Initiation of rubber biosynthesis: In vitro comparisons of benzophenone-modified diphosphate analogues in three rubber-producing species

Natural rubber is the main component of latex obtained from laticifer cells of Hevea brasiliensis. For improving rubber yield, it is essential to understand the genetic molecular mechanisms responsible for laticifer differentiation and rubber biosynthesis. Jasmonate enhances both secondary laticifer differentiation and rubber biosynthesis. Here, we carried out time-course RNA-seq analysis in suspension-cultured cells treated with methyljasmonic acid (MeJA) to characterize the gene expression profile. Gene Ontology (GO) analysis showed that the term “cell differentiation” was enriched in upregulated genes at 24 h after treatment, but inversely, the term was enriched in downregulated genes at 5 days, indicating that MeJA could induce cell differentiation at an early stage of the response. Jasmonate signaling is activated by MYC2, a basic helix–loop–helix (bHLH)-type transcription factor (TF). The aim of this work was to find any links between transcriptomic changes after MeJA application and regulation by TFs. Using an in vitro binding assay, we traced candidate genes throughout the whole genome that were targeted by four bHLH TFs: Hb_MYC2-1, Hb_MYC2-2, Hb_bHLH1, and Hb_bHLH2. The latter two are highly expressed in laticifer cells. Their physical binding sites were found in the promoter regions of a variety of other TF genes, which are differentially expressed upon MeJA exposure, and rubber biogenesis-related genes including SRPP1 and REF3. These studies suggest the possibilities that Hb_MYC2-1 and Hb_MYC2-2 regulate cell differentiation and that Hb_bHLH1 and Hb_bHLH2 promote rubber biosynthesis. We expect that our findings will help to increase natural rubber yield through genetic control in the future.

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UNRAVELING THE MYSTERY OF NATURAL RUBBER BIOSYNTHESIS. PART II: COMPOSITION AND GROWTH OF IN VITRO NATURAL RUBBER USING HIGH-RESOLUTION SIZE EXCLUSION CHROMATOGRAPHY

Rubber Chemistry and Technology ◽

10.5254/rct.14.87913 ◽

2014 ◽

Vol 87 (3) ◽

pp. 451-458 ◽

Cited By ~ 2

Author(s):

Cheng Ching K. Chiang ◽

Balaka Barkakaty ◽

Judit E. Puskas ◽

Wenshuang Xie ◽

Katrina Cornish ◽

...

Keyword(s):

High Resolution ◽

Natural Rubber ◽

Size Exclusion Chromatography ◽

Rubber Tree ◽

Size Exclusion ◽

Common Source ◽

Rubber Biosynthesis ◽

Flight Mass Spectrometry ◽

Exclusion Chromatography

ABSTRACT The superior properties of natural rubber (cis-1,4-polyisoprene [NR]) are a function of its structure and composition, properties that still remain a mystery and that are irreplaceable by any synthetic rubber. NR from guayule (Parthenium argentatum) has been gaining special interest for its hypoallergenic properties while maintaining superior mechanical properties that are commonly associated with the Brazilian rubber tree (Hevea brasiliensis), the most common source of NR. Techniques exist to isolate washed rubber particles (WRPs) that contain enzymatically active rubber transferase, to study NR biosynthesis, and previous work on the in vitro NR growth in Hevea has demonstrated the presence of around 50 wt% of a low molecular weight ([MW], Mn <10 000 g/mol) fraction. Structural and compositional analyses of this low MW fraction in Hevea are challenging due to the high protein content. We discuss the analysis and composition of guayule latex and WRPs using high-resolution Size Exclusion Chromatography. We also discuss the composition of the soluble fraction of inactive guayule latex using matrix-assisted laser desorption ionization/time of flight mass spectrometry.

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UNRAVELING THE MYSTERY OF NATURAL RUBBER BIOSYNTHESIS. PART I: INVESTIGATION OF THE COMPOSITION AND GROWTH OF IN VITRO NATURAL RUBBER USING HIGH RESOLUTION SIZE EXCLUSION CHROMATOGRAPHY

Rubber Chemistry and Technology ◽

10.5254/1.3570528 ◽

2011 ◽

Vol 84 (2) ◽

pp. 166-177 ◽

Cited By ~ 4

Author(s):

Chengching K. Chiang ◽

Wenshuang Xie ◽

Colleen McMahan ◽

Judit E. Puskas

Keyword(s):

High Resolution ◽

Natural Rubber ◽

Size Exclusion Chromatography ◽

Weight Fraction ◽

Size Exclusion ◽

Gravimetric Analysis ◽

In Vitro Synthesis ◽

Rubber Biosynthesis ◽

Exclusion Chromatography

Abstract Monitoring the growth of in vitro natural rubber was accomplished by high resolution size exclusion chromatography, SEC. Washed rubber particles isolated from H. brasiliensis latex, containing the rubber transferase enzyme, were used to catalyze the polymerization of synthetic isopentenyl pyrophosphate monomer in the presence of farnesyl pyrophosphate initiator. The high-resolution SEC was able to detect the formation of new rubber. Changes in the low molecular weight fraction were also detected. Gravimetric analysis revealed ∼30% mass gain after the in vitro synthesis. The overall gel content was found to be reduced, which further supported the formation of new rubber. This is the first report that utilizes high-resolution SEC to monitor the in vitro NR growth without the use of radiolabeling.

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Biosynthesis of Natural Rubber: Current State and Perspectives

International Journal of Molecular Sciences ◽

10.3390/ijms20010050 ◽

2018 ◽

Vol 20 (1) ◽

pp. 50 ◽

Cited By ~ 12

Author(s):

Xiao Men ◽

Fan Wang ◽

Guo-Qiang Chen ◽

Hai-Bo Zhang ◽

Mo Xian

Keyword(s):

Natural Rubber ◽

Growth Conditions ◽

Genetically Engineered ◽

Human Society ◽

Rubber Biosynthesis ◽

The Past ◽

Current State ◽

Natural Rubber Biosynthesis ◽

Chain Polymerization

Natural rubber is a kind of indispensable biopolymers with great use and strategic importance in human society. However, its production relies almost exclusively on rubber-producing plants Hevea brasiliensis, which have high requirements for growth conditions, and the mechanism of natural rubber biosynthesis remains largely unknown. In the past two decades, details of the rubber chain polymerization and proteins involved in natural rubber biosynthesis have been investigated intensively. Meanwhile, omics and other advanced biotechnologies bring new insight into rubber production and development of new rubber-producing plants. This review summarizes the achievements of the past two decades in understanding the biosynthesis of natural rubber, especially the massive information obtained from the omics analyses. Possibilities of natural rubber biosynthesis in vitro or in genetically engineered microorganisms are also discussed.

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The stereochemistry of rubber biosynthesis

Proceedings of the Royal Society of London Series B Biological Sciences ◽

10.1098/rspb.1966.0006 ◽

1966 ◽

Vol 163 (993) ◽

pp. 519-523 ◽

Cited By ~ 37

Keyword(s):

Natural Rubber ◽

Farnesyl Pyrophosphate ◽

Isopentenyl Pyrophosphate ◽

Rubber Biosynthesis

The stereochemistry of the formation of natural rubber and trans-trans -farnesyl pyrophosphate in latex has been studied in vitro using [2- 14 C-(4 R )-4- 3 H 1 ] and [2- 14 C-(4 S )-4- 3 H 1 ]mevalonates as substrates. The proton eliminated from C-2 of isopentenyl pyrophosphate during the formation of farnesyl pyrophosphate in latex has the same steric position as that released in the liver system, and is epimeric with that eliminated during the biosynthesis of rubber. Complete stereospecificity is exhibited by the enzymes in the two cases. In rubber biosynthesis isoprene residues are incorporated directly in an all- cis configuration without the intervention of trans structures.

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Stereochemistry of Rubber Biosynthesis

Rubber Chemistry and Technology ◽

10.5254/1.3539083 ◽

1967 ◽

Vol 40 (3) ◽

pp. 679-683

Author(s):

B. L. Archer ◽

D. Barnard ◽

E. G. Cockbain ◽

J. W. Cornforth ◽

Rita H. Cornforth ◽

...

Keyword(s):

Natural Rubber ◽

Farnesyl Pyrophosphate ◽

Isopentenyl Pyrophosphate ◽

Rubber Biosynthesis

Abstract The stereochemistry of the formation of natural rubber and trans-trans-farnesyl pyrophosphate in latex has been studied in vitro using [2−14C−(4R)−3H1] and [2−14C−(4S)−4−3H1]mevalonates as substrates. The proton eliminated from C-2 of isopentenyl pyrophosphate during the formation of farnesyl pyrophosphate in latex has the same steric position as that released in the liver system, and is epimeric with that eliminated during the biosynthesis of rubber. Complete stereospecificity is exhibited by the enzymes in the two cases. In rubber biosynthesis isoprene residues are incorporated directly in all-cis configuration without the intervention of trans structures.

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Magnesium ion regulation of in vitro rubber biosynthesis by Parthenium argentatum Gray

Phytochemistry ◽

10.1016/j.phytochem.2006.04.010 ◽

2006 ◽

Vol 67 (15) ◽

pp. 1621-1628 ◽

Cited By ~ 9

Author(s):

Bernardo M.T. da Costa ◽

Jay D. Keasling ◽

Colleen M. McMahan ◽

Katrina Cornish

Keyword(s):

Magnesium Ion ◽

Parthenium Argentatum ◽

Ion Regulation ◽

Rubber Biosynthesis

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Ultrastructural Investigations of the Contractile Filaments of Amoebae

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100050950 ◽

1974 ◽

Vol 32 ◽

pp. 188-189

Author(s):

P.L. Moore

Keyword(s):

Cytoplasmic Streaming ◽

Three Dimensional ◽

Freeze Fracture ◽

Amoeboid Movement ◽

Different Types ◽

Chemical Conditions ◽

Complete Interpretation

Previous freeze fracture results on the intact giant, amoeba Chaos carolinensis indicated the presence of a fibrillar arrangement of filaments within the cytoplasm. A complete interpretation of the three dimensional ultrastructure of these structures, and their possible role in amoeboid movement was not possible, since comparable results could not be obtained with conventional fixation of intact amoebae. Progress in interpreting the freeze fracture images of amoebae required a more thorough understanding of the different types of filaments present in amoebae, and of the ways in which they could be organized while remaining functional.The recent development of a calcium sensitive, demembranated, amoeboid model of Chaos carolinensis has made it possible to achieve a better understanding of such functional arrangements of amoeboid filaments. In these models the motility of demembranated cytoplasm can be controlled in vitro, and the chemical conditions necessary for contractility, and cytoplasmic streaming can be investigated. It is clear from these studies that “fibrils” exist in amoeboid models, and that they are capable of contracting along their length under conditions similar to those which cause contraction in vertebrate muscles.

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In Vitro Induction of Crystals of MT Protein by Vinblastine-Colcemid in Mouse Spermatids

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100050676 ◽

1974 ◽

Vol 32 ◽

pp. 132-133

Author(s):

John J. Wolosewick ◽

John H. D. Bryan

Keyword(s):

Endoplasmic Reticulum ◽

Smooth Endoplasmic Reticulum ◽

Nuclear Ring ◽

Rough Er ◽

Distal Direction ◽

In Vitro Induction

Early in spermiogenesis the manchette is rapidly assembled in a distal direction from the nuclear-ring-densities. The association of vesicles of smooth endoplasmic reticulum (SER) and the manchette microtubules (MTS) has been reported. In the mouse, osmophilic densities at the distal ends of the manchette are the organizing centers (MTOCS), and are associated with the SER. Rapid MT assembly and the lack of rough ER suggests that there is an existing pool of MT protein. Colcemid potentiates the reaction of vinblastine with tubulin and was used in this investigation to detect this protein.

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Induction and Differentiation of Dental Epithelium in Vivo and in Vitro

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100053425 ◽

1982 ◽

Vol 40 ◽

pp. 142-145

Author(s):

E. J. Kollar

Keyword(s):

Connective Tissue ◽

Oral Mucosa ◽

Basal Lamina ◽

Functional Derivatives ◽

Specific Source ◽

Dental Epithelium ◽

Connective Tissue Stroma

The differentiation and maintenance of many specialized epithelial structures are dependent on the underlying connective tissue stroma and on an intact basal lamina. These requirements are especially stringent in the development and maintenance of the skin and oral mucosa. The keratinization patterns of thin or thick cornified layers as well as the appearance of specialized functional derivatives such as hair and teeth can be correlated with the specific source of stroma which supports these differentiated expressions.

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