This document summarizes research on the cellulose synthase-like (CSL) family of genes, which encode glycosyltransferases that synthesize polysaccharide backbones in plant cell walls. There are several CSL subfamilies (A-J) that are involved in synthesizing different polysaccharides. Research has utilized seeds high in specific reserve polysaccharides to identify genes within subfamilies responsible for their synthesis. For example, CSLA genes were found to synthesize galactomannan backbones in guar seeds. CSLC and CSLF genes were shown to synthesize the xyloglucan and (1,3;1,4)-β-glucan backbones in nasturtium
Gmr2301 Breeding Transgenic Cattle For Human Therapeutics Avi Dey
Small breed cattle & pigs now can be part of small farm new product development via emerging agribio technology with recent breakthroughs in bioscience/bioengineering.
A number of developments have been made in the molecular biology of oat (Avena spp.) in recent years. Many of these were recently described at the Fourth International Oat Conference, held on 18 to 23 October, in Adelaide, South Australia. These advances include a report of oat transformation and regeneration, the characterisation of J3-glucanase genes in oat, the further development of a molecular genetic map in oats, and the characterisation of genes encoding novel oat grain proteins. A technique for assessing pedigrees in the oat and other cereal crops has been reported using a modified electrophoretic technique.
Cloning and extracellular expression of recombinant tissue plasminogen activa...bioejjournal
Tissue plasminogen activator (tPA) has noteworthy application in treatment of acute myocardial
infarctions. This study focuses on expression of rt-PA using microbial systems in order to reduce cost
without compromising on quality as an alternative to commercial (rt-PA)produced by using mammalian
host systems. In the present study, Pichia pastoris X-33strain was used as a host with pICZA expression
vector to obtain extracellular expression of full length tPA gene. Specific primers were designed in such a
way to get native tPA protein sequence in subsequent purification steps. Further, construct pICZA-tPA
was developed and electroporated into host to achieve stablert-PA gene by achieving genome integration.
The transformants were screened for phenotypic characters.Mut+phenotypic colony named Pichia tPA-3
showed expression of rt-PA at 66 kDa on SDS PAGE. Size Exclusion Chromatography (SEC) was
performed, resulting in product peak at same RT as reference standard. (alteplase).Cloning and expression
of rt-PA was successfully achieved in microbial system. Further process optimization at larger scales will
surely provide cost effective alternative to mammalian system forrt-PA production.
Gmr2301 Breeding Transgenic Cattle For Human Therapeutics Avi Dey
Small breed cattle & pigs now can be part of small farm new product development via emerging agribio technology with recent breakthroughs in bioscience/bioengineering.
A number of developments have been made in the molecular biology of oat (Avena spp.) in recent years. Many of these were recently described at the Fourth International Oat Conference, held on 18 to 23 October, in Adelaide, South Australia. These advances include a report of oat transformation and regeneration, the characterisation of J3-glucanase genes in oat, the further development of a molecular genetic map in oats, and the characterisation of genes encoding novel oat grain proteins. A technique for assessing pedigrees in the oat and other cereal crops has been reported using a modified electrophoretic technique.
Cloning and extracellular expression of recombinant tissue plasminogen activa...bioejjournal
Tissue plasminogen activator (tPA) has noteworthy application in treatment of acute myocardial
infarctions. This study focuses on expression of rt-PA using microbial systems in order to reduce cost
without compromising on quality as an alternative to commercial (rt-PA)produced by using mammalian
host systems. In the present study, Pichia pastoris X-33strain was used as a host with pICZA expression
vector to obtain extracellular expression of full length tPA gene. Specific primers were designed in such a
way to get native tPA protein sequence in subsequent purification steps. Further, construct pICZA-tPA
was developed and electroporated into host to achieve stablert-PA gene by achieving genome integration.
The transformants were screened for phenotypic characters.Mut+phenotypic colony named Pichia tPA-3
showed expression of rt-PA at 66 kDa on SDS PAGE. Size Exclusion Chromatography (SEC) was
performed, resulting in product peak at same RT as reference standard. (alteplase).Cloning and expression
of rt-PA was successfully achieved in microbial system. Further process optimization at larger scales will
surely provide cost effective alternative to mammalian system forrt-PA production.
Hybrid rice breeding problems, prospects and future strategies by Deepak SharmaDeepak Sharma
The presentation describes all the constraints worldwide regarding hybrids in rice and potential solutions. The material includes all the findings and the researches going on in the world. Material collection is surely going to be very helpful from conventional and molecular point of view and having all the recent achievement and work done .
The 1st recombinant drug .
A protein chain or peptide hormone.
A dimer of an A-chain & a B-chain linked together by disulfide bonds, composed of 110 aa & molecular mass is 5808 Da.
A product of commercially important fermentation process that produce recombinant products.
Naturally produced by beta cells of the islets of Langerhans in the pancreas & by Brockmann body in some teleost fish.
The preproinsulin precursor of insulin is encoded by the INS gene.
Important for metabolism and utilization of energy from the ingested nutrients – especially glucose.
Failure of control of insulin level causes diabetes mellitus.
R protein expression in rice in the recombinant protien which is expressed in rice to overcome all the abiotic factors which is a stress to the rice in some non ecological condition
Hybrid rice breeding problems, prospects and future strategies by Deepak SharmaDeepak Sharma
The presentation describes all the constraints worldwide regarding hybrids in rice and potential solutions. The material includes all the findings and the researches going on in the world. Material collection is surely going to be very helpful from conventional and molecular point of view and having all the recent achievement and work done .
The 1st recombinant drug .
A protein chain or peptide hormone.
A dimer of an A-chain & a B-chain linked together by disulfide bonds, composed of 110 aa & molecular mass is 5808 Da.
A product of commercially important fermentation process that produce recombinant products.
Naturally produced by beta cells of the islets of Langerhans in the pancreas & by Brockmann body in some teleost fish.
The preproinsulin precursor of insulin is encoded by the INS gene.
Important for metabolism and utilization of energy from the ingested nutrients – especially glucose.
Failure of control of insulin level causes diabetes mellitus.
R protein expression in rice in the recombinant protien which is expressed in rice to overcome all the abiotic factors which is a stress to the rice in some non ecological condition
Extranuclear inheritance or cytoplasmic inheritance is the transmission of genes that occur outside the nucleus. It is found in most eukaryotes and is commonly known to occur in cytoplasmic organelles such as mitochondria and chloroplasts or from cellular parasites like viruses or bacteria. Determining the contribution of organelle genes to plant phenotype is hampered by several factors, including the paucity of variation in the plastid and mitochondrial genomes. Mitochondria are organelles which function to transform energy as a result of cellular respiration. Chloroplasts are organelles which function to produce sugars via photosynthesis in plants and algae. The genes located in mitochondria and chloroplasts are very important for proper cellular function, yet the genomes replicate independently of the DNA located in the nucleus, which is typically arranged in chromosomes that only replicate one time preceding cellular division. The extranuclear genomes of mitochondria and chloroplasts however replicate independently of cell division. They replicate in response to a cell's increasing energy needs which adjust during that cell's lifespan. There is consistent difference between the results from reciprocal crosses; generally only the trait from female parent is transmitted. In most cases, there is no segregation in the F2 and subsequent generations.
Plant genetic engineering is one of the key technologies for crop improvement as well as an emerging approach for producing recombinant proteins in plants. Both plant nuclear and plastid genomes can be genetically modified, yet fundamental functional differences between the eukaryotic genome of the plant cell nucleus and the prokaryotic-like genome of the plastid will have an impact on key characteristics of the resulting transgenic organism. So, which genome, nuclear or plastid, to transform for the desired transgenic phenotype? In this paper we compare the advantages and drawbacks of engineering plant nuclear and plastid genomes to generate transgenic plants with the traits of interest, and evaluate the pros and cons of their use for different biotechnology and basic research applications. The chloroplast is a pivotal organelle in plant cells and eukaryotic algae to carry out photosynthesis, which provides the primary source of the world’s food. The expression of foreign genes in chloroplasts offers several advantages over their expression in the nucleus: high-level expression, no position effects, no vector sequences allowing stable transgene expression. In addition, transgenic chloroplasts are generally not transmitted through pollen grains because of the cytoplasmic localization. In the past two decades, great progress in chloroplast engineering has been made.
Access to large-scale omics datasets i.e. genomics, transcriptomics, proteomics, metabolomics, phenomics, etc. has revolutionized biology and led to the emergence of systems approaches to advance our understanding of biological processes. With decreasing time and cost to generate these datasets, omics data integration has created both exciting opportunities and immense challenges for biologists, computational biologists, biostatisticians and biomathematicians. Genomics, transcriptomics, proteomics, and metabolomics together they help to bring out the best of characters in plants.
Extraction of β-galactosidase and β-glucosidase from the seeds of Tamarindus ...Open Access Research Paper
The enzymes β–galactosidase and β–glucosidase were extracted from the tamarind seeds using different buffers at different pH. Highest activity was obtained with 10 mM sodium acetate buffer, pH 5.6 and 10 mM tris buffer, pH 7.4. The effect of NaCl and Triton X–100 at different concentrations on the extraction of the enzymes indicated 10 mM sodium acetate buffer, pH 5.6 containing 1 M NaCl as a better extractant of the enzyme. The enzyme assay was carried out using p–nitrophenyl–β–D–galactoside and p–nitrophenyl–β–D–glucoside as substrates. Highest enzyme activities were observed on 6th and 24th day of germination. The protein content gradually decreased upto 5th day of germination and suddenly increased on 6th day. However, on subsequent days of germination, the protein content greatly decreased upto 11th day. During the latter period of germination (18th day onwards) the content remained almost constant. The kinetic parameters varied for both β–galactosidase and β–glucosidase. The activity of β– galactosidase was show to have an optimal operating condition at pH 5.5 and a temperature of 500C. The thermostability of the enzyme was in the range of 400C – 700C with the pH stability in the range of 5.0 – 7.0. The Km and Vmax values for pNPGal were determined as 66μM and 2.27nmolesmin-1. In contrast the activity of β– glucosidase was shown to have an optimal operating condition at pH 5.0 and a temperature of 300C. The thermostability of the enzyme was in the range of 270C – 350C with the pH stability in the range of 4.0 – 7.0. The Km and Vmax values for pNPGlu were determined as 121μM and 5.26nmolesmin-1. The presented study is a preliminary work carried out for the standardization of protocols. The purification and characterization of β–galactosidase and β– glucosidase is under progress.
Extraction of β-galactosidase and β-glucosidase from the seeds of Tamarindus ...Innspub Net
The enzymes β–galactosidase and β–glucosidase were extracted from the tamarind seeds using different buffers at different pH. Highest activity was obtained with 10 mM sodium acetate buffer, pH 5.6 and 10 mM tris buffer, pH 7.4. The effect of NaCl and Triton X–100 at different concentrations on the extraction of the enzymes indicated 10 mM sodium acetate buffer, pH 5.6 containing 1 M NaCl as a better extractant of the enzyme. The enzyme assay was carried out using p–nitrophenyl–β–D–galactoside and p–nitrophenyl–β–D–glucoside as substrates. Highest enzyme activities were observed on 6th and 24th day of germination. The protein content gradually decreased upto 5th day of germination and suddenly increased on 6th day. However, on subsequent days of germination, the protein content greatly decreased upto 11th day. During the latter period of germination (18th day onwards) the content remained almost constant. The kinetic parameters varied for both β–galactosidase and β–glucosidase. The activity of β–galactosidase was show to have an optimal operating condition at pH 5.5 and a temperature of 500C. The thermostability of the enzyme was in the range of 400C – 700C with the pH stability in the range of 5.0 – 7.0. The Km and Vmax values for pNPGal were determined as 66μM and 2.27nmolesmin-1. In contrast the activity of β–glucosidase was shown to have an optimal operating condition at pH 5.0 and a temperature of 300C. The thermostability of the enzyme was in the range of 270C – 350C with the pH stability in the range of 4.0 – 7.0. The Km and Vmax values for pNPGlu were determined as 121μM and 5.26nmolesmin-1. The presented study is a preliminary work carried out for the standardization of protocols. The purification and characterization of β–galactosidase and β–glucosidase is under progress.
CRISPR/Cas systems: The link between functional genes and genetic improvement. The discovery and modification of CRISPR/Cas system, a nature-occurred gene editing tool, opens an era for studying gene function and precision crop breeding
cutting-edge biotechnological tool for crop improvement
Used for pathogen resistance, abiotic tolerance, plant development and morphology and even secondary metabolism and fiber development
Introduction
History
Geographical distribution
Genome Structure
Anatomy and Life Cycle
Significance of Arabidopsis in Plant Genetics
Conclusion
References.
Loss of pollen-specific phospholipase NOT LIKE DAD triggers gynogenesis in maizePGS
This lecture was a part of Plant Genetics Seminars - PGS 2017/2018 at Assiut University. These seminars organized by Dr. Ahmed Sallam, Department of Genetics, Faculty of Agriculture, Assiut University
Abstract
Gynogenesis is an asexual mode of reproduction common to animals and plants, in which stimuli from the sperm cell trigger the development of the unfertilized egg cell into a haploid embryo. Fine mapping restricted a major maize QTL (quantitative trait locus) responsible for the aptitude of inducer lines to trigger gynogenesis to a zone containing a single gene NOT LIKE DAD (NLD) coding for a patatin-like phospholipase A. In all surveyed inducer lines, NLD carries a 4-bp insertion leading to a predicted truncated protein. This frameshift mutation is responsible for haploid induction because complementation with wild-type NLD abolishes the haploid induction capacity. Activity of the NLD promoter is restricted to mature pollen and pollen tube. The translational NLD::citrine fusion protein likely localizes to the sperm cell plasma membrane. In Arabidopsis roots, the truncated protein is no longer localized to the plasma membrane, contrary to the wild-type NLD protein. In conclusion, an intact pollen-specific phospholipase is required for successful sexual reproduction and its targeted disruption may allow establishing powerful haploid breeding tools in numerous crops.
10 nazir ahmad malla and mudasir bashir 215 plant protein kinases in signal ...Dheeraj Vasu
ABSTRACT: A protein kinase is a enzyme that modifies other proteins by adding phosphate groups to them. This results in a functional change of the target protein by changing enzyme activity, cellular location, or association with other proteins. Cells can interact to environmental fluctuations by transduction of extracellular signals, to produce intracellular responses. Membrane-impermeable signal molecules are recognized by receptors, which are localized on the plasma membrane of the cell. Binding of a ligand can result in the stimulation of an intrinsic enzymatic activity of its receptor or the modulation of a transducing protein. This review discusses the various protein kinases and their role in plants.
Francesca Gottschalk - How can education support child empowerment.pptxEduSkills OECD
Francesca Gottschalk from the OECD’s Centre for Educational Research and Innovation presents at the Ask an Expert Webinar: How can education support child empowerment?
Biological screening of herbal drugs: Introduction and Need for
Phyto-Pharmacological Screening, New Strategies for evaluating
Natural Products, In vitro evaluation techniques for Antioxidants, Antimicrobial and Anticancer drugs. In vivo evaluation techniques
for Anti-inflammatory, Antiulcer, Anticancer, Wound healing, Antidiabetic, Hepatoprotective, Cardio protective, Diuretics and
Antifertility, Toxicity studies as per OECD guidelines
Safalta Digital marketing institute in Noida, provide complete applications that encompass a huge range of virtual advertising and marketing additives, which includes search engine optimization, virtual communication advertising, pay-per-click on marketing, content material advertising, internet analytics, and greater. These university courses are designed for students who possess a comprehensive understanding of virtual marketing strategies and attributes.Safalta Digital Marketing Institute in Noida is a first choice for young individuals or students who are looking to start their careers in the field of digital advertising. The institute gives specialized courses designed and certification.
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How to Make a Field invisible in Odoo 17Celine George
It is possible to hide or invisible some fields in odoo. Commonly using “invisible” attribute in the field definition to invisible the fields. This slide will show how to make a field invisible in odoo 17.
A Strategic Approach: GenAI in EducationPeter Windle
Artificial Intelligence (AI) technologies such as Generative AI, Image Generators and Large Language Models have had a dramatic impact on teaching, learning and assessment over the past 18 months. The most immediate threat AI posed was to Academic Integrity with Higher Education Institutes (HEIs) focusing their efforts on combating the use of GenAI in assessment. Guidelines were developed for staff and students, policies put in place too. Innovative educators have forged paths in the use of Generative AI for teaching, learning and assessments leading to pockets of transformation springing up across HEIs, often with little or no top-down guidance, support or direction.
This Gasta posits a strategic approach to integrating AI into HEIs to prepare staff, students and the curriculum for an evolving world and workplace. We will highlight the advantages of working with these technologies beyond the realm of teaching, learning and assessment by considering prompt engineering skills, industry impact, curriculum changes, and the need for staff upskilling. In contrast, not engaging strategically with Generative AI poses risks, including falling behind peers, missed opportunities and failing to ensure our graduates remain employable. The rapid evolution of AI technologies necessitates a proactive and strategic approach if we are to remain relevant.
A workshop hosted by the South African Journal of Science aimed at postgraduate students and early career researchers with little or no experience in writing and publishing journal articles.
A review of the growth of the Israel Genealogy Research Association Database Collection for the last 12 months. Our collection is now passed the 3 million mark and still growing. See which archives have contributed the most. See the different types of records we have, and which years have had records added. You can also see what we have for the future.
2. Glycosyltransferases
• Cell-wall polysaccharides are synthesized by glycosyltransferases
(GTs).
• GTs are grouped into families (>90 known) based on sequence
similarities, particular motifs etc (Carbohydrate Active enZymes
(CAZy database).
• Arabidopsis has 455 genes in 41 families that encode GTs
• Catalyse transfer of glycosyl (sugar) residues from nucleotide sugars
to acceptors
• Some transfer only a single glycosyl residue.
• Others “processive” or “polymerizing” transferases use the product
of one addition as the acceptor for the next, producing a chain
(polysaccharide synthases)
• Specificity is in the enzyme – no template; highly specific for
polymer and bond formed
3. • From genome sequence of Arabidopsis, Somerville identified
a family of genes of unknown function with sequence
similarity to cellulose synthase (Cutler & Somerville 1997;
Richmond & Somerville 2000): cellulose synthase-like (CSL)
genes (30 CSL + 10 CESA genes in Arabidopsis)
• Six CSL gene subfamilies identified in Arabidopsis
• CSL gene subfamilies identified by a letter code: A, B, C etc.
• CSL gene subfamilies also identified in the genome sequence
of rice (Ozya sativa) (Hazen et al. 2002), but differences.
• Unlike Arabidopsis rice has no subfamily B or G
• Unlike Arabidopsis rice has two additional subfamilies: F and
H
4. Nine subfamilies of CSL genes
Subfamily Arabidopsis Rice
A 9 9
B 6 -
C 5 6
D 6 5
E 1 3
F - 8
G 3 -
H - 2
Subfamily J also present in some Poaceae genomes, but not rice (Fincher 2009)
6. CSL genes and proteins
• Cellulose synthase/cellulose synthase-like superfamily
genes encode family GT2 proteins.
• Type III integral membrane proteins with 3-6 predicted
multiple transmembrane spanning domains towards
the COOH teminus and 1-2 (usually) towards the NH2
terminus
Burton et al 2008 Plant
Physiology 146, 1821-1833
Hordeum vulgare CSLF3
• All have a D, D, D, QxxRW motif found in all processive
family GT2 proteins
8. • When CSL genes first discovered, all known GT2
glycosyltransferases (cellulose synthases, chitin synthases etc)
synthesized polysaccharides with repeating β-glycosyl residues
(processive β-glycosyltransferases)
Cutler & Somerville (1997) Current Biology 7: R108-R111
• Postulated that the different CSL subfamilies function in the
synthesis of plant cell-wall polysaccharides with backbones of
repeating β-glycosyl residues:
– heteromannans
– xyloglucans
– (13),(14)--glucans (present only in Poaceae & related families)
– callose [(1→3)--glucan]
– heteroxylans
– (1→4)--galactans
9. Using seeds to discover the functions of CSL
gene products
• Identifying the genes encoding enzymes involved in the
biosynthesis of -glycans in cell walls has been done using
particular seeds.
• The cotyledons or endosperms of many seeds have thick, non-
lignified secondary walls.
• These walls usually contain one polysaccharide that functions
as a reserve and is metabolised during germination
• These polysaccharides include galactomannans, xyloglucans
and (13),(14)--glucans
10. CSLA subfamily
Dhugga et al. 2004 [Science 303, 363-366]
• Used seeds of guar (Cyamopsis tetragonoloba) (family
Fabaceae) to investigate the genes encoding the synthesis of
the backbone of galactomannans
• The seeds of this and related plants in the Fabaceae have
thick cell walls containing galactomannans which are used as
additives (thickeners, stabilizers, emulsifiers and gelling
agents) in the food industry.
11. • Membrane preparations from developing guar seeds showed
mannan synthase (ManS) activity which used GDP-mannose
as the substrate
• Made 3 cDNA libraries at different stages of development
• Found 15 ESTs that were similar to CESA (i.e. CSL)
• Abundance of these ESTs in different libraries mirrored
pattern of ManS activity
12. • A full-length cDNA was assembled from 12 over-lapping
sequence tags
• Phylogenetic analysis showed that the gene grouped with the
CSLA subfamily of Arabidopsis and rice
• To determine the function of the putative CtManS (CSLA)
gene, they transformed it into embryogenic soybean
suspension-culture cells under the control of a seed-specific
promoter (heterologous expression)
• The cells were allowed to develop into mature somatic
embryos and membrane preparations from these showed
ManS enzymic activity (normally did not) and the CtManS
(CSLA) gene was expressed (Northern blots)
13. • Enzyme specifically used GDP-mannose and the product was
hydrolysed by an endo--mannanase but not by a cellulose;
acid hydrolysis of the product yielded only mannose. Indicated
a (1→4)--mannan was formed.
• During guar seed development, another membrane-bound
enzyme, α-galactosyltransferase (a family GT34 enzyme),
which uses UDP-galactose, adds α-galactosyl residues to the
mannan backbone
14. Liepman et al. (2005) PNAS 102, 2221-2226
• Heterologously expressed Arabidopsis CSLA genes (AtCSLA2,
7, and 9) in Drosophila Schneider 2 (S2) cells
• Isolated microsomal membranes from S2 cells and incubated
these with GDP-mannose, which gave (1→4)--mannan
• Also incubated with a mixture of GDP-mannnose and GDP-
glucose and a glucomannan was produced
• Glucomannans occur in lignified, secondary walls of eudicots
(hard woods) and galactoglucomannans occur (in large
proportions in lignified, secondary walls of coniferous
gymnosperms (softwoods)
Glucomannnan
Galactoglucomannan
15. • Also heterogenously expressed AtCSLE1 and OsCSLH1
Plenty of recombinant protein was obtained but was not active
in the enzyme assay.
Liepman et al. (2007) Plant Phyiology 143, 1881-183
• In the same way, heterogenously expressed:
From the grass Oryza sativa OsCSLA1
From the gymnosperm Pinus taeda PtCSLA1
From the moss Physcomitrella patens PpCSLA1
• Isolated microsomal membranes and incubated with GDP-
mannose and formed (1→4)--mannans; incubated with both
GDP-mannose and GDP-glucose and formed glucomannans
16. CSLC subfamily
Cocuron et al. (2007) PNAS 104, 8550-8555
• Used a similar approach to identify the gene encoding the
enzyme that synthesizes the (1→4)-β-glucan backbone of
xyloglucans.
• Used developing nasturtium (Tropaeolum majus, family
Tropaeolaceae) seeds, which at maturity have cotyledons with
thick cell walls that contain reserve xyloglucans.
17. Reserve xyloglucans in seed walls lack the fucosyl residues
found in xyloglucans in the primary walls of vegetative
organs of the same plant.
Seed xyloglucan
Primary wall
xyloglucan
18. • mRNA was isolated from developing seeds at the stage of
maximum deposition of the xyloglucans.
• A cDNA library was made and partial sequences of 10,000
cDNA clones determined.
• A single CSLC gene was overrepresented in the cDNA library.
• Heterologously expressed this gene in the yeast Pichia
pastoris and analysed the polysaccharides.
• P. pastoris usually has large amounts of (1→3)-β-glucan in its
cell walls, but only insignificant amounts of (1→4)-β-glucan.
• However, transgenic P. pastoris contained (1→4)-β-glucan.
• Similar results were obtained with the Arabidopsis CSLC4
gene (AtCSL4), the gene with the highest sequence similarity
to the TmCSLC
19. CSLF, H & J subfamilies
• Genes encode enzymes involved in the synthesis of
(13),(14)--glucans (-glucans)
• In flowering plants, (13),(14)--glucans occur only in cell
walls of Poaceae and related families
• Barley grains ~4-7% (13),(14)--glucans
Oat grains ~3-6%
Wheat grains ~0.5-1.0%
Barley (Hordeum vulgare)
• These glucans occur particularly in cell walls of starchy
endosperm and aleurone of cereal grains
• Walls of starchy endosperm and aleurone in barley contain 75%
and 26%, respectively
20. Diagram of the barley grain, showing the different organs, tissues and cell types.
Reproduced from Briggs (1978) with permission.
From: Harris, P.J., and Fincher, G.B. (2009). In: Bacic, A., Fincher, G.B., and Stone, B.A. eds, Chemistry, biochemistry, and biology
of (1→3)-β-glucans and related polysaccharides. San Diego, USA: Academic Press, Elsevier Inc. 621-654.
21. The caryopsis (C) of wheat and its pericarp (A, B).
From: Esau K (1953) Plant anatomy. John Wiley, New York, p 583.
22. • Linear polysaccharides with (13)-links (~30%) & (14)-links
(~70%)
• These two different linkages are not randomly distributed but
occur as:
cellotriosyl units:
and cellotetraosyl units:
• These units are joined by (13)-links
23. • In the grains of barley and oats, a high proportion of the
(13),(14)--glucans are water soluble
• Form viscous solutions:
• Cause filtration problems in brewing
• Contribute to haze formation in beer
• Reduce growth rate of monogastric animals (anti-nutritive effect)
• But lower serum cholesterol and reduce glycaemic index
• In the grains of wheat <3% water soluble
24. CLSF subfamily
Burton et al. (2006) Science 311: 1940-1942
• Quantitative trait loci (QTL) for (13),(14)--glucan content
of grain have been identified for barley
• One QTL with a large effect on (13),(14)--glucan content is
on barley chromosome 2H
• Genetic mapping has shown genome structures of common
cereals are similar (synteny)
• Used comparative genomics to identify a corresponding region
in the rice genome; contains a group of 6CSLF genes
(CSLF1,2,3,4,8,&9)
• Possible role of CSLF genes in (13),(14)--glucan synthesis
tested by expressing them in Arabidopsis (heterologous
expression)
25. • Expressed OsCSLF2 and 4 and examined transgenic
Arabidopsis by immunogold microscopy using a monoclonal
antibody that specifically recognizes (13),(14)--glucans
Epidermal walls of wild type and transgenic plants
• Found formation of (13),(14)--glucans in low
concentrations
26. Burton et al. (2008) Plant Physiology 146: 1821-1833
• Mapped CSLF genes in barley and found 4 of the 7 in the QTL
locus originally identified on chromosome 2H
• Spatial and temporal transcription patterns in developing
grains determined; transcripts of HvCSLF6 and HvCSLF9 were
predominant
27. CLSH subfamily
• Only found so far in Poaceae, so is another candidate for
encoding enzymes involved in (13),(14)--glucan
synthesis
• Barley has only one gene in this subfamily
Doblin et al. (2009) PNAS 106: 5996-6001
• Expressed the HvCSLH1 in Arabidopsis and detected
(13),(14)--glucans in the transgenic plants using
immunogold microscroscopy
• Examined expression of HvCSLH1 in barley: only weakly
transcribed in developing grain endosperm; it was most highly
transcribed in leaf tips
28. CLSJ subfamily
• Been found in some members of Poaceae, including barley,
maize, sorghum and wheat, but not in Brachypodium or rice.
• “Preliminary association mapping data suggest that the
HvCSLJ genes could also be involved in (13),(14)--glucan
synthesis” (Fincher 2009).
29. CSLD subfamily
• Genes of this subfamily are most similar to CESA genes
• Been suggested it is involved in cellulose synthesis but the
evidence is inconclusive
• Expression of CSLD genes is associated with tip-growing cells
• Analysis of CSLD mutants also indicated a role in tip-growing
cells
• Recent indications that CSLD proteins may be involved in the
synthesis of heteromannans (Yin et al. 2011 Molecular Plant,
In Press)
31. Evidence for CSL genes involvement in cell-wall
polysaccharide synthesis
– Heteromannans YES
– Xyloglucans YES
– (13),(14)--glucans (present only in Poaceae & related
families) YES
– callose [(13)--glucan] NO (other genes found)
– Heteroxylans NO (other genes found)
– (14)--galactans NO (but other genes not found so far)
– Also possibly involved (with CESA genes) in cellulose
synthesis
32. Key papers for discussion
Burton RA, Wilson SM, Hrmova M, Harvey AJ, Shirley NJ, Medhurst A, Stone BA,
Newbigin EJ, Bacic A, Fincher GB (2006) Cellulose synthase-like CslF genes mediate the
synthesis of cell wall (1,3;1,4)-β-D-glucans. Science 311: 1940-1942
Cocuron J-C, Lerouxel O, Drakakaki G, Alonso AP, Liepman AH, Keegstra K, Raikhel N,
Wilkerson CG (2007) A gene from the cellulose synthase-like C family encodes a β-1,4
glucan synthase. Proceedings of the National Academy of Science USA 104: 8550-8555
Doblin MS, Pettolino FA, Wilson SM, Campbell R, Burton RA, Fincher GB, Newbigin E,
Antony Bacic A (2009) A barley cellulose synthase-like CSLH gene mediates (1,3;1,4)-β-D-
glucan synthesis in transgenic Arabidopsis. Proceedings of the National Academy of
Science USA 106: 5996-6001
Dhugga KS, Barreiro R, Whitten B, Stecca K, Hazebroek J, Randhawa GS, Dolan M, Kinney
AJ, Tomes D, Nichols S, Anderson P (2004) Guar seed β-mannan synthase is a member of
the cellulose synthase super gene family. Science 303: 363-366
Liepman AH, Wilkerson CG, Keegstra K (2005) Expression of cellulose synthase-like (Csl)
genes in insect cells reveal that CslA family members encode mannan synthases.
Proceedings of the National Academy of Science USA 102: 2221-2226
33. Papers for seminars
Burton RA, Jobling SA, Harvey AJ, Shirley NJ, Mather DE, Bacic A, Fincher GB (2008)
The genetics and transcriptional profiles of the cellulose synthase-like HvCslF gene
family in barley. Plant Physiology 146: 1821-1833
Burton RA, Collins HM, Kibble NAJ, Smith JA, Shirley NJ, Jobling SA, Henderson M,
Singh RR, Pettolino F, Wilson SM, Bird AR, Topping DL, Bacic A, Fincher GB (2011) Over-
expression of specific HvCslF cellulose synthase-like genes in transgenic barley
increases the levels of cell wall (1,3;1,4)-β-D-glucans and alters their fine structure.
Plant Biotechnology Journal 9: 117-135
Davis J, Brandizzi F, Liepman A, Keegstra K (2010) Arabidopsis mannan synthase CSLA9
and glucan synthase CSLC4 have opposite orientations in the Golgi membrane. Plant
Journal 64: 1028-1037
Dwivany FM, Yulia D, Burton RA, Shirley NJ, Wilson SM, Fincher GB, Bacic A, Newbigin
E, Doblin MS (2009) The CELLULOSE-SYNTHASE LIKE C (CSLC) family of barley includes
members that are integral membrane proteins targeted to the plasma membrane.
Molecular Plant 2: 1025-1039
Liepman AH, Nain CJ, Willats WGT, Sorensen I, Roberts AW, Keegstra K (2007)
Functional genomic analysis supports conservation of function among cellulose-
synthase-like A gene family members and suggests diverse roles of mannans in plants.
Plant Phyiology 143: 1881-1893
34. Nemeth C, Freeman J, Jones HD, Sparks C, Pellny TK, Wilkinson MD, Dunwell J,
Andersson AAM, Åman P, Guillon F, Saulnier L, Mitchell RAC, Shewry PR (2010)
Down-regulation of the CSLF6 gene results in decreased (1,3;1,4)-β-D-glucan in
endosperm of wheat. Plant Physiology 152: 1209-1218
Tonooka T, Aokil E, Yoshioka T, Taketa S (2009) A novel mutant gene for (1-3, 1-4)-β-D-
glucanless grain on barley (Hordeum vulgare L.) chromosome 7H. Breeding Science
59: 47-54
van Erp H, Walton JD (2009) Regulation of the cellulose synthase-like gene family by
light in the maize mesocotyl. Planta 229: 885-897
Wang W, Wang L, Chen C, Xiong G, Tan X-Y, Yang K-Z, Wang Z-C, Zhou Y, Ye D, Chen L-Q
(2011) Arabidopsis CSLD1 and CSLD4 are required for cellulose deposition and
normal growth of pollen tubes. Journal of Experimental Botany (In press) (NB pdf
is available from journal website)
Yin L, Verhertbruggen Y, Oikawa A, Manisseri C, Knierim B, Prak L, Jensen JK, Knox JP,
Auer M, Willats WGT, Scheller HV (2011) The cooperative activities of CSLD2,
CSLD3, and CSLD5 are required for normal Arabidopsis development. Molecular
Plant (In press) (NB pdf is available from journal website)
Yin Y, Huang J, Xu Y (2009) The cellulose synthase superfamily in fully sequenced
plants and algae. BMC Plant Biology 9: 99
35. Reviews
(NB only parts of these reviews are about CSL genes and their products; you need to
read selectively)
Carpita NC (2011) Update on mechanisms of plant cell wall biosynthesis: how plants
make cellulose and other (1→4)-β-D-glycans. Plant Physiology 155: 171-184
Doblin MS, Pettolino F, Bacic A (2010) Plant cell walls: the skeleton of the plant world.
Functional Plant Biology 37: 357-381
Fincher GB (2009) Exploring the evolution of (1,3;1,4)-β-D-glucans in plant cell walls:
comparative genomics can help! Current Opinion in Plant Biology 12: 140-147
Fincher GB (2009) Revolutionary times in our understanding of cell wall biosynthesis
and remodeling in the grasses. Plant Physiology 149: 27-37
36. Other resource papers
Cutler S, Somerville C (1997) Cellulose synthesis: cloning in silico. Current Biology 7:
R108-R111
Harris PJ, Fincher GB (2009) Distribution, fine structure and function of (1,3;1,4)-β-
glucans in the grasses and other taxa. In: Bacic A, Fincher GB, Stone BA (eds)
Chemistry, biochemistry, and biology of (1→3)-β-glucans and related
polysaccharides. Academic Press, Elsevier Inc., San Diego, USA, pp 621-654
Hazen SP, Scott-Craig JS, Walton JD (2002) Cellulose synthase-like genes of rice. Plant
Phyiology 128: 336-340
Richmond TA, Somerville CR (2000) The cellulose synthase superfamily. Plant
Physiology 124: 495-498
Richmond TA, Somerville CR (2001) Integrative approaches to determining Csl
function. Plant Phyiology 47: 131-143
37.
38. Families of CSL genes
Family Arabidopsis Rice Function
A 9 9 Heteromannan
synthesis
B 6 - ?
C 5 6 Synthesis of xyloglucan
main chain
D 6 5 Cellulose synthesis in tip-
growing cells?
E 1 3 ?
F - 8 Synthesis of
(1→3)(1→4)-β-glucans
G 3 - ?
H - 2 Synthesis of
(1→3)(1→4)-β-glucans
39. • From genome sequence of Arabidopsis, Somerville identified
a family of genes of unknown function with sequence
similarity to cellulose synthase (Cutler & Somerville 1997;
Richmond & Somerville 2000): cellulose synthase-like (CSL)
genes (30 CSL + 10 CESA genes in Arabidopsis)
• Postulated they encode enzymes that synthesize non-
cellulosic polysaccharides.
• Six CSL gene subfamilies identified in Arabidopsis and
Somerville speculated each responsible for biosynthesis:
Callose
Xyloglucan
Heteroxylan
Homogalacturonan (HG)
Rhamnogalacturonan I Pectic polysaccharides
Rhamnogalacturonan II