and correspond to its necklace-like morphology. Genes involved in defense against biotic stresses and rubber biosynthesis were highly up-regulated in the primary laticifers, whereas genes involved in abiotic stresses and dormancy were up-regulated in the secondary laticifers, suggesting that the primary laticifers are more adequately prepared to defend against biotic stresses, while the secondary laticifers are more adequately prepared to defend against abiotic stresses. Therefore, the two types of laticifers are morphologically and functionally distinct.

Dehydrin is a type of late embryogenesis abundant (LEA) protein. The dehydrin genes, HbDHN1 and HbDHN2, in Hevea brasiliensis were previously found to be induced at the wounding site of epicormic shoots, with local tissue dehydration identified as the key signal for laticifer differentiation. However, the exact role of the HbDHNs remains unknown. In this study, HbDHN1 and HbDHN2 expression was examined under multiple abiotic stresses; namely, cold, salt, drought, wounding, abscisic acid (ABA), ethylene (ET), and jasmonic acid (JA) treatment. Although, both HbDHNs were defined as SK2-type dehydrin, they showed different cellular localizations. Overexpression of the HbDHNs in Arabidopsis thaliana further revealed a significant increase in tolerance to salt, drought and osmotic stresses. Increased accumulation of proline and a reduction in electrolyte leakage were also observed under salt and drought stress, and a higher water content was indicated under osmotic stress. The transgenic plants also showed higher activity levels of ascorbate peroxidase (APX), superoxide dismutase (SOD) and catalase, and accumulated less hydrogen peroxide (H2O2) and superoxide (O2−). Given that reactive oxygen species (ROS) are thought to be a key signal for laticifer differentiation, these findings suggest that HbDHNs act as ROS scavengers, directly or indirectly affecting laticifer differentiation. Both HbDHNs therefore influence physiological processes, improving plant tolerance to multiple abiotic stresses.

Recent studies have shown the laticifers of Apocynaceae, previously classified as nonarticulated, indeed are articulated, anastomosing laticifers whose transverse walls dissolve rapidly and entirely, although doubts about their growth mode still persist. To better understand the mode of laticifer growth and differentiation in this family, we studied its development in Allamanda blanchetii using anatomical and ultrastructural analyses. Our results showed that laticifers are formed by a row of cells that join each other through dissolution of the transverse walls from the center to the periphery. The laticifers originate from ground meristem and procambium; the laticifers in the different tissues connect through lateral fusion, generating a laticifer network. The laticifers occur in the cortex, pith, and vascular system of the shoot, mesophyll, and vascular bundles of the leaves. There is no apical growth, and all the organelles observed in the apices of the laticifers play a role in the production of latex or in the dissolution of the terminal walls between the cells that compose the laticifer. The latex is composed of many metabolites produced mainly in the endoplasmic reticulum and plastids. Mitochondria are abundant, and dictyosomes are scarce. The vacuome is prominent from the start of laticifer differentiation, and many small vacuoles and vesicles transport the secretion from cytosol into the central vacuole, where an emulsion of substances is stored. The articulated, anastomosing laticifers of A. blanchetii have no subcellular mechanism for production of a cell wall in a polarized manner or dissolution of middle lamella of the cells which surround the laticifer tip, proving that there is no intrusive growth in this secretory structure.

[EN] In the latex-bearing plants, the laticiferous system is the tubing structure that contains the latex and is constituted of living cells (laticifers). While laticifers are present only in a small percentage of the flowering plant species, they represent a type of specialized tissue within the plant where a myriad of metabolites are synthesized, some of them of considerable commercial importance. In this mini-review we synopsize the present knowledge about laticifer cells and discuss about their particular features as well as some evolutionary and ecophysiological cues and the potential exploitation of the knowledge generated around this peculiar type of plant cell. We illustrate some of these questions with the experience in Euphorbia lathyris laticifers and latex., This work was supported by the Spanish MINECO (BFU2015-68199-R to P.V.) and Generalitat Valenciana (Prometeo 2014/024 to P.V.)

Studies investigating the resistance-susceptibility of crop insects to proteins found in latex fluids have been reported. However, latex-bearing plants also host insects. In this study, the gut proteolytic system of Pseudosphinx tetrio, which feeds on Plumeria rubra leaves, was characterized and further challenged against the latex proteolytic system of its own host plant and those of other latex-bearing plants. The gut proteolytic system of Danaus plexippus (monarch) and the latex proteolytic system of its host plant (Calotropis procera) were also studied. The latex proteins underwent extensive hydrolysis when mixed with the corresponding gut homogenates of the hosted insects. The gut homogenates partially digested the latex proteins of foreign plants. The fifth instar of D. plexippus that were fed diets containing foreign latex developed as well as those individuals who were fed diets containing latex proteins from their host plant. In vitro assays detected serine and cysteine peptidase inhibitors in both the gut homogenates and the latex fluids. Curiously, the peptidase inhibitors of caterpillars did not inhibit the latex peptidases of their host plants. However, the peptidase inhibitors of laticifer origin inhibited the proteolysis of gut homogenates. In vivo analyses of the peritrophic membrane proteins of D. plexippus demonstrate resistance against latex peptidases. Only discrete changes were observed when the peritrophic membrane was directly treated with purified latex peptidases in vitro. This study concludes that peptidase inhibitors are involved in the defensive systems of both caterpillars and their host plants. Although latex peptidase inhibitors inhibit gut peptidases (in vitro), the ability of gut peptidases to digest latex proteins (in vivo) regardless of their origin seems to be important in governing the resistance-susceptibility of caterpillars.

This work aims to characterize the anatomy of leaves of wild Manihot species (Euphorbiaceae) and identify anatomical characteristics of taxonomic importance. Samples of M. violacea var. cecropiaefolia, M. violacea var. divergens, M. violacea ssp. jacobinensis (Lencois) and M. violacea were collected in cerrado (savanna), and samples of M. violacea ssp. jacobinensis and M. violacea ssp. jacobinensis (Mucuge) were collected in campo rupestre (rocky land) areas in northeastern Brazil. The leaf anatomy of these species was analyzed using optical and scanning electron microscopy in order to obtain useful characters for the taxonomy of the genus. Information acquired confirms the records from the literature for the genus. The taxonomic characters that were considered to be the most important are as follows: number of vascular bundles and petiole outline; distribution of secretory idioblasts; and presence of papillae and midrib outline. All of these species have secretory idioblasts, crystals, laticifers and isobilateral mesophyll. Most of the species present a cylindrical petiole outline. The number of vascular bundles on the petiole varies between five and ten. The similarity analysis reinforces M. jacobinensis as a distinct species.

Cytosolic acetyl-CoA is involved in the synthesis of a variety of compounds, including waxes, sterols and rubber, and is generated by the ATP citrate lyase (ACL). Plants over-expressing ACL were generated in an effort to understand the contribution of ACL activity to the carbon flux of acetyl-CoA to metabolic pathways occurring in the cytosol. Transgenic Arabidopsis plants synthesizing the polyester polyhydroxybutyrate (PHB) from cytosolic acetyl-CoA have reduced growth and wax content, consistent with a reduction in the availability of cytosolic acetyl-CoA to endogenous pathways. Increasing the ACL activity via the over-expression of the ACLA and ACLB subunits reversed the phenotypes associated with PHB synthesis while maintaining polymer synthesis. PHB production by itself was associated with an increase in ACL activity that occurred in the absence of changes in steady-state mRNA or protein level, indicating a post-translational regulation of ACL activity in response to sink strength. Over-expression of ACL in Arabidopsis was associated with a 30% increase in wax on stems, while over-expression of a chimeric homomeric ACL in the laticifer of roots of dandelion led to a four- and two-fold increase in rubber and triterpene content, respectively. Synthesis of PHB and over-expression of ACL also changed the amount of the cutin monomer octadecadien-1,18-dioic acid, revealing an unsuspected link between cytosolic acetyl-CoA and cutin biosynthesis. Together, these results reveal the complexity of ACL regulation and its central role in influencing the carbon flux to metabolic pathways using cytosolic acetyl-CoA, including wax and polyisoprenoids.

Two Apocynaceae species from savanna (Cerrado) area of Sao Paulo State, Brazil were studied. In both Mandevilla species examined, the laticifer systems are of non-articulated branched type, characteristic of the family. In vegetative organs reported was the occurrence of a primary laticifer system of which the cells were differentiated in the embryo. However, additional laticifer cells were always produced during the growth of the shoot apex. A secondary laticiferous system produced by the cells of vascular cambium was identified in the tuberous root and in the stem. A proposed discussion on this apparently unique record in Apocynaceae was arisen to evaluate the main distinction between articulated and non-articulated laticifers.

About 90% of natural rubber is obtained from tapping of the rubber tree (Hevea brasiliensis) for latex, which is circulated in the laticifers. Nowadays, the world supply of natural rubber is not sufficient for global demand, thus the increased latex yield was significantly underlined. In addition, the demand for wood (as a xylem part) from rubber tree was also increased as renewable resource for various applications. Laticifers are found in the secondary phloem containing tissue of rubber tree trunk-inner soft bark. The number of laticifers varies in consistent with latex yield and in responses to jasmonic acid level. This present study was committed to comparative transcriptome analysis in laticifers containing mature phloem, mature xylem and newly developed stem tissues of high latex-yield clone (RRIT251, with more laticifers) and high wood-yield clone (RRIT402, with less laticifers) of rubber tree to classify the genes and pathways involved with phloem (with laticifers) and xylem cell differentiation. There were 49, 54, 46 and 50 of vascular development-related genes in primary and secondary tissues of phloem and xylem, respectively. Differentially expressed genes in jasmonic acid signaling pathway was established with their highest expression in phloem tissue with laticifer cells of RRIT251, while genes in auxin signaling and secondary cell wall biosynthetic pathways were up-regulated in xylem tissue of RRIT402 for high wood yield. Promoter analysis of candidate-differentially expressed genes suggested the related pathway and putative regulatory elements for gene regulation. This genome-wide exploration of vascular development-related genes unraveled a largely unknown gap of this special vascular development containing laticifers in rubber tree.

Alkaloids attract great attention due to their valuable therapeutic properties. Stepharine, an aporphine alkaloid of Stephania glabra plants, exhibits anti-aging, anti-hypertensive, and anti-viral effects. The distribution of aporphine alkaloids in cell cultures, as well as whole plants is unknown, which hampers the development of bioengineering strategies toward enhancing their production. The spatial distribution of stepharine in cell culture models, plantlets, and mature micropropagated plants was investigated at the cellular and organ levels. Stepharine biosynthesis was found to be highly spatially and temporally regulated during plant development. We proposed that self-intoxication is the most likely reason for the failure of the induction of alkaloid biosynthesis in cell cultures. During somatic embryo development, the toxic load of alkaloids inside the cells increased. Only specialized cell sites such as vascular tissues with companion cells (VT cells), laticifers, and parenchymal cells with inclusions (PI cells) can tolerate the accumulation of alkaloids, and thus circumvent this restriction. S. glabra plants have adapted to toxic pressure by forming an additional transport secretory (laticifer) system and depository PI cells. Postembryonic growth restricts specialized cell site formation during organ development. Future bioengineering strategies should include cultures enriched in the specific cells identified in this study.

Transcription factors of AP2/ERF superfamily are generally involved in defense responses of plants to biotic and abiotic stresses. Although, defense proteins are present in abundance in laticifers of rubber tree, little is known about their transcriptional regulation. In this study, a full length cDNA, referred to as HbEREBP2 was characterized by means of bioinformatic analysis and quantitative real-time RT-PCR. The HbEREBP2 was 786-bp in length and contained a 480-bp open reading frame (ORF) encoding a protein of 159 amino acid residues. Bioinformatic analysis showed that the deduced amino acid sequence of HbEREBP2 had a specific domain of AP2 superfamily and shared relative high identity with members of CBF/DREB subfamily from different plant species. Quantitative real-time RT-PCR revealed that methyl jasmonate was more effective than ethylene and rapidly than mechanical wounding on upregulating HbEREBP2 expression. The results suggest that HbEREBP2 may be involved in the regulation of jasmonate-mediated defense responses in laticifers of rubber tree. Key words: Hevea brasiliensis, Laticifer, defense proteins, AP2/ERF transcription factor, Methyl jasmonates, Ethephon, mechanical wounding.

Sapium haematospermum Müll.Arg. presents two different laticifer systems, distinguishable on the basis of their structure and ontogeny. Although they are both articulated anastomosing laticifers, produced from procambium and (or) ground meristem, they vary in diameter and in the presence or absence of starch. One of them, originating near the promeristem and leaf primordia, is composed of straight, wide laticifers with rod-shaped starch grains, whereas the other one is made up of narrow and sinuous laticifers that arise in expanding organs. The two systems are observed in almost all tissues of the leaf and in the cortical parenchyma and vascular tissue of the stem. Our results show the presence of two different laticifer systems for the first time in Sapium and contradict former reports that described nonarticulated laticifers in this genus. The occurrence of two different laticifer types in the same genus, and indeed the same plant, is extremely rare. The milky latex of the two laticifer systems of S. haematospermum, which consists of terpenoids, fatty acids, phenolic compounds, proteins, and polysaccharides (including mucilage), plays an efficient role in sealing wounds, blocking microorganisms, and avoiding herbivory.

http://dx.doi.org/10.5902/198050989294O presente estudo teve por objetivos descrever anatomicamente o lenho de Phyllanthus sellowianus Müll. Arg. (Phyllanthaceae), bem como investigar caracteres anatômicos associados à flexibilidade de seus caules, característica desejável para obras de bioengenharia em taludes fluviais. Pertencente ao grupo das reófilas, e com até 3 metros de altura, a espécie ocorre naturalmente no sul do Brasil, Uruguai e Argentina. As exsicatas e as amostras de madeira, provenientes de um exemplar localizado no município de Santa Maria-RS (Brasil), foram acondicionadas no Herbário e Xiloteca da Universidade Federal de Santa Maria, sob os números HDCF 5587 e 5588. Para a confecção das lâminas histológicas de madeira, seguiu-se a técnica padrão. Para a maceração, utilizou-se o método de Jeffrey. A descrição anatômica da madeira seguiu as recomendações do IAWA Committee (1989). A análise microscópica do lenho revelou a presença de poros com pequeno diâmetro (< 60 μm), dispostos em múltiplos radiais de 2 – 4 unidades, e de placas de perfuração simples, oblíquas; parênquima axial ausente, raios heterogêneos, com 1 – 3 células de largura, e fibras gelatinosas. A ausência de parênquima axial e de tubos laticíferos é compatível com o grupo das Phyllanthaceae. A presença de células perfuradas de raio constitui-se, possivelmente, no principal elemento anatômico para a diferenciação de P. sellowianus em relação às demais espécies do gênero. A presença abundante de fibras gelatinosas é seguramente uma das principais características anatômicas relacionadas à elevada flexibilidade de seu caule.

An osmotin (CpOsm) from the latex of Calotropis procera has been crystallized in both tetragonal and trigonal forms suitable for structure determination. Crystallographic studies of CpOsm are of great interest because limited information is available concerning the structure of latex proteins and CpOsm has previously been shown to interact with the spore membranes of some plant pathogenic fungi, thus impairing spore germination and hyphal growth. CpOsm crystals were grown using 0.1 M HEPES buffer pH 7.5, 26% PEG 4000, 0.2 M ammonium sulfate (space group P4(3)) or using 0.1 M HEPES buffer pH 7.5, 35% MPD, 0.7 M ammonium sulfate (space group P3(1)12). X-ray diffraction data were collected to 2.17 Å (P4(3)) and 1.80 Å (P3(1)12) resolution and molecular-replacement analyses produced initial phases for both crystal forms.

A fundamental understanding of mango flowering in the tropics is essential to efficiently utilize cropping management systems which could extend both the flowering and crop production seasons. However, the information and appreciation of the floral biology of this popular fruit species is still lacking. Therefore, the objective of this work was to observe the morphology of Chok Anan mango flowers using scanning electron microscope. The Chok Anan mango flower is monoecius where both the male and hermaphrodite flowers exist in the same panicle. The fruit develops from the hermaphrodite flower while the male flower contributes the pollen when pollination occurred. The Chok Anan mango flowers contained five sepals and five petals arranged in a whorl. There were not much differences in the structure of sepals and petals between the male and hermaphrodite flowers. The structure of the petals is not flat but billowy. Both sepals and petals consist of ground parenchyma tissue with laticifer and idioblast cells, starch granules and vascular bundle tissues. Trichomes are present in both sepals and petals. The male flower has the same structure as the hermaphrodite flower except that it does not have carpel. Further study is needed to understand the peculiarities of the floral morphology to allow us to predict mango production. Key words: Pollen, sepal, petal, hermaphrodite, floral biology.

The secondary laticifer in rubber tree (Hevea brasiliensis Muell. Arg.) is a specific tissue within the secondary phloem. This tissue differentiates from the vascular cambia, and its function is natural rubber biosynthesis and storage. Given that jasmonates play a pivotal role in secondary laticifer differentiation, we established an experimental system with jasmonate (JA) mimic coronatine (COR) for studying the secondary laticifer differentiation: in this system, differentiation occurs within five days of the treatment of epicormic shoots with COR. In the present study, the experimental system was used to perform transcriptome sequencing and gene expression analysis. A total of 67,873 unigenes were assembled, and 50,548 unigenes were mapped at least in one public database. Of these being annotated unigenes, 15,780 unigenes were differentially expressed early after COR treatment, and 19,824 unigenes were differentially expressed late after COR treatment. At the early stage, 8,646 unigenes were up-regulated, while 7,134 unigenes were down-regulated. At the late stage, the numbers of up- and down-regulated unigenes were 7,711 and 12,113, respectively. The annotation data and gene expression analysis of the differentially expressed unigenes suggest that JA-mediated signalling, Ca2+ signal transduction and the CLAVATA-MAPK-WOX signalling pathway may be involved in regulating secondary laticifer differentiation in rubber trees.

Latex exploitation-caused latex flow is effective in enhancing latex regeneration in laticifer cells of rubber tree. It should be suitable for screening appropriate reference gene for analysis of the expression of latex regeneration-related genes by quantitative real-time PCR (qRT-PCR). In the present study, the expression stability of 23 candidate reference genes was evaluated on the basis of latex flow by using geNorm and NormFinder algorithms. Ubiquitin-protein ligase 2a (UBC2a) and ubiquitin-protein ligase 2b (UBC2b) were the two most stable genes among the selected candidate references in rubber tree clones with differential duration of latex flow. The two genes were also high-ranked in previous reference gene screening across different tissues and experimental conditions. By contrast, the transcripts of latex regeneration-related genes fluctuated significantly during latex flow. The results suggest that screening reference gene during latex flow should be an efficient and effective clue for selection of reference genes in qRT-PCR.

The movement of water is a matter of great importance to all plant species. This is particularly true for the rubber tree ( Hevea brasiliensis Muell. Arg.) which is tapped for the laticifer cytoplasm in the form of aqueous latex. Due to the lack of plasmodesmata, the laticifer water balance is mediated largely by aquaporins (AQPs), a class of channel-forming integral membrane proteins that transport water and other small solutes. Our studies reported the characterization of 51 full-length AQP genes from the rubber tree genome, and further assigned them to five subfamilies, i.e. plasma membrane intrinsic proteins (PIPs: 15), tonoplast intrinsic proteins (TIPs: 17), NOD26-like intrinsic proteins (NIPs: 9), small basic intrinsic proteins (SIPs: 4) and X intrinsic proteins (XIPs: 6). Although the classification of subfamily/subgroup is the same as in Ricinus communis , genome-wide comparative analysis revealed the PIP and TIP subfamilies in rubber tree are highly expanded, corresponding to their high water permeability and the particular importance of water balance in a big tree and a highly differentiated laticifer tissue. Functional prediction based on the analysis of the aromatic/arginine (ar/R) selectivity filter, Froger’s positions and specificity-determining positions (SDPs) further suggested the potentially key role of HbPIPs and HbTIPs in the laticifer water balance. Moreover, deep sequencing of the latex transcriptome and qRT-PCR analysis supported a crucial role of several PIP subfamily members in the laticifer water balance under both normal and ethephon-stimulated conditions.

Made available in DSpace on 2018-12-11T17:28:18Z (GMT). No. of bitstreams: 0 Previous issue date: 2016-06-01 Premise of research. In Apocynaceae, nonarticulated laticifers have been recorded in most species studied. Interpretation of the mode of development of laticifers, whether articulated or nonarticulated, is controversial, possibly because of the rapid changes that occur in the early differentiation stages of the structures. Here, we describe laticifers in the embryo, seedling, and plant of Tabernaemontana catharinensis A.DC. (Apocynaceae), aiming to understand the structure and developmental mechanism of the laticifer system. Methodology. We prepared samples of mature embryos, 20-d-old seedlings, and 80-d-old plants of T. catharinensis according to conventional light microscopy techniques for anatomical and histochemical analysis. Pivotal results. Articulated anastomosing laticifers with intrusive growth, producing proteins, lipids, and terpenes, are present from mature embryos. Alkaloids are present in the laticifer protoplast of the older portions of the stem. Laticifers of the primary system originate from the ground meristem and procambium, and those of the secondary system originate from the vascular cambium toward the secondary phloem. In the embryo, laticifers are found in the ground meristem and procambium but do not occur in the promeristem; the nodal region exhibits lateral projections between laticifers and ground meristem cells. In seedlings and plants, laticifers are immersed in the parenchyma tissue and associated with the primary and secondary phloem of the root and shoot systems. The anatomical evidence suggests the incorporation of meristematic and parenchyma cells within the laticifer system. Conclusions. The laticifer system of T. catharinensis has a complex structure and developmental mechanism involving protoplast fusion, the addition of cells, and intrusive growth. We recorded, for the first time, the occurrence of intrusive growth associated with articulated anastomosing laticifers in a member of Apocynaceae. Our findings highlight the importance of detailed anatomical analysis to properly classify laticifers and to access their origin and development. Departamento de Botânica Instituto de Biociências Univ Estadual Paulista (UNESP) Departamento de Botânica Instituto de Biociências UNESP Departamento de Botânica Instituto de Biociências Univ Estadual Paulista (UNESP) Departamento de Botânica Instituto de Biociências UNESP

Sticky disease, which is caused by Papaya meleira virus (PMeV), is a significant papaya disease in Brazil and Mexico, where it has caused severe economic losses, and it seems to have spread to Central and South America. Studies assessing the pathogen-host interaction at the nano-histological level are needed to better understand the mechanisms that underlie natural resistance. In this study, the topography and mechanical properties of the leaf midribs and latex of healthy and PMeV-infected papaya plants were observed by atomic force microscopy and scanning electron microscopy. Healthy plants displayed a smooth surface with practically no roughness of the leaf midribs and the latex and a higher adhesion force than infected plants. PMeV promotes changes in the leaf midribs and latex, making them more fragile and susceptible to breakage. These changes, which are associated with increased water uptake and internal pressure in laticifers, causes cell disruption that leads to spontaneous exudation of the latex and facilitates the spread of PMeV to other laticifers. These results provide new insights into the papaya-PMeV interaction that could be helpful for controlling papaya sticky disease.

Vinblastine and vincristine are secondary metabolites from Madagascar periwinkles that have a very high economic value as chemotherapy drugs. These compounds are naturally produced in a very low quantity in planta . One promising alternative method for vinblastine and vincristine production is to use a treatment that can trigger plant stress response in vitro . This study has been done to evaluate the effect of drought stress using polyethylene glycol (PEG) on vinblastine a nd vincristine production in the C. roseus callus culture , which were grown on medium Zenk supplemented with plant growth regulators (PGR) 1 μM NAA + 10 μM Kinetin to induce laticifer and idioblast differentiation. 13 -week-old callus culture s were then treated with 0% , 6% , 9%, a nd 12% (w/v) PEG4000 each for 0 , 24 , 48, and 72 hours . Biochemical analysis was performed using HPLC to determine the levels of vinblastine and vincristine, while the presence of differentiated cells (idioblasts and laticifers) was determined using a histochemical method. Protein profiles of the culture were determined by SDS-Page. The results showed that drought treatment with PEG4000 , until the concentration was 12% (w/v), did not significantly affect the production of vinblastine and vincristine, but might affect terpenoid production. Histochemical analysis confirmed the presence of idioblasts, non-elongated laticifers, and laticifers that were producing and accumulating terpenoids highest in t he 12% PEG treatment. PEG treatments also did not change the protein profile of callus.

Catharanthus roseus (L.) G. Don is a medicinal plant well known for producing antitumor drugs such as vinblastine and vincristine, which are classified as terpenoid indole alkaloids (TIAs). The TIA metabolic pathway in C. roseus has been extensively studied. However, the localization of TIA intermediates at the cellular level has not been demonstrated directly. In the present study, the metabolic pathway of TIA in C. roseus was studied with two forefront metabolomic techniques, that is, Imaging mass spectrometry (MS) and live Single-cell MS, to elucidate cell-specific TIA localization in the stem tissue. Imaging MS indicated that most TIAs localize in the idioblast and laticifer cells, which emit blue fluorescence under UV excitation. Single-cell MS was applied to four different kinds of cells [idioblast (specialized parenchyma cell), laticifer, parenchyma, and epidermal cells] in the stem longitudinal section. Principal component analysis of Imaging MS and Single-cell MS spectra of these cells showed that similar alkaloids accumulate in both idioblast cell and laticifer cell. From MS/MS analysis of Single-cell MS spectra, catharanthine, ajmalicine, and strictosidine were found in both cell types in C. roseus stem tissue, where serpentine was also accumulated. Based on these data, we discuss the significance of TIA synthesis and accumulation in the idioblast and laticifer cells of C. roseus stem tissue.

Rubber tree is a major commercial source of natural rubber. Latex coagulation is delayed by thiols, which belong to the important type of antioxidants in laticifer submembrane, and is composed of glutathione (GSH), cysteine, and methionine. The rate-limiting enzyme,γ-ECS, plays an important role in regulating the biosynthesis of glutathione under any environment conditions. To understand the relation betweenγ-ECS and thiols and to correlate latex flow with one-time tapping and continuous tapping, we cloned and derived the full length of oneγ-ECS from rubber tree latex (Hbγ-ECS1). According to qPCR analysis, the expression levels ofHbγ-ECS1were induced by tapping and Ethrel stimulation, and the expression was related to thiols content in the latex. Continuous tapping induced injury, and the expression ofHbγECS1increased with routine tapping and Ethrel-stimulation tapping (more intensive tapping). According to expression in long-term flowing latex, the gene was related to the duration of latex flow.HbγECS1was expressed inE. coliRosetta using pET-sumo as an expression vector and the recombinant enzyme was purified; then we achieved 0.827 U/mg specific activity and about 66 kDa molecular weight. The present study can help us understand the complex role ofHbγ-ECSin thiols biosynthesis, which is influenced by tapping.

Ethylene is commonly used as a latex stimulant of Hevea brasiliensis by application of ethephon (chloro-2-ethylphosphonic acid); however, the molecular mechanism by which ethylene increases latex production is not clear. To better understand the effects of ethylene stimulation on the laticiferous cells of rubber trees, a latex expressed sequence tag (EST)-based complementary DNA microarray containing 2,973 unique genes (probes) was first developed and used to analyze the gene expression changes in the latex of the mature virgin rubber trees after ethephon treatment at three different time-points: 8, 24 and 48 h. Transcript levels of 163 genes were significantly altered with fold-change values ≥ 2 or ≤ -2 (q-value < 0.05) in ethephon-treated rubber trees compared with control trees. Of the 163 genes, 92 were up-regulated and 71 down-regulated. The microarray results were further confirmed using real-time quantitative reverse transcript-PCR for 20 selected genes. The 163 ethylene-responsive genes were involved in several biological processes including organic substance metabolism, cellular metabolism, primary metabolism, biosynthetic process, cellular response to stimulus and stress. The presented data suggest that the laticifer water circulation, production and scavenging of reactive oxygen species, sugar metabolism, and assembly and depolymerization of the latex actin cytoskeleton might play important roles in ethylene-induced increase of latex production. The results may provide useful insights into understanding the molecular mechanism underlying the effect of ethylene on latex metabolism of H. brasiliensis.

X-Intrinsic Proteins (XIP) were recently identified in a narrow range of plants as a full clade within the aquaporins. These channels reportedly facilitate the transport of a wide range of hydrophobic solutes. The functional roles of XIP in planta remain poorly identified. In this study, we found three XIP genes (HbXIP1;1, HbXIP2;1 and HbXIP3;1) in the Hevea brasiliensis genome. Comprehensive bioinformatics, biochemical and structural analyses were used to acquire a better understanding of this AQP subfamily. Phylogenetic analysis revealed that HbXIPs clustered into two major groups, each distributed in a specific lineage of the order Malpighiales. Tissue-specific expression profiles showed that only HbXIP2;1 was expressed in all the vegetative tissues tested (leaves, stem, bark, xylem and latex), suggesting that HbXIP2;1 could take part in a wide range of cellular processes. This is particularly relevant to the rubber-producing laticiferous system, where this isoform was found to be up-regulated during tapping and ethylene treatments. Furthermore, the XIP transcriptional pattern is significantly correlated to latex production level. Structural comparison with SoPIP2;1 from Spinacia oleracea species provides new insights into the possible role of structural checkpoints by which HbXIP2;1 ensures glycerol transfer across the membrane. From these results, we discuss the physiological involvement of glycerol and HbXIP2;1 in water homeostasis and carbon stream of challenged laticifers. The characterization of HbXIP2;1 during rubber tree tapping lends new insights into molecular and physiological response processes of laticifer metabolism in the context of latex exploitation.

Hevea brasiliensis is an important plant species currently cultivated for the commercial production of natural rubber. As the demand for rubber continues to increase, it is important to identify alternative sources of natural rubber and to increase plant rubber content using molecular approaches. Taraxacum kok-saghyz, a Russian dandelion, produces natural rubber that is of high quality. In this study, the SMALL RUBBER PARTICLE PROTEIN (SRPP) promoter from H. brasiliensis was characterized to determine its suitability for the expression of latex-specific genes in Taraxacum brevicorniculatum which is another Russian dandelion species of T. kok-saghyz from the similar geographical areas. Studies using transgenic Taraxacum plants carrying the SRPP promoter::β-glucuronidase (GUS) sequence indicate that the SRPP promoter does induce gene expression primarily in laticiferous tissues. Additionally, the promoter was regulated by various external conditions including light, tapping, and cold. These findings suggest that the SRPP promoter will be a useful molecular tool for the manipulation of gene expression in the laticiferous tissues of Taraxacum plant species.

Aiming to complement the group’s taxonomy, the objective of this work is to describe and characterise, searching for morpho-anatomical patterns, the leaf anatomy of the Manilkara species occurring in the northeast region of Brazil. The leaves of all Manilkara species were analysed, using herbaria specimens and collections made in the field by the authors. The characterisation of the leaf blades was performed according to the usual protocols in vegetal anatomy. As diagnosed by anatomical characters, the Manilkara Adans. genus presents petioles with a thick cuticle, a uniseriate epidermis, a sclerenchymatous sheath involving the vascular bundle, and laticifers on the cortical and medullar regions. The anatomical data increase the M. elata and M. huberi synonymisation and show the importance of the anatomy as a complementary tool to the taxonomy based on external morphology.

Laticifer proteins (LP) of Calotropis procera were fractionated by ion-exchange chromatography, and the influence of a sub-fraction (LP(PI)) on the inflammatory response of Swiss mice challenged by Salmonella enterica Ser. Typhimurium was investigated.Mice (n = 10) received LP(PI) (30 or 60 mg/kg) in a single inoculum by the intraperitoneal route 24 h before infection. To investigate the relevance of the proteolytic activity, three additional groups were included: the first one received heat-treated LP (30 mg/kg-30 min at 100 °C), the second received LP (30 mg/kg) inactivated by iodoacetamide, and a control group received only phosphate-buffered saline (PBS).The survival rate reached 100 % in mice treated with LP(PI) and was also observed with the other treatment, whereas the PBS group died 1-3 days after infection. The neutrophil infiltration into the peritoneal cavity of pretreated mice was enhanced and accompanied by high bacterial clearance from the bloodstream. Tumor necrosis factor-alpha mRNA transcripts, but not interferon-gamma, were detected early in spleen cells of pretreated mice after infection; however, the nitric oxide contents in the bloodstream were decreased in comparison to the PBS group.The inflammatory stimulus of C. procera proteins increased phagocytosis and balanced the nitric oxide release in the bloodstream, preventing septic shock induced by Salmonella infection.

Tapping panel dryness (TPD) is a complex physiological syndrome found widely in rubber tree (Hevea brasiliensis) plantations that causes severe yield loss in natural rubber-producing countries. In an earlier study, we confirmed that there is a negative correlation between HbMyb1 expression and TPD severity. To further investigate the function of HbMyb1 in TPD, HbMyb1 was over-expressed in tobacco controlled by a CaMV 35S promoter. In transgenic plants expressing HbMyb1, cell death induced by UV-B irradiation, paraquat and the hypersensitive reaction to necrotrophic fungal infection (Botrytis cinerea) was suppressed with a close correlation between HbMyb1 protein levels and the extent of suppression. In addition the nuclear condensation and degradation were observed in laticifer cells of TPD trees, while the nucleus of laticifer cells of healthy trees was morphologically normal. On the basis of the results described above, we propose that HbMyb1 maybe suppress stress induced cell death in rubber trees.

A laticifer is a cell involved in plant defense against biotic stresses such as herbivores and microorganisms; however, its gene expression is poorly understood. We compared protein accumulation and transcriptomes among laticifers of lignified and unlignified organs of mulberry (Morus alba), which has a non-articulated, branched type of laticifer. LA-a (equivalent to MLX56) and its homolog LA-b (insecticidal chitinase-like proteins containing two chitin-binding domains) were major proteins in laticifers of unlignified organs, and another protein (LA-c) was a major protein in laticifers of lignified organs. Purification, cDNA cloning, and bioassay of LA-c revealed that LA-c was an acidic class I chitinase having antifungal but not insecticidal activity. Comparative mRNA-Seq analysis using a GS-FLX revealed transcripts of other possible defense-related proteins. Jacalin-like lectin, galacturonase-inhibitor, and pathogenesis-related proteins were also abundant; however, the relative amounts differed among laticifers of lignified and unlignified organs. The results suggest a discontinuous laticifer network in planta and adaptation to different potential enemies among these organs.

MYC2 transcription factor is a key component of the core module COI1-JAZ-MYC2 of jasmonate signaling in Arabidopsis, but the MYC transcription factor (s) associated with jasmonate signaling in jasmonate-responsive laticifer cells remains to be identified. Two full-length cDNAs, designated HblMYC1 and HblMYC2, were isolated from laticifer cells in Hevea brasiliensis by the method of RACE. HblMYC1 contained 1431bp ORF encoding a putative protein of 476 amino acids while HblMYC2 contained 1428bp ORF encoding a putative protein of 475 amino acids. Bioinformatic analysis showed that the putative proteins, HblMYC1 and HblMYC2, possessed a bHLH domain and were most related to the MYC2 among the selected 27 MYC members with identified functions in Arabidopsis. In addition to the presence of cis-regulatory elements involving jasmonate responsiveness in the promoter regions of HblMYC1 and HblMYC2, the abscisic acid-, salicylic acid- and gibberellin-responsive elements were found in the promoter region of HblMYC1. Transcripts of HblMYC1 and HblMYC2 were most abundant in latex, relatively low in male flowers and nearly undetected in bark tissues and roots by real-time RT-PCR analysis. Regular tapping, mechanical wounding, and ethrel remarkably up-regulated HblMYC1 expression, but had little effect on the expression of HblMYC2 in laticifer cells. Successive tapping, however, significantly down-regulated the expression of HblMYC2 while up-regulating the expression of HblMYC1. The HblMYC2 expression took a mutual ebb and flow relationship with the HblMYC1 expression upon treatment with methyl jasmonate. Characterization of HblMYC1 and HblMYC2 will contribute to the understanding of jasmonate signaling in laticifiers, a kind of specialized tissue for natural rubber biosynthesis in Hevea brasiliensis.

AP2/ERF transcription factors play an important role in regulation of the cross-talk between ethylene and jasmonate signaling pathways mediating defense responses of plants to biotic and abiotic stresses. In this study, an AP2/ERF transcription factor gene was isolated and characterized from laticifers of rubber tree by using RACE and real time PCR. The full length cDNA, referred to as HbEREBP1, was 1,095 bp in length and contained a 732 bp open reading frame encoding a putative protein of 243 amino acid residues. The molecular mass of the putative protein is 26.4 kDa with a pI of 9.46. The deduced amino acid sequence had a specific domain of AP2 superfamily and an ethylene-responsive element binding factor-associated amphiphilic repression motif, sharing 42.4, 39.1, and 38.0% identity with that of AtERF11, AtERF4, and AtERF8 in Arabidopsis, respectively. HbEREBP1 expression was down-regulated by tapping and mechanical wounding in the laticifers of adult trees. It was also down-regulated at early stage while up-regulated at late stage upon treatment with exogenous ethephon or methyl jasmonate, which was reverse to the case of defense genes in laticifers of epicormic shoots of rubber tree. Our results suggest that HbEREBP1 may be a negative regulator of defense genes in laticifers.

Proteins from latex of Calotropis procera (CpLP), Plumeria rubra (PrLP), Carica candamarcensis (P1G10) and Euphorbia tirucalli (EtLP) were tested for antifungal activity against phytopathogens. CpLP and P1G10 inhibited each fungi analyzed. PrLP and EtLP did not exert inhibition. CpLP and P1G10 exhibited preferential inhibitory activity towards R. solani (IC₅₀ = 20.7 and 25.3 µg/ml, respectively). The inhibitory activity was lost after heat treatment or proteolysis, providing evidence for the involvement of proteins in the inhibitory effect. Treatment of CpLP or P1G10 with Dithiothreitol improved both, the endogenous proteolytic activity and the antifungal properties. Conversely, pre-treatment of CpLP or P1G10 with iodoacetamide drastically reduced endogenous proteolytic activities and partially abrogated antifungal activity. Similar results were observed when spores were challenged to germinate in the presence of laticifer proteins. The purified cysteine proteinase CMS2MS2 from Carica candamarcensis latex or papain (E.C. 3.4.22.2), a cysteine proteinase from latex of Carica papaya L., but not trypsin (EC 3.4.21.4) or chymotrypsin (EC 3.4.21.1), two serine proteases, replicated the results obtained with CpLP or P1G10, thus restricting the antifungal property to latex plant cysteine proteinases. CpLP, CMS2MS2 and papain induced production of reactive oxygen species in spores of F. solani, suggesting that inhibition could be linked to oxidative stress. Proteome analysis of CpLP by 2-D electrophoresis and MALDI-TOF-TOF confirmed the existence of various pathogenic-related proteins such as chitinases, peroxidases and osmotins. The results support that laticifer proteins are part of plant defense repertoire against phytopathogenic fungi.

The plant secondary metabolites glucosinolates (GSL) have important functions in plant resistance to herbivores and pathogens. We identified all major GSL that accumulated in S-cells in Arabidopsis by MALDI-TOF MS, and estimated by LC-MS that the total GSL concentration in these cells is >130 mM. The precise locations of the S-cells outside phloem bundles in rosette and cauline leaves and in flower stalks were visualised using sulphur mapping by cryo-SEM/energy-dispersive X-ray analysis. S-cells contain up to 40% of the total sulphur in flower stalk tissues. S-cells in emerging flower stalks and developing leaf tissues show typical signs of programmed cell death (PCD) or apoptosis, such as chromatin condensation in the nucleus and blebbing of the membranes. TUNEL staining for DNA double-strand breaks confirmed the occurrence of PCD in S-cells in post-meristematic tissues in the flower stalk as well as in the leaf. Our results indicate that S-cells in post-meristematic tissues show an extreme degree of metabolic specialisation in addition to PCD. Accumulation and maintenance of a high concentration of GSL in these cells are accompanied by degradation of a number of cell organelles. The substantial changes in cell composition during S-cell differentiation indicate the importance of this particular GSL-based phloem defence system. The specific anatomy of the S-cells and the ability to accumulate specialised secondary metabolites is similar to that of the non-articulated laticifer cells in latex plants, suggesting a common evolutionary origin.

Lettuce is an economically important leafy vegetable that accumulates a milk-like sap called latex in the laticifer. Previously, we conducted a large-scale lettuce latex proteomic analysis. However, the identified proteins were obtained only from lettuce ESTs and proteins deposited in NCBI databases. To extend the number of known latex proteins, we carried out an analysis identifying 302 additional proteins that were matched to the NCBI non-redundant protein database. Interestingly, the newly identified proteins were not recovered from lettuce EST and protein databases, indicating the usefulness of this hetero system in MudPIT analysis. Gene ontology studies revealed that the newly identified latex proteins are involved in many processes, including many metabolic pathways, binding functions, stress responses, developmental processes, protein metabolism, transport and signal transduction. Application of the non-redundant plant protein database led to the identification of an increased number of latex proteins. These newly identified latex proteins provide a rich source of information for laticifer research.

Efficient sucrose loading in rubber-producing cells (laticifer cells) is essential for retaining rubber productivity in Hevea brasiliensis, but the molecular mechanisms underlying the regulation of this process remain unknown. Here, we functionally characterized a putative Hevea SUT member, HbSUT3, mainly in samples from regularly exploited trees. When expressed in yeast, HbSUT3 encodes a functional sucrose transporter that exhibits high sucrose affinity with a K(m) value of 1.24 mm at pH 4.0, and possesses features typical of sucrose/H(+) symporters. In planta, when compared to the expression of other Hevea SUT genes, HbSUT3 was found to be the predominant member expressed in the rubber-containing cytoplasm (latex) of laticifers. The comparison of HbSUT3 expression among twelve Hevea tissues demonstrates a relatively tissue-specific pattern, i.e. expression primarily in the latex and in female flowers. HbSUT3 expression is induced by the latex stimulator Ethrel (an ethylene generator), and relates to its yield-stimulating effect. Tapping (the act of rubber harvesting) markedly increased the expression of HbSUT3, whereas wounding alone had little effect. Moreover, the expression of HbSUT3 was found to be positively correlated with latex yield. Taken together, our results provide evidence favouring the involvement of HbSUT3 in sucrose loading into laticifers and in rubber productivity.

Although jasmonate (JA) signaling has recently been well elucidated in Arabidopsis thaliana, the JASMONATE ZIM domain proteins (JAZs) are suggested to have an important role in regulating specific JA responses in plants, not much is known about how the specific JA responses are regulated in plants. Laticifer is the sole site of natural rubber biosynthesis in rubber tree. Here, the full-length cDNA and promoter region of HbJAZ1 gene were cloned and characterized from the laticifer cells in rubber tree by a combination of molecular techniques and bioinformatic analyses. The cDNA was 1,442-bp long containing an 852-bp open reading frame (ORF) encoding a putative protein of 284 amino acids. The predicted HbJAZ1 protein possessed the conservative ZIM and Jas domains, and was homologous to the JAZ members from Arabidopsis and the JAZ homologs from other plant species. The 976-bp promoter region of HbJAZ1 was isolated using PCR-based DNA Walking. TATA box and other core configurations as well as some special cis-acting elements involving in gibberellin-, jasmonic acid- and wound-responsiveness were found within this region. Transcription pattern analysis revealed that HbJAZ expression was strongly up-regulated by tapping and mechanical wounding, but not by ethrel application. Characterization of HbJAZ1 will attribute to understand JA signaling in laticifers, a specialized tissue for natural rubber biosynthesis in rubber trees.

Background: Telosma pallida (L.) Kurz., is one of the traditional medicinal plants from Asclepiadaceae family. It is abundantly available in Gujarat, India during monsoon. It is a slender and deciduous climber used traditionally in the treatment of various diseases such as cough, cold, skin diseases etc., Aims: To study the leaves of Telosma pallida (L.) Kurz. for detailed Pharmacognostic characters and Analytical Profile. Materials and Methods: In this work pharmacognostic, preliminary phytochemical screening, fluorescence analysis, and HPTLC profile of leaves were determined following standard procedures. Results: The leaves are simple, entire, opposite, cordate in shape, the upper surface is dark green and lower surface is light green in colour. The Transverse section of a leaf through petiole shows the presence of uniseriate multicellular warty trichomes and laticifer cells at the adjacent side of the phloem. Surface study shows paracytic and anomocytic type of stomata. Preliminary physicochemical analysis shows water soluble extractive value as 28.44 ± 2.89% w/w and alcohol soluble extractive value as 14.60 ± 0.79% w/w. Preliminary phytochemical analysis of extracts shows the presence of glycosides, alkaloids, carbohydrates, proteins, amino acids and flavonoids. Conclusion: The information obtained from this research work may be useful to establish the botanical as well as analytical standards for the leaves of T. pallida.