Wheat-Embryo Ribonucleates. I. Subcellular Localization of a Satellite Polyribonucleotide

1973 ◽  
Vol 51 (5) ◽  
pp. 606-612 ◽  
Author(s):  
A. A. Azad ◽  
B. G. Lane
Keyword(s):  
Microsomal Fraction ◽  
Large Subunit ◽  
Satellite Rna ◽  
Selective Precipitation ◽  
Ribosomal Subunits ◽  
Wheat Embryo ◽  
Wheat Embryos ◽  
The Subject ◽  
Rna Complex ◽  
Electrophoretic Component

(1) When wheat embryos are extracted with aqueous phenol and the aqueous phase is made 2.5 M with respect to NaCl at 0 °C, there is selective precipitation of about 80% of the total RNA. The 18 S and 26 S RNA species from the wheat-embryo ribosomes comprise a preponderant mass fraction (ca. 80%) of this NaCl-insoluble RNA (iRNA). A small amount of a rapidly migrating electrophoretic component (iRMEC) can be released by aqueous denaturation of wheat-embryo NaCl-insoluble RNA and because it is specifically complexed with 26 S RNA, the iRMEC component has been termed a "satellite" of 26 S RNA.(2) The wheat-embryo satellite RNA has been shown to be present in the microsomal fraction recovered from cell-free homogenates of wheat embryos.(3) The wheat-embryo satellite RNA has been shown to be differentially localized in the large subunit of wheat-embryo ribosomes where it presumably exists as part of the same intermolecular 26 S RNA complex that can be isolated by directly extracting the whole embryos with aqueous phenol.(4) During preparation of the ribosomal subunits, there is substantial degradation of the component ribonucleates and the nature of this degradation is the subject of a brief discussion.

Wheat embryo ribonucleates. XIV. Mass isolation of mRNA from wheat germ and comparison of its translational capacity with that of mRNA from imbibing wheat embryos

1979 ◽  
Vol 57 (9) ◽  
pp. 1170-1175 ◽  
Author(s):  
A. C. Cuming ◽  
T. D. Kennedy ◽  
B. G. Lane
Keyword(s):  
Wheat Germ ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sucrose Density Gradient ◽  
Wheat Embryo ◽  
Molecular Weights ◽  
Wheat Embryos ◽  
The Subject ◽  
Mass Isolation ◽  
Gradient Fractionation ◽  
Rabbit Reticulocytes

Commercially milled wheat germ is shown to be a convenient source material for facile recovery of mass (milligram) quantities of highly purified poly(A)-rich RNA. This poly(A)-rich RNA is efficiently translated in a nuclease-treated extract of rabbit reticulocytes. By sucrose density gradient fractionation of bulk poly(A)-rich RNA from wheat germ, it has been possible to show that there is a direct relationship between the molecular weights of the polypeptide products of cell-free synthesis and the molecular weights of the wheat mRNA molecules which program their synthesis. As assessed by SDS – polyacrylamide gel electrophoresis, the same array of polypeptides is synthesized when nuclease-treated reticulocyte extract is programmed by poly(A)-rich RNA from either commercially supplied or laboratory-prepared wheat embryos. Significantly, there are gross quantitative if not qualitative differences between the translational capacities of poly(A)-rich RNA from dry and imbibing wheat embryos, and the possible importance of these differences for interpreting a changing pattern of polypeptide synthesis in imbibing wheat embryos is the subject of a brief discussion.

Wheat embryo ribonucleates. VIII. The presence of 7-methylguanosine 'cap structures' in the RNA of imbibing wheat embryos

1977 ◽  
Vol 55 (8) ◽  
pp. 819-824 ◽  
Author(s):  
M. S. Saini ◽  
B. G. Lane
Keyword(s):  
Ribosomal Rna ◽  
Messenger Rna ◽  
Large Fraction ◽  
Deae Cellulose ◽  
Careful Analysis ◽  
Significant Fraction ◽  
Unbound Fraction ◽  
Wheat Embryo ◽  
Wheat Embryos ◽  
The Subject

1. By imbibing wheat embryos in media that contain methyl-labelled methionine, it is possible to label both terminal and nonterminal 7-methylguanosine constituents in NaCl-insoluble (2.5 M, 0 °C) RNA (iRNA).2. Most of the 7-[Me-14C]methylguanosine in wheat embryo i[Me-14C]RNA is present in nonterminal positions of polynucleotide chains, probably in ribosomal RNA.3. By passage through a column of oligo-dT-cellulose, it is possible to show that most of the 7-[Me-3H]methylguanosine in a 'bound' fraction of i[Me-3H]RNA from imbibing wheat embryos is present in terminal 'cap' structures, probably in messenger RNA.4. Although most of the 7-[Me-3H]methylguanosine in the 'unbound' (to oligo-dT-cellulose) fraction of i[Me-3H]RNA was present in nonterminal positions, there was also a highly significant fraction of 7-[Me-3H]methylguanosine in terminal 'cap' structures. Although it will be a subject of continued investigation, possible reasons why a large fraction of the total 7-[Me-3H]-methylguanosine was present in the 'unbound' fraction, in this present study, are a subject of discussion.5. Careful analysis failed to reveal the presence of any N6,O2′-di[Me-3H]methyladenosine in the 'unbound' fraction of i[Me-3H]RNA.6. Factors that might influence the binding of 'cap' oligonucleotides to DEAE-cellulose are the subject of a brief discussion.

Chain Termini of the Satellite RNA from Yeast Ribosomes

1973 ◽  
Vol 51 (5) ◽  
pp. 520-528 ◽  
Author(s):  
K. M. Oakden ◽  
B. G. Lane
Keyword(s):  
Group Analysis ◽  
Formal Structure ◽  
Yeast Cells ◽  
Satellite Rna ◽  
Dominant Form ◽  
Selective Precipitation ◽  
Specific Complex ◽  
Yeast Ribosomes ◽  
Electrophoretic Component ◽  
End Group

(1) When yeast cells are extracted with aqueous phenol and the aqueous phase is made 2.5 M with respect to NaCl at 0°, there is selective precipitation of about 80% of the total RNA. The 17 S and 26 S RNA from yeast ribosomes comprise a preponderant mass-fraction (ca. 90%) of this NaCl-insoluble RNA. A small amount (ca. 3%) of a rapidly migrating electrophoretic component (iRMEC) can be released by aqueous denaturation of yeast NaCl-insoluble RNA (iRNA), and because it forms a specific complex with 26 S RNA, this iRMEC component can be appropriately described as a "satellite" of 26 S RNA.(2) Following its release by aqueous denaturation of yeast NaCl-insoluble RNA, the satellite RNA has been subjected to end-group analysis, and it has been found to have a formal structure that is based on a repeating 5′-mononucleotide unit, i.e. (pN)n, where n = 160–200. On the basis of an analysis of its principal termini, the dominant form of the satellite RNA is[Formula: see text](3) In an allied study, a "rapidly labelled" fraction of yeast NaCl-insoluble RNA has been subjected to aqueous denaturation and end-group analysis.

A Possible Role for 5 S rRNA as a Bridge Between Ribosomal Subunits

1973 ◽  
Vol 51 (12) ◽  
pp. 1669-1672 ◽  
Author(s):  
A. A. Azad ◽  
B. G. Lane
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Large Subunit ◽  
Ribosomal Subunit ◽  
Small Subunit ◽  
Low Molecular Weight ◽  
Large Ribosomal Subunit ◽  
Ribosomal Subunits ◽  
Wheat Embryo ◽  
Present Context

18 S rRNA is a high molecular weight polyribonucleotide found in the small subunit, and 26 S rRNA is a high molecular weight polyribonucleotide found in the large subunit, whereas 5 S rRNA and 5.8 S rRNA are low molecular weight ("satellite") polyribonucleotides confined to the large subunit of wheat-embryo ribosomes. Under the same conditions in which 5.8 S rRNA is known to complex efficiently and preferentially with 26 S rRNA, it has been observed that 5 S rRNA complexes efficiently and preferentially with 18 S rRNA. Since 5 S rRNA is a component of the large ribosomal subunit, but it complexes preferentially with 18 S rRNA, which is a component of the small ribosomal subunit, it has been proposed that 5 S rRNA may serve as a "bridge" to mediate reversible association between the small and large ribosomal subunits. The possible role that a polycistronic precursor of rRNA might be visualized to play in the biogenesis and assembly of reversibly associating ribosomal subunits is alluded to in the present context.

Wheat-Embryo Ribonucleates. IV. Factors That Influence the Formation and Stability of a Complex Between 5S rRNA and 18S rRNA

1975 ◽  
Vol 53 (3) ◽  
pp. 320-327 ◽  
Author(s):  
A. A. Azad ◽  
B. G. Lane
Keyword(s):  
18S Rrna ◽  
5S Rrna ◽  
23S Rrna ◽  
Wheat Embryo ◽  
Physiological Importance ◽  
5.8S Rrna ◽  
Wheat Embryos ◽  
The Subject ◽  
26S Rrna ◽  
Source Materials

Under the conditions used in this study, wheat-embryo 5S rRNA complexes with its homologous 18S rRNA from wheat embryos and with heterologous 18S rRNA from other eukaryotic source materials such as yeast, L cells, and HeLa cells, but it does not complex with heterologous 16S rRNA from a prokaryote such as Escherichia coli or with homologous or heterologous 26S(23S) rRNA of either eukaryotic or prokaryotic origin.If a solution of wheat-embryo rRNA is simply made 0.3 M with respect to NaCl and then heated at 60 °C for 3 min before quick cooling to room temperature (ca. 20 °C), there is both preferential and efficient complex formation between 5S and 18S rRNA and between 5.8S and 26S rRNA.The laboratory-prepared' complex between wheat-embryo 5S rRNA and its homologous 18S rRNA is more thermostable in 0.1 M NaCl solution than is the 'natural' complex between wheat-embryo 5.8S rRNA and its homologous 26S rRNA, and both complexes 'melt' over a narrow range of temperature.The possible physicochemical and physiological importance of both homologous and heterologous rRNA complexes is the subject of a brief discussion.

Synthesis and turnover of proteins and mRNA in germinating wheat embryos

1982 ◽  
Vol 60 (3) ◽  
pp. 389-397 ◽  
Author(s):  
Zbyszko F. Grzelczak ◽  
Mark H. Sattolo ◽  
Linda K. Hanley-Bowdoin ◽  
Theresa D. Kennedy ◽  
Byron G. Lane
Keyword(s):  
Growth Phase ◽  
Time Course ◽  
Phase 1 ◽  
Major Product ◽  
Lag Phase ◽  
Wheat Embryo ◽  
Phase Development ◽  
Wheat Embryos

The most prominent methionine-labeled protein made when cell-free systems are programmed with bulk mRNA from dry wheat embryos has been identified with what may be the most abundant protein in dry wheat embryos. The protein has been brought to purity and has a distinctive amino acid composition, Gly and Glx accounting for almost 40% of the total amino acids. Designated E because of its conspicuous association with early imbibition of dry wheat embryos, the protein and its mRNA are abundant during the "early" phase (0–1 h) of postimbibition development, and easily detected during "lag" phase (1–5 h), but they are almost totally degraded soon after entry into the "growth" phase of development, by about 10 h postimbibition.The most prominent methionine-labeled protein peculiar to the cell-free translational capacity of bulk mRNA from "growth" phase embryos is not detected as a product of in vivo synthesis. Its electrophoretic properties and its time course of emergence, after 5 h postimbibition development, suggest that this major product of cell-free synthesis may be an in vitro counterpart to a prominent methionine-labeled protein made only in vivo, by "growth" phase embryos. Designated G because of its conspicuous association with "growth" phase development, the cell-free product does not comigrate with any prominent dye-stained band in electrophoretic distributions of wheat proteins. The suspected cellular counterpart to G, also, does not comigrate with a prominent dye-stained wheat protein during electrophoresis, and although found in particulate as well as soluble fractions of wheat embryo homogenates it is not concentrated in either nuclei or mitochondria, as isolated.

scholarly journals Genetics of in vitro organogenesis and precocious germination of wheat embryos

1998 ◽  
Vol 21 (1) ◽  
pp. 87-92 ◽  
Author(s):  
Claudia E. Lange ◽  
Luiz C. Federizzi ◽  
Fernando I.F. Carvalho ◽  
Ana L.C. Dornelles ◽  
Cristine L. Handel
Keyword(s):  
Gene Action ◽  
Precocious Germination ◽  
Genetic Determination ◽  
Wheat Embryo ◽  
In Vitro Organogenesis ◽  
Generation Means ◽  
Generation Means Analysis ◽  
Wheat Embryos ◽  
Segregating Population

The genetic bases of in vitro organogenesis and precocious germination of embryos in immature wheat embryo culture were investigated using six Brazilian genotypes and their F1, F2, BC1F1 and BC2F1 generations in a generation means analysis. Four parents and one set of F1’s were also analyzed in a diallel experiment. The results indicated a complex gene action controlling both traits, with additive, dominant and epistatic effects. High broad sense heritability values were found, indicating genetic determination. Considering the complexity of gene control, genetic gain could be achieved by selecting for the traits in advanced generations of the segregating population. Low correlation values between organogenesis, precocious germination, regeneration and somatic embryogenesis (data shown in a previous report) indicated the possibility of obtaining recombinant genotypes.

scholarly journals The Nuclear Export Receptors TbMex67 and TbMtr2 Are Required for Ribosome Biogenesis in Trypanosoma brucei

mSphere ◽  
2019 ◽  
Vol 4 (4) ◽  
Author(s):  
Constance Rink ◽  
Martin Ciganda ◽  
Noreen Williams
Keyword(s):  
Trypanosoma Brucei ◽  
Nuclear Export ◽  
Ribosome Biogenesis ◽  
Biological Process ◽  
Large Subunit ◽  
Critical Role ◽  
Ribosomal Subunits ◽  
Content Type ◽  
Protein Components ◽  
Export Receptors

ABSTRACT Ribosomal maturation is a complex and highly conserved biological process involving migration of a continuously changing RNP across multiple cellular compartments. A critical point in this process is the translocation of individual ribosomal subunits (60S and 40S) from the nucleus to the cytoplasm, and a number of export factors participate in this process. In this study, we characterize the functional role of the auxiliary export receptors TbMex67 and TbMtr2 in ribosome biogenesis in the parasite Trypanosoma brucei. We demonstrate that depletion of each of these proteins dramatically impacts the steady-state levels of other proteins involved in ribosome biogenesis, including the trypanosome-specific factors P34 and P37. In addition, we observe that the loss of TbMex67 or TbMtr2 leads to aberrant ribosome formation, rRNA processing, and polysome formation. Although the TbMex67-TbMtr2 heterodimer is structurally distinct from Mex67-Mtr2 complexes previously studied, our data show that they retain a conserved function in ribosome biogenesis. IMPORTANCE The nuclear export of ribosomal subunits (60S and 40S) depends in part on the activity of the essential auxiliary export receptors TbMtr2 and TbMex67. When these proteins are individually depleted from the medically and agriculturally significant parasite Trypanosoma brucei, distinct alterations in the processing of the rRNAs of the large subunit (60S) are observed as well as aberrations in the assembly of functional ribosomes (polysomes). We also established that TbMex67 and TbMtr2 interact directly or indirectly with the protein components of the 5S RNP, including the trypanosome-specific P34/P37. The critical role that TbMex67 and TbMtr2 play in this essential biological process together with their parasite-specific interactions may provide new therapeutic targets against this important parasite.

"Cap" sequences is poly(A)-rich RNA from dry wheat embryos

1981 ◽  
Vol 59 (10) ◽  
pp. 868-870 ◽  
Author(s):  
Byron G. Lane
Keyword(s):  
High Performance Liquid Chromatography ◽  
Liquid Chromatography ◽  
Messenger Rna ◽  
High Performance ◽  
Higher Plants ◽  
Rna Molecules ◽  
Wheat Embryos ◽  
The Subject ◽  
Widespread Interest

Although template-active RNA in dry seeds and embryos has attracted widespread interest, there have been no published reports about 5′-terminai "capping" sequences in such RNA. Boro[3H]hydride labeling of periodate-oxidized termini and high performance liquid chromatography of cap oligonucleotides have been used to compare terminal sequences in poly(A)-rich RNA from dry and germinating embryos. As is the case in germinating embryos, poly(A)-rich RNA from dry embryos contains only "type 0" cap sequences, i.e., m7G(5′)ppp(5′)N, in which m7G is the 7-methylguanosine cap and N is any of the classical ribonucleosides: adenosine (A), guanosine (G), cytidine (C), and uridine (U). Striking differences between the cell-free translational capacities of bulk messenger RNA (mRNA) populations from dry and germinating embryos are not reflected in signal differences in their proportions of "type 0" cap structures: in general, there is approximately 40% m7G(5′)ppp(5′)A, with roughly equivalent amounts of m7G(5′)ppp(5′)G and m7G(5′)ppp(5′)C accounting for most of the remaining sequences. The findings with mRNA from dry plant embryos serve to emphasize interesting differences between patterns of methylation in the capped and uncapped RNA molecules in higher plants and animals; these differences have not been previously noted in the literature and are the subject of brief comment in this paper.

scholarly journals The proteins of Xenopus ovary ribosomes

1971 ◽  
Vol 125 (4) ◽  
pp. 1091-1107 ◽  
Author(s):  
P J Ford
Keyword(s):  
Potassium Chloride ◽  
Ribosomal Proteins ◽  
Large Subunit ◽  
Buoyant Density ◽  
Small Subunit ◽  
Chemical Methods ◽  
Ribosomal Subunits ◽  
Molecular Weights ◽  
Caesium Chloride ◽  
Chloride Treatment

1. The preparation of ribosomes and ribosomal subunits from Xenopus ovary is described. 2. The yield of once-washed ribosomes (buoyant density in caesium chloride 1.601g·cm-3; 44% RNA, 56% protein by chemical methods) was 10.1mg/g wet wt. of tissue. 3. Buoyant density in caesium chloride and RNA/protein ratios by chemical methods have been determined for ribosome subunits produced by 1.0mm-EDTA or 0.5m-potassium chloride treatment and also for EDTA subunits extracted with 0.5m-, 1.0m- or 1.5m-potassium chloride, 4. Analysis of ribosomal protein on acrylamide gels at pH4.5 in 6m-urea reveals 24 and 26 bands from small and large EDTA subunits respectively. The actual numbers of proteins are greater than this, as many bands are obviously doublets. 5. Analysis of the proteins in the potassium chloride extract and particle fractions showed that some bands are completely and some partially extracted. Taking partial extraction as an indication of possible doublet bands it was found that there were 12 and 20 such bands in the small and large subunits respectively, making totals of 36 and 46 proteins. 6. From the measured protein contents and assuming weight-average molecular weights for the proteins of large and small subunits close to those observed for eukaryote ribosomal proteins it is possible to compute the total numbers of protein molecules per particle. It appears that too few protein bands have been identified on acrylamide gels to account for all the protein in the large subunit, but probably enough for the small subunit.

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