The latex collected from the Hevea brasiliensis tree is today the only commercial source of natural rubber (NR), the cis-1,4-polyisoprene polymer, a strategic raw material. The Hevea latex is a very complex material both in its structure and composition. In terms of structure, it is a colloidal dispersion where various micrometric objects, mainly rubber particles and lutoids, are dispersed in the cytoplasmic serum (C-serum). The rubber fraction is the most abundant, followed by the C-serum and the lutoids. In terms of composition, the fresh latex contains about 60% of water, 35% of cis-1,4-polyisoprene and 5% of non-isoprene molecules. These non-isoprenes are biochemical compounds mostly including proteins, lipids, carbohydrates and minerals, and their distribution in the fractions of latex is not homogeneous. Although the non-isoprenes represent a minor part of the latex, some of them are retained in NR after latex processing and recognized to play a crucial role on the NR properties. Actually, the non-isoprene molecules are likely behind the better mechanical properties of NR over its synthetic counterpart, but they are also responsible for the high variability of NR quality. This variability of NR quality is a major drawback in NR industry and is directly linked to the latex composition, which is influenced by various physical and physiological parameters. The biochemical composition of latex matters, and this chapter is thus an overview of the protein, lipid, carbohydrate and mineral contents in latex, as well as their distribution in the three main fractions of latex.

Biochemical composition of Hevea brasiliensis latex: A focus on the protein, lipid, carbohydrate and mineral contents

Salinity is one of the most severe forms of abiotic stress and affects crop yields worldwide. Plants respond to salinity stress via a sophisticated mechanism at the physiological, transcriptional and metabolic levels. However, the molecular regulatory networks involved in salt and alkali tolerance have not yet been elucidated. We developed an RNA-seq technique to perform mRNA and small RNA (sRNA) sequencing of plants under salt (NaCl) and alkali (NaHCO3) stress in tobacco. Overall, 8064 differentially expressed genes (DEGs) and 33 differentially expressed microRNAs (DE miRNAs) were identified in response to salt and alkali stress. A total of 1578 overlapping DEGs, which exhibit the same expression patterns and are involved in ion channel, aquaporin (AQP) and antioxidant activities, were identified. Furthermore, genes involved in several biological processes, such as “photosynthesis” and “starch and sucrose metabolism,” were specifically enriched under NaHCO3 treatment. We also identified 15 and 22 miRNAs that were differentially expressed in response to NaCl and NaHCO3, respectively. Analysis of inverse correlations between miRNAs and target mRNAs revealed 26 mRNA-miRNA interactions under NaCl treatment and 139 mRNA-miRNA interactions under NaHCO3 treatment. This study provides new insights into the molecular mechanisms underlying the response of tobacco to salinity stress.

https://www.mdpi.com/1422-0067/20/10/2391/pdf

Integration of mRNA and miRNA Analysis Reveals the Molecular Mechanism Underlying Salt and Alkali Stress Tolerance in Tobacco.

The miR169n-NF-YA8 regulation module involved in drought resistance in Brassica napus L.

The genus Eucalyptus occurs in a wide range of environmental conditions, including rainforests, subal-pine, arid/semi-arid and moist temperate zones. It includes species with the capacity to cope with extremely low water potential. This study aims to screen water stress tolerance in two Eucalyptus species under nursery conditions. Inter-specific variation in morphological, physiological, biochemical and molecular parameters in two Eucalyptus species (E. tereticornis and E. camaldulensis) with contrasting levels of tolerance to progressive short term water-deprived condition was evaluated. Water stress reduced growth measured in terms of root:shoot ratio and specific leaf area (SLA), photosynthetic parameters, leaf water potential and relative water content (RWC) in both genotypes. Biochemical parameters including total sugars, phenol, phytohormones (indole acetic acid and abscisic acid) and proline were found to significantly increase during stress in both genotypes. Water responsive transcripts like osmotin and DREB/CBF registered significant expression variation in the two genotypes, suggesting their key role in regulating water stress tolerance in Eucalyptus.

/pdf/variation-in-morpho-physiological-biochemical-and-molecular-1nkw8o22w3.pdf

Variation in morpho-physiological, biochemical and molecular responses of two Eucalyptus species under short-term water stress

To better understand how diatoms are capable of responding to environmental stress, protein expression during heat treatment of a thermo-intolerant (Phaeodactylum tricornutum) and thermo-tolerant (Chaetoceros muelleri) diatom (Chrysophyta) was investigated. The stress response is a universal and conserved mechanism of cell survival to unfavorable conditions. Typically, a 10 to 15° C temperature elevation above cell growth optimal causes constitutively expressed proteins to decrease and heat shock proteins (HSPs) to increase. HSPs are categorized by molecular weight among five classes with each apparently specialized for a particular function that enhances cell survival. One-dimensional SDS-PAGE of diatoms subjected to heat treatment revealed that P. tricornutum exhibited a typical stress response, but C. muelleri did not exhibit a characteristic response even at a greatly elevated temperature (50° C). This result was confirmed by total soluble protein assays. Chaetoceros muelleri may contain higher basal levels of HSPs than P. tricornutum allowing C. muelleri to better tolerate elevated temperatures. Western blot analysis using pea HSP70 (70 kDa) antisera of heat-treated P. tricornutum and C. muelleri validated the hypothesis that thermo-tolerant cells contain higher levels of constitutively expressed HSPs. Two-dimensional gel electrophoresis of heat-treated cells indicate that the small HSPs (17–30 kDa) played a role in the stress response similar to that found in vascular plants. Ongoing work is focused on the manipulation of the stress response through over-expression of key hsp genes.

Protein expression during heat stress in thermo‐intolerant and thermo‐tolerant diatoms

In order to meet the ever rising global demand for natural rubber, cultivation of Hevea is being extended to non-traditional regions of India where extreme climatic conditions like drought and low temperature negatively influence the crop performance. In order to ensure maximum productivity, identification of drought tolerant clones of Hevea which can cope up with stress and give better crop yield is essential. Several attempts have been made previously to identify genes that are associated with drought tolerance in Hevea. In the present study, quantitative expression analysis was made using quantitative PCR for seven drought associated transcripts in four clones of Hevea with varying levels of drought tolerance. Among the seven genes studied, Mitogen Activated Protein (MAP) kinase, Myeloblastosis (Myb) transcription factor, C-repeat responsive element/Dehydration Responsive Element (CRT/DRE) binding factor and Nuclear Factor Y subunit A (NFYA) showed a positive association with drought tolerance. Transcripts of ascorbate peroxidase and heat shock protein 70 (HSP 70) did not show any correlation with drought tolerance. Interestingly, catalase gene was found down regulated in all the clones under drought condition. The possible role of these genes based on their level of gene expression in four different clones of Hevea with varying levels of drought tolerance is discussed.

Quantitative expression analysis of drought responsive genes in clones of Hevea with varying levels of drought tolerance.

Cold stress is one of the major abiotic factors that influence the productivity and geographical distribution of many agriculturally important crops like Hevea brasiliensis. Cultivation of H. brasiliensis in India is being extended to northeastern regions, where low temperature during winter adversely affects its survival, growth, and productivity. Developing cold-tolerant genotypes is a primary requisite to maximize the productivity under such challenging environmental conditions. However, lack of methods for early evaluation of cold tolerance in the newly developed clones and the extensive time required for assessing their tolerance in the field are major constraints for clonal selection. The present study was initiated with an objective to identify and characterize cold stress responsive miRNAs from H. brasiliensis that show stronger association with cold tolerance. Next generation sequencing using Illumina HiSeq method revealed the expression of 21 and 29 conserved miRNA (from clone RRIM 600) families in cold-stressed and control samples, respectively. Forty-two novel miRNAs were identified from this study. Upon differential expression analysis, eight conserved miRNAs were found commonly expressed in both the samples. When expression analyses were performed subsequently with six selected miRNAs in two Hevea clones (viz. RRII 105 and RRIM 600), miR169 showed a strong association with cold tolerance. miRNAs such as miR482 and miR159 also exhibited association with cold tolerance. This study suggests the possibility of employing these miRNAs as markers for cold tolerance after validation in more number of genotypes with varying levels of cold tolerance.

Identification and validation of cold responsive microRNAs of Hevea brasiliensis using high throughput sequencing

MicroRNAs (miRNAs) in plants play critical role in regulating gene expression at the post-transcriptional level in a number of biological processes including plant growth, development and defense responses against biotic and abiotic stresses. Recent reports indicate the role of miRNAs in regulating genes associated with various metabolic as well as abiotic stress responsive pathways in Hevea. The present study was initiated with an objective to identify drought responsive miRNAs from Hevea and to elucidate their role in stress response/tolerance in four clones of Hevea with varying levels of drought tolerance. miRNAs were isolated from leaves of Hevea and annotated after sequencing. The selected miRNAs were quantified using real time PCR. Four of the miRNAs displayed definite pattern of expression. The miR164, miR167 and the novel miRNA (HbmiRn_42) were down regulated in susceptible clones. HbmiRn_42 got upregulated in tolerant clones, indicating its strong association with stress tolerance. On the contrary, miR482 was down regulated in tolerant clones, while there was no change in its expression level in susceptible clones. This study throws light on stress responsive miRNAs of Hevea and their regulation under drought conditions.

http://nopr.niscair.res.in/bitstream/123456789/35549/1/IJBT%2015%282%29%20153-160.pdf

Expression of miRNAs of Hevea brasiliensis under drought stress is altered in clones with varying levels of drought tolerance

Drought, in combination with high temperature and low humidity affects the productivity of Hevea brasiliensis, the natural rubber tree and its expansion to non-traditional regions. The genotypes of H. brasiliensis that perform well in traditional regions often failed in non-traditional regions thus necessitating breeding for stress-tolerant genotypes. This can be accomplished by adopting molecular-assisted selection method. Recent developments in identification of drought-responsive transcripts from H. brasiliensis and the findings on role of small RNAs indicate the possibility of employing them as markers for identification of suitable genotypes. In this study, we attempted to identify drought-responsive miRNAs from H. brasiliensis through next-generation sequencing (Illumina HiSeq) method. The results revealed the expression of 33 conserved and 32 novel drought-responsive miRNAs. Further, validation of differentially expressed miRNAs by quantitative expression analysis indicated the association of two novel miRNAs, viz., HbmiRn_63 and HbmiRn_42 and two conserved miRNAs, viz., miR168 and miR160 miRNAs with drought tolerance. These miRNAs can be employed as markers for drought tolerance after validation in a larger set of genotypes. This study opens up the possibility of employing miRNAs as markers for drought tolerance in Hevea.

Identification and validation of drought-responsive microRNAs from Hevea brasiliensis

Rootstock influences gene expression in scion in Hevea brasiliensis.

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