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J L Riechmann

Publications and source records attributed to J L Riechmann.

At least 19 recordsLinked to original sources
PubMed2001

Regulation of flowering in Arabidopsis by an FLC homologue.

The Arabidopsis FLC gene encodes a MADS domain protein that acts as a repressor of flowering. Late-flowering vernalization-responsive ecotypes and mutants have high steady-state levels of FLC transcript, which decrease during the promotion of flowering by vernalization. Therefore, FLC has a central role in regulating the response to vernalization. We have isolated an Arabidopsis gene, MAF1, which encodes a protein that is closely related to FLC. Overexpression studies demonstrate that MAF1 produces comparable effects to FLC, and likely has a similar function in the regulation of flowering. In contrast to FLC, however, MAF1 expression shows a less clear correlation with the vernalization response. In addition, MAF1 overexpression does not influence FLC transcript levels. Thus, MAF1 likely acts downstream or independently of FLC transcription. We further report identification of a cluster of four additional FLC-like genes in the Arabidopsis genome.

Amino Acid Sequence↗
PubMed2000

Arabidopsis transcription factors: genome-wide comparative analysis among eukaryotes.

The completion of the Arabidopsis thaliana genome sequence allows a comparative analysis of transcriptional regulators across the three eukaryotic kingdoms. Arabidopsis dedicates over 5% of its genome to code for more than 1500 transcription factors, about 45% of which are from families specific to plants. Arabidopsis transcription factors that belong to families common to all eukaryotes do not share significant similarity with those of the other kingdoms beyond the conserved DNA binding domains, many of which have been arranged in combinations specific to each lineage. The genome-wide comparison reveals the evolutionary generation of diversity in the regulation of transcription.

Amino Acid Motifs↗
PubMed2000

A genomic perspective on plant transcription factors.

Data from the Arabidopsis genome project suggest that more than 5% of the genes of this plant encode transcription factors. The necessity for the use of genomic analytical approaches becomes clear when it is considered that less than 10% of these factors have been genetically characterized. A variety of tools for functional genomic analyses in plants have been developed over the past few years. The availability of the full complement of Arabidopsis transcription factors, together with the results of recent studies that illustrate some of the challenges to their functional characterization, now provides the basic framework for future analyses of transcriptional regulation in plants.

Arabidopsis↗
PubMed2000

Identification of a pathogenicity determinant of Plum pox virus in the sequence encoding the C-terminal region of protein P3+6K(1).

A full-length genomic cDNA clone of a plum pox potyvirus (PPV) isolate belonging to the M strain (PPV-PS) has been cloned downstream from a bacteriophage T7 polymerase promoter and sequenced. Transcripts from the resulting plasmid, pGPPVPS, were infectious and, in herbaceous hosts, produced symptoms that differed from those of virus progeny of pGPPV, a full-length genomic cDNA clone of the D strain PPV-R. Viable PPV-R/-PS chimeric viruses were constructed by recombination of the cDNA clones in vitro. Analysis of plants infected with the different chimeras indicated that sequences encoding the most variable regions of the potyvirus genome, the P1 and capsid protein coding sequences, were not responsible for symptom differences between the two PPV isolates in herbaceous hosts. On the contrary, complex symptomatology determinants seem to be located in the central region of the PPV genome. The results indicate that a genomic fragment that encodes 173 aa from the C-terminal part of the P3+6K(1) coding region is enough to confer, on a PPV-R background, a PS phenotype in Nicotiana clevelandii. This pathogenicity determinant also participates in symptom induction in Pisum sativum, although the region defining the PS phenotype in this host is probably restricted to 74 aa.

Amino Acid Sequence↗
PubMed1999

Non-AUG initiation of AGAMOUS mRNA translation in Arabidopsis thaliana.

The MADS box organ identity gene AGAMOUS (AG) controls several steps during Arabidopsis thaliana flower development. AG cDNA contains an open reading frame that lacks an ATG triplet to function as the translation initiation codon, and the actual amino terminus of the AG protein remains uncharacterized. We have considered the possibility that AG translation can be initiated at a non-AUG codon. Two possible non-AUG initiation codons, CUG and ACG, are present in the 5' region of AG mRNA preceding the highly conserved MADS box sequence. We prepared a series of AG genomic constructs in which these codons are mutated and assayed their activity in phenotypic rescue experiments by introducing them as transgenes into ag mutant plants. Alteration of the CTG codon to render it unsuitable for acting as a translation initiation site does not affect complementation of the ag-3 mutation in transgenic plants. However, a similar mutation of the downstream ACG codon prevents the rescue of the ag-3 mutant phenotype. Conversely, if an ATG is introduced immediately 5' to the disrupted ACG codon, the resulting construct fully complements the ag-3 mutation. The AG protein synthesized in vitro by initiating translation at the ACG position is active in DNA binding and is of the same size as the AG protein detected from floral tissues, whereas AG polypeptides with additional amino-terminal residues do not appear to bind DNA. These results indicate that translation of AG is initiated exclusively at an ACG codon and prove that non-AUG triplets may be efficiently used as the sole translation initiation site in some plant cellular mRNAs.

AGAMOUS Protein, Arabidopsis↗
PubMed1998

The AP2/EREBP family of plant transcription factors.

AP2 (APETALA2) and EREBPs (ethylene-responsive element binding proteins) are the prototypic members of a family of transcription factors unique to plants, whose distinguishing characteristic is that they contain the so-called AP2 DNA-binding domain. AP2/ REBP genes form a large multigene family, and they play a variety of roles throughout the plant life cycle: from being key regulators of several developmental processes, like floral organ identity determination or control of leaf epidermal cell identity, to forming part of the mechanisms used by plants to respond to various types of biotic and environmental stress. The molecular and biochemical characteristics of the AP2/EREBP transcription factors and their diverse functions are reviewed here, and this multigene family is analyzed within the context of the Arabidopsis thaliana genome sequence project.

Amino Acid Sequence↗
PubMed1997

Determination of floral organ identity by Arabidopsis MADS domain homeotic proteins AP1, AP3, PI, and AG is independent of their DNA-binding specificity.

The MADS domain homeotic proteins APETALA1 (AP1), APETALA3 (AP3), PISTILLATA (PI), and AGAMOUS (AG) combinatorially specify the identity of Arabidopsis floral organs. AP1/AP1, AG/AG, and AP3/PI dimers bind to similar CArG box sequences; thus, differences in DNA-binding specificity among these proteins do not seem to be the origin of their distinct organ identity properties. To assess the overall contribution that specific DNA binding could make to their biological specificity, we have generated chimeric genes in which the amino-terminal half of the MADS domain of AP1, AP3, PI, and AG was substituted by the corresponding sequences of human SRF and MEF2A proteins. In vitro DNA-binding assays reveal that the chimeric proteins acquired the respective, and distinct, DNA-binding specificity of SRF or MEF2A. However, ectopic expression of the chimeric genes reproduces the dominant gain-of-function phenotypes exhibited by plants ectopically expressing the corresponding Arabidopsis wild-type genes. In addition, both the SRF and MEF2 chimeric genes can complement the pertinent ap1-1, ap3-3, pi-1, or ag-3 mutations to a degree similar to that of AP1, AP3, PI, and AG when expressed under the control of the same promoter. These results indicate that determination of floral organ identity by the MADS domain homeotic proteins AP1, AP3, PI, and AG is independent of their DNA-binding specificity. In addition, the DNA-binding experiments show that either one of the two MADS domains of a dimer can be sufficient to confer a particular DNA-binding specificity to the complex and that sequences outside the amino-terminal basic region of the MADS domain can, in some cases, contribute to the DNA-binding specificity of the proteins.

AGAMOUS Protein, Arabidopsis↗
PubMed1997

MADS domain proteins in plant development.

The MADS domain (MCM1, AGAMOUS, DEFICIENS, and SRF, serum response factor) is a conserved DNA-binding/dimerization region present in a variety of transcription factors from different kingdoms. MADS box genes represent a large multigene family in vascular plants. In angiosperms, many of the genes of the MADS family are involved in different steps of flower development, most notably in the determination of floral meristem and organ identity. The roles that MADS box genes play, however, are not restricted to control the development of the plant reproductive structures. The genetic, molecular, and biochemical basis of the action of the MADS domain proteins in the plant life cycle are reviewed here.

Amino Acid Sequence↗
PubMed1996

DNA-binding properties of Arabidopsis MADS domain homeotic proteins APETALA1, APETALA3, PISTILLATA and AGAMOUS.

The MADS domain proteins APETALA1 (AP1), APETALA3 (AP3), PISTILLATA (PI), and AGAMOUS (AG) specify the identity of Arabidopsis floral organs. AP1 and AG homocomplexes and AP3-PI heterocomplexes bind to CArG-box sequences. The DNA-binding properties of these complexes were investigated. We find that AP1, AG and AP3-PI are all capable of recognizing the same DNA-binding sites, although with somewhat different affinities. In addition, the three complexes induce similar conformational changes on a CArG-box sequence. Phasing analysis reveals that the induced distortion is DNA bending, oriented toward the minor groove. The molecular dissection of AP1, AP3, PI and AG indicates that the boundaries of the dimerization domains of these proteins vary. The regions required to form a DNA-binding complex include, in addition to the MADS box, the entire L region (which follows the MADS box) and the first putative amphipathic helix of the K box in the case of AP3-PI, while for AP1 and AG only a part of the L region is needed. The similarity of the DNA-binding properties of AP1, AP3-PI and AG is discussed with regard to the biological specificity that these proteins exhibit.

AGAMOUS Protein, Arabidopsis↗
PubMed1996

Dimerization specificity of Arabidopsis MADS domain homeotic proteins APETALA1, APETALA3, PISTILLATA, and AGAMOUS.

The MADS domain homeotic proteins APETALA1 (AP1), APETALA3 (AP3), PISTILLATA (PI), and AGAMOUS (AG) act in a combinatorial manner to specify the identity of Arabidopsis floral organs. The molecular basis for this combinatorial mode of action was investigated. Immunoprecipitation experiments indicate that all four proteins are capable of interacting with each other. However, these proteins exhibit "partner-specificity" for the formation of DNA-binding dimers; only AP1 homodimers, AG homodimers, and AP3/PI heterodimers are capable of binding to CArG-box sequences. Both the AP3/PI heterodimer and the AP1 or AG homodimers are formed when the three corresponding proteins are present together. The use of chimeric proteins formed by domain swapping indicates that the L region (which follows the MADS box) constitutes a key molecular determinant for the selective formation of DNA-binding dimers. The implications of these results for the ABC genetic model of flower development are discussed.

AGAMOUS Protein, Arabidopsis↗
PubMed1995

Processing of the plum pox virus polyprotein at the P3-6K1 junction is not required for virus viability.

Proteolytic processing of the potyvirus polyprotein is mainly performed by the virus-encoded NIa protease, whose cleavage sites are characterized by conserved heptapeptide sequences. Partial processing at the cleavage site present between the P3 and 6K1 cistrons by the plum pox potyvirus (PPV) NIa protease has been previously shown to occur in vitro. We have now studied the role of polyprotein processing at the P3-6K1 junction in vivo, using a full-length PPV cDNA clone. PPV mutant transcripts containing a histidine for glutamine substitution in the cleavage site sequence (a change that abolishes in vitro processability) are able to infect Nicotiana clevelandii plants, indicating that normal processing at the P3-6K1 junction is not required for virus viability. However, disease symptoms were not detected and virus accumulation occurred after a second site mutation was introduced into the 6K1 cistron during replication. This additional change did not restore the in vitro processability of the mutant heptapeptide. Changes at other positions in the heptapeptide (that only slightly altered the in vitro processability of this NIa site) were also engineered and it was found that these mutations affected the time course and severity of the symptom induction process. A possible regulatory effect on the function of the potyvirus P3 + 6K1 protein by processing at the P3-6K1 junction is discussed in light of our present results with PPV.

Animals↗
PubMed1993

Inhibitory effects of human cystatin C on plum pox potyvirus proteases.

The effect of different protease inhibitors on the proteolytic processing of the plum pox potyvirus (PPV) polyprotein has been analyzed. Human cystatin C, an inhibitor of cysteine proteases, interfered with the autoprocessing of the viral papain-like cysteine protease HCPro. Unexpectedly, it also had an inhibitory effect on the autocatalytic cleavage of the Nla protease which, although it has a Cys residue in its active center, has been described as structurally related to serine proteases. Other protease inhibitors tested had no effect on any of the cleavage events analyzed.

Cystatin C↗
PubMed1993

3'-terminal sequence of the plum pox virus PS and ŏ6 isolates: evidence for RNA recombination within the potyvirus group.

The sequence of the 3'-terminal 1768 nucleotides of the PS and ŏ6 isolates of plum pox virus (PPV) has been determined and compared with that of the equivalent regions of other PPV isolates sequenced previously. The sequenced region is part of the PPV open reading frame encoding the last 186 amino acids of the NIb protein and the coat protein (CP, 330 amino acids), followed by a non-coding region of 220 nucleotides and a poly(A) tail. PPV-PS and PPV(-)ŏ6, just like PPV-El Amar, show rather high levels of nucleotide diversity in the sequence encoding the C-terminal region of the NIb protein (19.4 to 31%) and the N terminus of CP (22.8 to 41.1%) when compared with PPV-Rankovic, PPV-D and PPV-NAT, whereas the level of diversity in the rest of the CP sequence and the 3' non-coding region is low (8 to 10.8% and 5.5 to 7.7%, respectively). However, the first 429 sequenced nucleotides of PPV(-)ŏ6 are very similar to those of the PPV-Rankovic, PPV-D and PPV-NAT isolates, whereas the rest of the sequence clearly resembles PPV-PS. Thus, PPV(-)ŏ6 seems to be the result of a natural recombination event between two wild strains of PPV. To our knowledge this is the first evidence of homologous RNA recombination (a process which could play an important role in the evolution of RNA viruses) within the potyvirus group.

Amino Acid Sequence↗
PubMed1992

Proteolytic processing of the plum pox potyvirus polyprotein by the NIa protease at a novel cleavage site.

The expression of potyvirus genomic RNA takes place through translation of its unique long and functional open reading frame into a large polyprotein that undergoes extensive proteolytic processing. Most of the cleavages are performed by the virus-encoded NIa protease, which cuts the polyprotein at defined sites that are characterized by conserved heptapeptide sequences. We have demonstrated in vitro cleavage activity by the plum pox potyvirus (PPV) NIa protease at a novel site, previously identified by sequence analysis, thus allowing a further refinement of the potyviral genetic map. This novel site is located 52 amino acids upstream from the site corresponding to the N-terminus of the CI protein (the NIa cleavage site previously considered the closest to the beginning of the polyprotein). The specificity of the processing was demonstrated by its abolishment when the Gln at position -1 of the cleavage site was changed to His. This novel NIa cleavage site was only partially processed, a characteristic that was not altered when its heptapeptide sequence was modified to become that of the efficiently cleaved NIb-CP junction. On the contrary, substitutions at the nonconserved position +3 had notable effects, positive or negative, on the efficiency of processing. These results show the relevance of sequence and/or conformational context outside the conserved heptapeptide for modulating the cleavage reaction catalyzed by the NIa protease.

Amino Acid Sequence↗
PubMed1991

Identification of the initiation codon of plum pox potyvirus genomic RNA.

The expression of plum pox potyvirus (PPV) genomic RNA takes place through translation of its unique long and functional open reading frame (ORF) into a large polyprotein that undergoes extensive proteolytic processing. In this paper we show that the AUG recognized as the initiation codon of the PPV ORF by in vitro translation systems is the one found at nucleotide position 147, in spite of the presence at position 36 of an in-phase AUG that marks the start of the ORF. Deletion of a substantial part of the PPV 5' nontranslated region (5'-NTR), from nucleotide 19 to 101, does not impair the in vitro translation of PPV synthetic transcripts. By introduction of mutations that disrupt either of these two AUGs into a full-length PPV cDNA clone, it is shown that, while alteration of the first AUG does not have any effect on virus viability, growth, or symptom induction, destruction of the second renders the viral RNA noninfectious. This result indicates that the AUG employed in vivo is also the second. The hypothesis that this AUG could be recognized through a ribosomal internal entry mechanism has been tested in vitro using various bicistronic transcripts in which the PPV 5'-NTR was internally placed. The second cistron of these bicistronic RNAs was translated, but only at low levels, indicating that the PPV 5'-NTR is not able to drive in vitro an efficient internal entry of the ribosomes and suggesting that PPV RNA translation might proceed through a conventional leaky scanning mechanism.

Base Sequence↗
PubMed1991

Novel catalytic activity associated with positive-strand RNA virus infection: nucleic acid-stimulated ATPase activity of the plum pox potyvirus helicaselike protein.

The cylindrical inclusion protein of potyviruses contains the so-called nucleoside triphosphate binding motif, an amino acid sequence motif present in proteins encoded by most positive-strand RNA viruses, some double-strand RNA viruses, apparently all groups of double-strand DNA viruses, and also several single-strand DNA viruses. Further sequence analysis has allowed to include the cylindrical inclusion protein of potyviruses as a member of a superfamily of helicaselike proteins. In this paper we show that the purified cylindrical inclusion protein of plum pox potyvirus interacts with RNA and ATP and copurifies with a nucleic acid-stimulated ATPase activity. To our knowledge, this is the first time that this kind of enzymatic activity has been experimentally associated with a positive-strand RNA virus-encoded protein.

Adenosine Triphosphatases↗