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Biomedical subjects

H Saedler

Publications and source records attributed to H Saedler.

At least 73 records · Page 4Linked to original sources
PubMed1993

Mobility of the maize transposable element En/Spm in Arabidopsis thaliana.

The autonomous element En-1 of the maize En/Spm transposable element system is capable of frequent somatic and germinal excision in the heterologous host Arabidopsis thaliana. The pattern of En-homologous transcripts generated in transgenic Arabidopsis resembles En transcription in maize. An excision reporter construct based on NPT-II gene (pKEn2) can be used reliably for the isolation of En-1 germinal revertants by seed germination on kanamycin-containing medium. Re-insertion after germinal excision is apparently frequent. A dSpm receptor element can be efficiently trans-activated in Arabidopsis either by En-1 or by expressing cDNAs of tnpA and tnpD. Excision and re-insertion of En/Spm take place with similar characteristics as in maize. This is the first description of En/Spm transposition in Arabidopsis and the parameters analysed here suggest that transposon tagging with En should be feasible in this species.

Arabidopsis↗
PubMed1992

Characterization of the Antirrhinum floral homeotic MADS-box gene deficiens: evidence for DNA binding and autoregulation of its persistent expression throughout flower development.

We have determined the structure of the floral homeotic deficiens (defA) gene whose mutants display sepaloid petals and carpelloid stamens, and have analysed its spatial and temporal expression pattern. In addition, several mutant alleles (morphoalleles) were studied. The results of these analyses define three functional domains of the DEF A protein and identify in the deficiens promoter a possible cis-acting binding site for a transcription factor which specifically upregulates expression of deficiens in petals and stamens. In vitro DNA binding studies show that DEF A binds to specific DNA motifs as a heterodimer, together with the protein product of the floral homeotic globosa gene, thus demonstrating that the protein encoded by deficiens is a DNA binding protein. Furthermore, Northern analysis of a temperature sensitive allele at permissive and non-permissive temperatures provides evidence for autoregulation of the persistent expression of deficiens throughout flower development. A possible mechanism of autoregulation is discussed.

Alleles↗
PubMed1992

Bracteomania, an inflorescence anomaly, is caused by the loss of function of the MADS-box gene squamosa in Antirrhinum majus.

Anomalous flowering of the Antirrhinum majus mutant squamosa (squa) is characterized by excessive formation of bracts and the production of relatively few and often malformed or incomplete flowers. To study the function of squamosa in the commitment of an inflorescence lateral meristem to floral development, the gene was cloned and its genomic structure, a well as that of four mutant alleles, was determined. SQUA is a member of a family of transcription factors which contain the MADS-box, a conserved DNA binding domain. In addition, we analysed the temporal and spatial expression pattern of the squa gene. Low transcriptional activity of squa is detectable in bracts and in the leaves immediately below the inflorescence. High squa transcript levels are seen in the inflorescence lateral meristems as soon as they are formed in the axils of bracts. Squa transcriptional activity persists through later stages of floral morphogenesis, with the exception of stamen differentiation. Although necessary for shaping a normal racemose inflorescence, the squa function is not absolutely essential for flower development. We discuss the function of the gene during flowering, its likely functional redundancy and its possible interaction with other genes participating in the genetic control of flower formation in Antirrhinum.

Alleles↗
PubMed1992

GLOBOSA: a homeotic gene which interacts with DEFICIENS in the control of Antirrhinum floral organogenesis.

GLOBOSA (GLO) is a homeotic gene whose mutants show sepaloid petals and carpelloid stamens. The similarity of Glo mutants to those of the DEFICIENS (DEFA) gene suggests that the two genes have comparable functions in floral morphogenesis. The GLO cDNA has been cloned by virtue of its homology to the MADS-box, a conserved DNA-binding domain also contained in the DEFA gene. We have determined the structure of the wild type GLO gene as well as of several glo mutant alleles which contain transposable element insertions responsible for somatic and germinal instability of Glo mutants. Analyses of the temporal and spatial expression patterns of the DEFA and GLO genes during development of wild type flowers and in flowers of various stable and unstable defA and glo alleles indicate independent induction of DEFA and GLO transcription. In contrast, organ-specific up-regulation of the two genes in petals and stamens depends on expression of both DEFA and GLO. In vitro DNA-binding studies were used to demonstrate that the DEFA and GLO proteins specifically bind, as a heterodimer, to motifs in the promoters of both genes. A model is presented which proposes both combinatorial and cross-regulatory interactions between the DEFA and GLO genes during petal and stamen organogenesis in the second and third whorls of the flower. The function of the two genes controlling determinate growth of the floral meristem is also discussed.

Alleles↗
PubMed1992

Endogenous and environmental factors influence 35S promoter methylation of a maize A1 gene construct in transgenic petunia and its colour phenotype.

30,000 transgenic petunia plants carrying a single copy of the maize A1 gene, encoding a dihydroflavonol reductase, which confers a salmon red flower colour phenotype on the petunia plant, were grown in a field test. During the growing season plants with flowers deviating from this salmon red colour, such as those showing white or variegated phenotypes and plants with flowers exhibiting only weak pigmentation were observed with varying frequencies. While four white flowering plants were shown at the molecular level to be mutants in which part of the A1 gene had been deleted, other white flowering plants, as well as 13 representative plants tested out of a total of 57 variegated individuals were not mutants but rather showed hypermethylation of the 35S promoter directing A1 gene expression. This was in contrast to the homogeneous fully red flowering plants in which no methylation of the 35S promoter was observed. While blossoms on plants flowering early in the season were predominantly red, later flowers on the same plants showed weaker coloration. Once again the reduction of the A1-specific phenotype correlated with the methylation of the 35S promoter. This variation in coloration seems to be dependent not only on exogenous but also on endogenous factors such as the age of the parental plant from which the seed was derived or the time at which crosses were made.

Alcohol Oxidoreductases↗
PubMed1991

Molecular analysis of tap2, an anther-specific gene from Antirrhinum majus.

Deficiens is a floral homeotic gene of Antirrhinum majus, mutation of which results in transformation of petals to sepals and stamens to carpels. In a search for putative target genes, controlled by this regulatory locus, cDNA clones representing genes, that are expressed in wild type but not in the deficiens mutant flowers, were isolated by differential screening. The molecular structure and the expression pattern of one of these genes, tap2, is described. Tap2 is transiently and tissue-specifically expressed in the tapetum of the anthers. It encodes a 131 amino-acids-long protein with a hydrophobic N-terminus, displaying all characteristic features of a signal peptide. This indicates that the TAP2 protein may be secreted.

Amino Acid Sequence↗
PubMed1991

The duplicated chalcone synthase genes C2 and Whp (white pollen) of Zea mays are independently regulated; evidence for translational control of Whp expression by the anthocyanin intensifying gene in.

Two chalcone synthase genes in maize have been cloned and molecularly characterized to be the C2 and the Whp (white pollen) locus. The two genes have highly homologous exon sequences but differ considerably in sequences 5' upstream and 3' downstream of the coding region, as well as in their introns. Northern and Western experiments of chalcone synthase expression in various tissues and in different genotypes indicated that C2 and Whp are differently regulated. The expression of Whp in maize aleurone is dependent on the presence of the recessive allele of the gene intensifier (in). The regulatory effect of in on Whp expression is not detectable at the transcriptional level, but seems to take place during translation.

Acyltransferases↗
PubMed1991

Molecular characterization of two stamen-specific genes, tap1 and fil1, that are expressed in the wild type, but not in the deficiens mutant of Antirrhinum majus.

Deficiens, a homeotic gene involved in the genetic control of flower development, codes for a putative transcription factor. Upon mutation of the gene, petals are transformed to sepals and stamens to carpels, indicating that deficiens is essential for the activation of genes required for petal and stamen formation. In a search for putative target genes of deficiens, several stamen- and petal-specific genes were cloned that are expressed in wild type but not in the deficiensglobifera mutant. In this report the molecular characterization of two of these genes, tap1 and fil1, is presented. They are transiently expressed during flower development. In situ hybridization data demonstrate that tap1 is expressed in the tapetum of the anthers and fil1 in the filament of the stamen and at the bases of the petals. Both genes encode small proteins with N-terminal hydrophobic domains suggesting that they are secreted. We discuss possible functions of the gene products and their relationship to the deficiens gene.

Amino Acid Sequence↗
PubMed1991

The transposable element Tam1 from Antirrhinum majus shows structural homology to the maize transposon En/Spm and has no sequence specificity of insertion.

We present the genomic structure of Tam1, a transposable element from Antirrhinum majus. The Tam1 element is 15.2 kb long and includes two genes that are transcribed to produce a 2.4 kb (tnp1) and a 5 kb mRNA (tnp2). These transcripts partially overlap and the exons are scattered over the whole element. Tnp1 encodes a 53 kDa protein as deduced from the cDNA sequence. The 5 kb transcript of tnp2 contains an open reading frame that shares 45% homology with part of the tnpD gene of En/Spm from maize and 48% homology with an open reading frame of the Tgm element from Glycine max. We discuss the possible functions of these genes by analogy with En/Spm. Additionally, a number of flanking sequences of Tam1 insertions were analysed to investigate the sequence specificity of insertion. From these studies we conclude that Tam1 transposes predominantly into AT-rich regions that can be unique as well as repetitive. No specific target sequence of insertion could be found.

Amino Acid Sequence↗
PubMed1991

Molecular analysis of the Ubiquitous (Uq) transposable element system of Zea mays.

The Uq transposable element of maize is the most widely dispersed among different maize populations and genetic testerstrains. Despite intensive genetic characterization, little is known about its molecular structure. In order to obtain information relevant to this topic, we have cloned and sequenced three ruq receptors. Surprisingly, they are all Ds1-like receptor types of the Ac-Ds transposon family. Based on our molecular data, we present a model to explain the functional differences associated with the differential expression of the Uq and Ac transposon systems.

Base Sequence↗
PubMed1990

En/Spm encoded tnpA protein requires a specific target sequence for suppression.

The En/Spm encoded suppressor function has been reconstituted in transgenic tobacco protoplasts. The suppressor affects genes which contain an En/Spm responsive transposable element in the transcribed sequences. The En/Spm encoded protein tnpA binds a defined cis element in the inserted transposon, repressing expression of the adjacent gene. This was shown by monitoring transient expression of a bacterial marker gene (GUS) expressed from a strong plant viral promoter. Suppressible variants of the marker gene were produced by inserting I element sequences into the untranslated sequences of the GUS transcript. Comparison of transient expression of these variants in wildtype tobacco protoplasts with their expression in protoplasts transgenic for tnpA protein demonstrates that tnpA is the suppressor. In addition, the minimal cis element required for suppression has been defined as a dimer consisting of two 12 bp tnpA binding sequences in a particular inverted orientation. One of these dimers occurs in each En/Spm end close to the characteristic 13 bp terminal inverted repeat. TnpA binding sites in different arrangements do not respond as well to tnpA. The implications of this observation are discussed. This system can be used to analyse tnpA-DNA interactions involved in gene regulation further.

Alleles↗
PubMed1990

The En/Spm transposable element of Zea mays contains splice sites at the termini generating a novel intron from a dSpm element in the A2 gene.

The A2 locus of Zea mays, identified as one of the genes affecting anthocyanin biosynthesis, was cloned using the transposable elements rcy and dSpm as gene tags. The A2 gene encodes a putative protein of 395 amino acids and is devoid of introns. Two a2-m1 alleles, containing dSpm insertions of different sizes, were characterized. The dSpm element from the original state allele has perfect termini and undergoes frequent transposition. The element from the class II state allele is no longer competent to transpose. It has retained the 13 bp terminal inverted repeat but has lost all subterminal sites at the 5' end, which are recognized by tnpA protein, the most abundant product of the En/Spm transposable element system. The relatively high A2 gene expression of one a2-m1 allele is due to removal of almost all dSpm sequences by splicing. The slightly altered A2 enzyme is still functional as shown by complementation of an a2 mutant with the corresponding cDNA. The 5' and 3' splice sites are constituted by the termini of the dSpm element; it therefore represents a novel intron of the A2 gene.

Alleles↗
PubMed1990

Excision of the En/Spm transposable element of Zea mays requires two element-encoded proteins.

An excision assay system for En/Spm was developed in transgenic tobacco. The characteristics of excision and integration are similar to the natural system of Zea mays. In this transgenic model system two En/Spm encoded trans-acting functions, TNPA and TNPD, are required for excision. A biochemical model for transposition is proposed that might also be applicable to other transposable elements.

Cloning, Molecular↗
PubMed1990

Molecular analysis of the C1-I allele from Zea mays: a dominant mutant of the regulatory C1 locus.

The C1 locus of Zea mays (maize) controls the expression of genes involved in anthocyanin biosynthesis in aleurone and scutellar tissue and encodes a protein with the features of a transcriptional activator. C1-I is a dominant negative mutant which inhibits pigment formation. The structure of the C1-I allele was determined by cloning and sequencing of this allele and of two distinct C1-I derived cDNAs. C1-I has two major and several minor sequence differences with respect to the wild-type C1 allele. Transcription initiation occurs at the same position as in wild-type but transcription yields two different products, one major RNA of 1.3 kb and one minor RNA of 1.45 kb in length, encoding two proteins of 252 and 108 amino acids respectively. The longer 252 amino acid C1-I protein differs from the 273 amino acid wild-type C1 protein at several positions but most prominently at its carboxy terminus, resulting in reduced acidity of the C1-I protein. A similar change in acidity of the Gal4 protein of yeast converted this transcriptional activator into a repressor protein. We discuss the dominant phenotype of C1-I with respect to its possible repressor function in contrast to the activator function of the C1 gene product.

Alleles↗
PubMed1990

Deficiens, a homeotic gene involved in the control of flower morphogenesis in Antirrhinum majus: the protein shows homology to transcription factors.

Deficiens (defA+) is a homeotic gene involved in the genetic control of Antirrhinum majus flower development. Mutation of this gene (defA-1) causes homeotic transformation of petals into sepals and of stamina into carpels in flowers displaying the 'globifera' phenotype, as shown by cross sections and scanning electronmicroscopy of developing flowers. A cDNA derived from the wild type defA+ gene has been cloned by differential screening of a subtracted 'flower specific' cDNA library. The identity of this cDNA with the defA+ gene product has been confirmed by utilizing the somatic and germinal instability of defA-1 mutants. According to Northern blot analyses the defA+ gene is expressed in flowers but not in leaves, and its expression is nearly constant during all stages of flower development. The 1.1 kb long mRNA has a 681 bp long open reading frame that can code for a putative protein of 227 amino acids (mol. wt 26.2 kd). At its N-terminus the DEF A protein reveals homology to a conserved domain of the regulatory proteins SRF (activating c-fos) in mammals and GRM/PRTF (regulating mating type) in yeast. We discuss the structure and the possible function of the DEF A protein in the control of floral organogenesis.

Amino Acid Sequence↗
PubMed1990

Epigenetic changes in the expression of the maize A1 gene in Petunia hybrida: role of numbers of integrated gene copies and state of methylation.

The Petunia hybrida mutant RL01 is white flowering due to a genetic block in the anthocyanin pathway. The introduction of the maize A1 cDNA under the control of the CaMV 35S RNA promoter leads to the production of pelargonidin derivatives, resulting in a brick red flower phenotype. Among the transgenic petunia plants the pigmentation of the petals exhibited different expression patterns which could be categorized into the 'red', the 'variegated', and the 'white' phenotype. This system proved to be especially suitable for the investigation of gene expression by simply looking at the pigmentation pattern of the petals. The uniformity of floral pelargonidin pigmentation is inversely correlated with the number of integrated A1 copies. Furthermore, a correlation was found between the methylation status of the 35S RNA promoter and the instability of the floral pelargonidin coloration. The status of promoter methylation controlling the expression of the A1 gene seems to be influenced by the copy number and the chromosomal position of the transferred gene.

Anthocyanins↗