The mosaic of ancestral karyotype blocks in the Sinapis alba L. genome

The organisation of the Sinapis alba genome, comprising 12 linkage groups (n = 12), was compared with the Brassicaceae ancestral karyotype (AK) genomic blocks previously described in other crucifer species. Most of the S. alba genome falls into conserved triplicated genomic blocks that closely match the AK-defined genomic blocks found in other crucifer species including the A, B, and C genomes of closely related Brassica species. In one instance, an S. alba linkage group (S05) was completely collinear with one AK chromosome (AK1), the first time this has been observed in a member of the Brassiceae tribe. However, as observed for other members of the Brassiceae tribe, ancestral genomic blocks were fragmented in the S. alba genome, supporting previously reported comparative chromosome painting describing rearrangements of the AK karyotype prior to the divergence of the Brassiceae from other crucifers. The presented data also refute previous phylogenetic reports that suggest S. alba was more closely related to Brassica nigra (B genome) than to B. rapa (A genome) and B. oleracea (C genome). A comparison of the S. alba and Arabidopsis thaliana genomes revealed many regions of conserved gene order, which will facilitate access to the rich genomic resources available in the model species A. thaliana for genetic research in the less well-resourced crop species S. alba.     

Myrosinase in Sinapis alba L.

Extracts from developing seeds, seedings, and different organsof mature plants of Sinapis alba L. were tested for the presenceof myrosinase. Three different patterns of isoenzymes were distinguishedby isoelectric focusing. One was found in developing seeds andseedlings, a second in mature leaves, stems, flowers, and podsand a third in roots. Sections of all the parts tested were examined for the presenceof myrosin cells by light microscopy. Developing seeds and seedlingsshowed a good correlation between the appearance of myrosinaseand the presence of myrosin cells. No myrosin cells were foundin root or flower parts although they showed myrosinase activity.     

Turgor Pressure and Phototropism in Sinapis alba L. Seedlings

Rich, T. C. G. and Tomos, A. D. 1988. Turgor pressure and phototropismin Sinapis alba L. seedlings.—J. exp. Bot 39: 291-299. Phototropic responses were studied in light-grown mustard hypocotyls.Phototropism was induced by adding 0.27 µmol m^–2s^–1 unilateral blue light to a background of low pressuresodium (SOX) lamp light. Curvatures of some 6° from thevertical were reached by 60 min, the curvature rate between20 min and 60 min being 0.14° min^–1. From the axialgrowth rate and tissue geometry the local growth rates of illuminatedand shaded sides of the hypocotyl were calculated to be 1.5and 4.5 µmin^–1 respectively. Turgor pressures ofexpanding cells in control plants and in the shaded and illuminatedsides of the blue light illuminated hypocotyls were measuredto be 0.40-0.55 MPa with a pressure probe. No changes in turgorpressure were observed on initiation of curvature. The decayof pressure in the cells of non-transpiring plants followingexcision indicated that the yield stress threshold of the tissuemay be as low as 0.1 MPa. These results indicate that the phototropicgrowth response in this tissue is not mediated by changes inturgor pressure. Key words: Sinapis alba L., phototropism, turgor pressure     

Dark Reversion of Phytochrome in Sinapis alba L

Phytochrome in Sinapis alba L. (white mustard) seedlings undergoes both decay and reversion after an exposure to red light. This is typical of other crucifers and of dicotyledons in general. In the presence of sodium azide, decay is inhibited, and reversion continues at about the same rate as in buffer alone. The reversion has been demonstrated both in cotyledon plus hypocotyl hook and in hypocotyl hook samples alone and is of the same order of magnitude in both. Contrary conclusions in the literature that there is no reversion in Sinapis are based on indirect measurements and are unjustified.     

Intergeneric Somatic Hybridization Between Brassica napus L. and Sinapis alba L.

Electrically induced protoplast fusion was used to produce somatic hybrids between Brassica napus L. and Sinapis alba L. Seven hybrids were obtained and verified by the simple sequence repeat and cleaved amplified polymorphic sequence analysis of the genefael, indicating that the characteristic bands from S. alba were present in the hybrids. The hybridity was also confirmed by chromosome number counting because the hybrids possessed 62 chromosomes, corresponding to the sum of fusion-parent chromosomes. Chromosome pairing at meiosis was predominantly normal, which led to high pollen fertility,ranging from 66% to 77%. All hybrids were grown to full maturity and could be fertilized and set seed after self-pollination or back-crosses with B. napus. The morphology of the hybrids resembled characteristics from both parental species. An analysis of the fatty acid composition in the seeds of F1 plants was conducted and the seeds were found to contain different amounts of erucic acid, ranging from 11.0% to 52.1%.     

Intergeneric Somatic Hybridization Between Brassica napus L. And Sinapis alba L.

Electrically induced protoplast fusion was used to produce somatic hybrids between Brassica napus L. and Sinapis alba L. Seven hybrids were obtained and verified by the simple sequence repeat and cleaved amplified polymorphic sequence analysis of the genefael, indicating that the characteristic bands from S. alba were present in the hybrids. The hybridity was also confirmed by chromosome number counting because the hybrids possessed 62 chromosomes, corresponding to the sum of fusion-parent chromosomes. Chromosome pairing at meiosis was predominantly normal, which led to high pollen fertility,ranging from 66% to 77%. All hybrids were grown to full maturity and could be fertilized and set seed after self-pollination or back-crosses with B. napus. The morphology of the hybrids resembled characteristics from both parental species. An analysis of the fatty acid composition in the seeds of F1 plants was conducted and the seeds were found to contain different amounts of erucic acid, ranging from 11.0% to 52.1%.     

Biosynthesis of wax esters in tissues of Sinapis alba L. seeds

The biosynthesis of wax esters has been investigated in maturing seeds of Sinapis alba. Exogenous long-chain alcohols are incorporated exclusively into alkyl moieties of wax esters. Oxidation of the long-chain alcohols is not detected. Exogenous fatty acids are incorporated into acyl moieties of wax esters to a low extent. A reduction of fatty acids to alcohols is not observed. Synthesis of wax esters is localized exclusively in the testa; both outer and inner integument are equally active in wax ester biosynthesis. The biosynthesis of wax esters is specific with regard to both chain length and degree of unsaturation of long-chain alcohols. Exogenous and endogenous sterols are not esterified.     

High irradiance response promotion of a subsequent light induction response in Sinapis alba L.

Relative quantum responsivity curves for inhibition of hypocotyl elongation in Sinapis alba L. seedlings previously grown in white light confirm that a marked end of day inhibition response can be induced by a monochromatic light treatment (30 min) at the end of the light period. In dark grown seedlings, however, no growth inhibition can be induced by a 30 min monochromatic light treatment. A prerequisite for an induction response appears to be a pretreatment with continuous light. Far red light is most effective with blue and red light showing a lesser effectiveness. The light pretreatment also shows a marked fluence rate dependency with respect to its ability to allow an induction response to manifest itself. The pretreatment required shows all the characteristics of a classical HIR response. The appearance of the effect in plants treated with the herbicide SAN 9789 seems to exclude chlorophyll as being the photoreceptor.Abbreviations SAN 9789 4-chloro-5-(methylamino)-2-(, , -trifluoro-m-tolyl)-3(2H)-pyridazinone - RG9 light long wavelength far red light (Schott RG9 colour glass) - FR far red light - WL white light - BL blue light - RL red light - D darkness - P_tot total phytochrome - P_fr far red absorbing form of phytochrome - HSR high irradiance response     

Bee visits and the nectar of Echium vulgare L. and Sinapis alba L.

Abstract. 1. The time-course of anthesis of Echium vulgare is described.
2. Diel changes in the sugar concentration of the nectar, the quantity of nectar and the quantity of sugar per flower are illustrated for E.vulgare and for Sinapis alba.
3. These changes are interpreted in terms of (a) the periodicity of secretion and (b) the influence of microclimate and insect visits on post-secretory changes in the composition and volume of nectar.
4. There was hour-to-hour and day-to-day variation in the species composition and the proportion of workers taking nectar rather than nectar plus pollen among the social bees visiting E.vulgare.
5. Honeybee visits to E.vulgare were more numerous in humid weather, when there was enough nectar per flower for their relatively short tongues to reach, and in an area where the corollas grew shorter than they did elsewhere.
6. The significance of changes in the caloric content, volume, concentration, viscosity and sugar composition is discussed from the points of view of insects and ecologists.     

Differentiation of a Functional Aleurone Layer Within the Seed Coat of Sinapis alba L.

The hitherto unresolved ontogenetic origin of the aleurone layerin mustard (Sinapis alba L.) seeds was investigated with lightand electron microscopy. Contrary to previous views, this layerof storage cells is neither derived from the endosperm nor fromthe nucellus, but from a particular cell layer within the innerintegument of the seed coat. These cells differentiate and becomefilled with storage protein and fat concurrently with the maturationof the embryo. They survive seed desiccation and become depletedof storage materials during seed germination. Temporally correlatedwith the germinating embryo, the aleurone cells produce microbodyenzymes, which are controlled by light in a similar fashionin both tissues. Sinapis alba L., mustard, aleurone layer, seed coat, seed formation, germination     

Markierungsmuster nach H3-Thymidineinbau in Kernen der Hypokotylzellen von Sinapis alba L.

DNA-synthesis in the hypocotyls of Sinapis alba L. was studied with H³-thymidine labelling. Cells from hypocotyl segments were stained by the Feulgen-method and squash preparations were made. The following labelling patterns were observed: 1. Labelling of the chromocentres only. 2. Nuclear area evenly labelled. 3. No radioactivity in the chromocentres. This pattern was rarely seen. — The frequency of the first two types in different tissue segments is not equal. In segments with more differentiated cells there was an increase in the percentage of nuclei with radioactivity only in the chromocentres. This could be due to a prolongation of the phase of synthesis in the chromocentres in this tissue. — The total number of labelled nuclei decreases basipetally as well as with the age of the hypocotyl. In hypocotyls of seedlings older than 52 hrs radioactivity appeared only sporadically in the nuclei. The decrease in the number of labelled nuclei is faster than the decline of the corresponding measurable total DNA synthesis in the hypocotyl. This can either be due to extra nuclear DNA synthesis or depend on an increase in DNA synthesis in the later replicating heterochromatic region of the nucleus.     

The appearance of competence for phytochrome-mediated anthocyanin synthesis in the cotyledons of Sinapis alba L.

Summary It is demonstrated that phytochrome-mediated anthocyanin synthesis in the epidermal cells of mustard seedling cotyledons takes place only 27 h after sowing onwards (at 25°C). This starting point cannot be shifted by light treatments or by nutrients. The late appearance of competence for P _fr (P _r and P _fr, red- and far-red absorbing forms of phytochrome, respectively) with regard to anthocyanin synthesis is not related to the phytochrome system per se (P _rP _fr) as this is fully functional immediately after sowing of the seed; nor is it related to the primary reaction of phytochrome: P _fr+XP _fr XP _fr X (X, X, two forms of a receptor for P _fr) or to the initial action of P _fr X:P _fr X+KY (K, coupling element, leading to the product Y, which is no longer photoreversible). Rather, the starting point is determined by internal factors only and is thus not accessible to any specific control by external factors. On the other hand, however, the beginning of the initial action of P _fr X (coupling point) can be shifted by light via phytochrome under high irradiance conditions. Moreover, it is shown that there is no phytochrome-independent effect of blue light on photomorphogenesis in the young mustard seedling and that there is no rapid dark reversion of P _fr which can be detected by physiological means, at least duringAbbreviations P _r red-absorbing forms of phytochrome - P _fr far-red-absorbing forms of phytochrome - P ₁ total spectrophotometrically detectable phytochrome - HS Hoagland's nutrient solution - HIR high irradiance response     

Formation of protein storage bodies during embryogenesis in cotyledons of Sinapis alba L.

An electron microscopic investigation of fine structural changes in post-meristematic cotyledon mesophyll cells during the period of storage protein accumulation (16–32 d after pollination) showed that the rough ER, the Golgi apparatus and the developing vacuome are intimately involved in the formation of storage protein bodies (aleurone bodies). At the onset of storage protein accumulation (16–18 d after pollination) storage protein-like material appears within Golgi vesicles and preformed vacuoles. At a later stage (24 d after pollination) similar material can also be detected within vesicles formed directly by the rough endoplasmic reticulum (ER). It is concluded that there are two routes for storage protein transport from its site of synthesis at the ER to its site of accumulation in the vacuome. The first route involves the participation of dictyosomes while the second route bypasses the Golgi apparatus. It appears that the normal pathways of membrane flow in the development of central vacuoles in post-meristematic cells are used to deposit the storage protein within the protein bodies. Thus, the protein body can be regarded as a transient stage in the process of vacuome development of these storage cells.Abbreviation ER endoplasmic reticulum     

Specificity of induction responses in Sinapis alba L.: Plant growth and development

Plant defenses are expected to be negatively correlated with plant growth, development and reproduction. In a recent study, we investigated the specificity of induction responses of chemical defenses in the Brassicaceae Sinapis alba.¹ It was shown that glucosinolate levels and myrosinase activities increased to different degrees after 24-hours-feeding by a specialist or generalist herbivore or mechanical wounding. Here, we present the specific influences of these treatments on organ biomasses which were recorded as a measure of growth. Directly after the treatments, organ biomasses were reduced locally and systemically by herbivore feeding, but not by mechanical wounding compared to control plants. Induction of glucosinolates, which increased in all treatments, is thus not necessarily expressed as cost in terms of reduced growth in S. alba. No significant long-term differences in plant development between herbivore treated and control plants were found. Thus, tissue loss and increased investments in chemical defenses could be compensated over time, but compensation patterns depended on the inducing agent. Furthermore, herbivore treatments resulted in an increased mechanical defense, measured as abaxial trichome densities. Plants respond highly dynamic with regard to defense and growth allocation and due to different inductors.Key words: Brassicaceae, organ biomass, plant development, specialist, generalist, herbivore, mechanical wounding, costs, trichome densityPlant defenses are generally thought to impose costs in relation to growth and fitness.² The ability to increase defense levels only after herbivory, i.e., induction, is one possible mechanism of lowering these allocation costs.³ In Brassicaceae, the glucosinolate-myrosinase system is known to hold a defensive function.⁴ The constitutive and induced production of glucosinolates and myrosinases is thought to be connected to allocation and ecological costs.^2,5In a recent study, we investigated the specificity of short-term induction patterns of chemical defenses in Sinapis alba L. var. Silenda damaged by a glucosinolate-sequestering specialist herbivore (turnip sawfly, Athalia rosae (L.), Hymenoptera), a generalist herbivore (fall armyworm, Spodoptera frugiperda J. E. Smith, Lepidoptera) or mechanical wounding (cork borer).¹ Feeding by the specialist as well as mechanical wounding led to 3-fold increases in both glucosinolate- and myrosinase-levels, whereas generalist feeding induced up to 2-fold increases in glucosinolates only.Different strengths of plant chemical responses might be mirrored in differences of subsequent fitness-related parameters of the plants.⁶ To assess short-term effects within 24 hours of induction on organ growth in S. alba, organ dry biomasses were calculated from the previous plant set.¹ Water content was determined of the organ halves which were freeze-dried and analyzed for glucosinolate content¹ and organ dry weights were calculated from water content and total organ fresh weight. The percentage of removed tissue area was determined by photo analysis and organ dry weights of treated leaves were corrected for the respective area. The percentage of lost area in damaged leaves was 7.9 ± 0.5 % after mechanical wounding, 15.1 ± 2.3 % after feeding by S. frugiperda and 15.6 ± 2.3 % after feeding by A. rosae (mean values ± SE, n = 7–8). The plants'' habits and total number of leaves did not vary between the tested plant groups (Fig. 1B; ANOVA: f = 2.36, df = 3, p = 0.095).Open in a separate windowFigure 1Organ dry biomasses of leaves and stems (A) and total numbers of leaves (B) of Sinapis alba cv. Silenda directly after induction. The second youngest leaves of three weeks old plants were treated with either mechanical wounding (cork borer), one Spodoptera frugiperda caterpillar (third instar) or one larva of Athalia rosae (third instar) enclosed in a muslin bag for 24 hours. Bagged leaves without any further treatment served as controls (mean values ± SE, n = 6–8 per treatment). Letters above bars indicate significant differences (ANOVA, Tukey-HSD tests: p < 0.05; n.s., not significant). DL, damaged leaf; OL, older leaf; YL, younger leaf; OS, older stem; YS, younger stem.The short-term growth responses were highly specific between treatments. Herbivore damage did not only result in reduced organ biomass growth of the damaged leaf (ANOVA: f = 11.29, df = 3, p < 0.001), but also of adjacent tissues compared to organs from bag treated and mechanically wounded plants after 24 hours of treatment (Fig. 1A; older leaf - ANOVA: f = 3.87, df = 3, p = 0.021; younger leaf - ANOVA: f = 6.02, df = 3, p = 0.003; younger stem - ANOVA: f = 4.12, df = 3, p = 0.017). Significant differences from bag treated control plants were found for damaged and systemic younger leaves of plants treated with A. rosae larvae. Differences of organ dry biomasses between mechanically wounded and herbivore treated plants were more pronounced, with reduced growth in the latter of 15 to 36 % in leaves and 23 to 48 % in stem parts. This specificity in growth response could be brought about by elicitors introduced to the wounded plant tissues from the herbivores'' saliva which can influence C-allocation to roots.⁷ The reduced growth of organ biomasses observed in herbivore treated leaves could be the result of specifically saliva elicited resource allocation away from leaf tissue,⁸ and might not represent costs of increased chemical defense.Long-term effects of herbivore feeding on development of S. alba were monitored in a second set of plants which were treated (as described previously in ref. ¹) for 24 hours with either the specialist or the generalist, enclosed in a bag. About three weeks later, on the day when the first flower opened, several parameters were recorded (9^,10 Thereby, thresholds for damage seem to exist, beyond which no compensation of tissue loss is possible.¹¹ The percentages of damage in S. alba were, however, below the threshold values reported for other Brassicaceae.¹¹ Influences on growth rates can be obviously transitory. In Arabidopsis thaliana (L.) Heynh., reduced growth rates were observed directly after treatment, but later growth increased so much, that these plants overcompensated and were even larger than control plants.⁹ Such plastic plant responses can be again modified by elicitors.^7,12

Table 1

Developmental responses of 3-week-old Sinapis alba plants treated for 24 hours with either one larva of the specialist Athalia rosae or one caterpillar of the generalist Spodoptera frugiperda

Characterization of five new low-molecular-mass trypsin inhibitors from white mustard (Sinapis alba L.) seed.

Five new low-molecular-mass trypsin inhibitors belonging to the RTI/MTI-2 family were identified from white mustard (Sinapis alba L. ; MTI-2) seed. Purified MTI-2 consisted of a peptide mixture, displaying Ile or Arg at position 43, Trp or kynurenine (Kyn) at position 44, and C-terminal ragged ends. The occurrence of Ile or Arg at position 43 suggested that MTI-2 inhibitors originated from different genes. The presence of 5-oxo-proline (pyroglutamic acid; 5-oxoPro1) and Kyn44 reflected post-translational processing of the serine proteinase inhibitor. MTI-2 showed approximately 70% amino-acid identity with low-molecular-mass trypsin inhibitors isolated from oil rape (Brassica napus var. oleifera; RTI-III) seed and with serine proteinase inhibitors mapped in Arabidopsis thaliana chromosome II (ATTI). Furthermore, MTI-2 was homologous to brazzein, the sweet-tasting protein from Pentadiplandra brazzeana Baillon fruit ( approximately 30% amino-acid identity). Although snake-venom toxins showed a low amino-acid identity (< 20%) with MTI-2, RTI-III, and ATTI, some structurally relevant residues were conserved. The disulfide bridge pattern of MTI-2 (Cys5-Cys27, Cys18-Cys31, Cys42-Cys52, and Cys54-Cys57) corresponded to that of RTI-III and of snake-venom toxins, being different from that of brazzein. Therefore, protein similarity might be attributable to the three-dimensional arrangement rather than to the amino-acid sequence. Values of Ka for MTI-2 binding to bovine beta-trypsin (trypsin) and bovine alpha-chymotrypsin were 6.3 x 109 M-1 and 2.0 x 106 M-1, respectively, at pH 8.0 and 21.0 degrees C. Moreover, values of kon for MTI-2 binding to trypsin and of koff for the dissociation of the serine proteinase:inhibitor complex were 5.6 x 105 M-1.s-1 and 8.9 x 10-5 M-1.s-1, respectively, at pH 8.0 and 21.0 degrees C. Despite the heterogeneity of the purified inhibitor peptide mixture, the inhibition properties of the different MTI-2 inhibitors were indistinguishable.     

Specificity of sterol-glucosylating enzymes from Sinapis alba and Physarum polycephalum.

1. Sinapis alba L. seedlings contain glycosyltransferase catalyzing the synthesis of sterol glucosides in the presence of UDPglucose as sugar donor. The major activity occurs in the membranous fraction sedimenting at 300--9000 x g. Successive treatment of the particulate enzyme fraction with acetone and Triton X-100 affords a soluble glucosyltransferase preparation which can be partly purified by gel filtration on Sephadex G-150. Molecular weight of the glucosyltransferase is 1.4 . 10(5). Apparent Km values for UDPglucose and sitosterol are 8.0 . 10(-5) M and 5.0 . 10(-6) M, respectively. 2. Comparison was made of the S. alba glucosyltransferase with a similar sterol-glucosylating enzyme isolated from non-photosynthesizing organism Physarum polycephalum (Myxomycetes). UDPglucose was the most efficient glucose donor in both cases but the enzyme from Ph. polycephalum can also utilize CDPglucose and TDPglucose. Glucose acceptors are, in case of both enzymes, sterols containing a beta-OH group at C-3 and a planar ring system (5 alpha-H or double bond at C-5). The number and position of double bonds in the ring system and in the side chain, as well as the presence of additional alkyl groups in the side chain at C-24 are of secondary importance. 3. The present results indicate that both enzymes can be regarded as specific UDPglucose:sterol glucosyltransferases. Certain differences in their specificity towards donors and acceptors of the glucosyl moiety suggest, however, a different structure of the active sites in both enzymes.     

Phytochrome control of plastid mRNA in mustard (Sinapis alba L.)

The steady-state levels of plastid RNA sequences in dark-grown and light-grown mustard (Sinapis alba L.) seedlings have been compared. Total cellular RNAs were labeled in vitro with ³²P and hybridized to separated restriction fragments of plastid DNA. Cloned DNA fragments which encode the large subunit (LS) of ribulose-1,5-bisphosphate carboxylase [3-phospho-D-glycerate carboxylase (dimerizing), EC 4.1.1.39] and a 35,000 plastid polypeptide were used as probes to assess the levels of these two plastid mRNAs. The 1.22-kilobase-pair mRNA for the 35,000 polypeptide is almost undetectable in dark-grown seedlings, but is a major plastid mRNA in light-grown seedlings. The hybridization analysis of RNA from seedlings which were irradiated with red and far-red light indicates that the level of this mRNA, but not of LS mRNA, is controlled by phytochrome.Abbreviations LS large subunit - RuBP ribulose-1,5-bisphosphate - ptDNA plastid DNA