The Arabidopsis PHD-finger protein SHL is required for proper development and fertility.

The SHL gene from Arabidopsis thaliana encodes a small nuclear protein that contains a BAH domain and a PHD finger. Both domains are found in numerous (putative) transcriptional regulators and chromatin-remodeling factors. Different sets of transgenic lines were established to analyze the physiological relevance of SHL. SHL expression driven by the CaMV 35S promoter results in reduced growth, early flowering, early senescence, and impaired flower and seed formation. Antisense inhibition of SHL expression gives rise to dwarfism and delayed development. In-frame N-terminal fusion of the SHL protein to beta-glucuronidase (GUS) directs GUS to the nucleus of stably transformed Arabidopsis plants. Thus, SHL encodes a novel putative regulator of gene expression, which directly or indirectly influences a broad range of developmental processes.

Brassinosteroids rescue the deficiency of CYP90, a cytochrome P450, controlling cell elongation and de-etiolation in Arabidopsis.

The cpd mutation localized by T-DNA tagging on Arabidopsis chromosome 5-14.3 inhibits cell elongation controlled by the ecdysone-like brassinosteroid hormone brassinolide. The cpd mutant displays de-etiolation and derepression of light-induced genes in the dark, as well as dwarfism, male sterility, and activation of stress-regulated genes in the light. The CPD gene encodes a cytochrome P450 (CYP90) sharing homologous domains with steroid hydroxylases. The phenotype of the cpd mutant is restored to wild type both by feeding with C23-hydroxylated brassinolide precursors and by ectopic overexpression of the CPD cDNA. Brassinosteroids also compensate for different cell elongation defects of Arabidopsis det, cop, fus, and axr2 mutants, indicating that these steroids play an essential role in the regulation of plant development.

Blood-flow sensing by anionic biopolymers.

Using 23Na-NMR techniques we could show that the polyanion proteoheparan sulfate integrated into the membrane of endothelial cells may serve as "flow sensor'. Based on its viscoelastic properties, heparan sulfate proteoglycan is present as a random coil under "no flow' conditions, whereby most of its polyanionic sites undergo intramolecular hydrogen bonding. With increasing flow the macromolecule becomes unfolded into a filamentous structure. Additional anionic binding sites to which Na+ ions from the blood bind are released by this shear stress-dependent conformational change. The Na+ binding triggers the signal transduction chain for a vasodilatory vessel reaction. Decrease in flow effects, for reasons of the intramolecular elastic recoil forces of the macromolecules, an entropic coiling, the release of Na+ ions and thus an interruption of the signal chain. Proteoheparan sulfate adsorbed onto a hydrophobic surface in physiological Krebs solution at pH 7.3 demonstrated clearly its characteristic as a Na+ sensor. While Ca2+ ions modulated the adsorption (promotion with increasing Ca2+ concentrations) by changing the conformation of the sensor molecule, the adsorbed amount was determined preferably by the Na+ concentration. K+ and Mg2+ ions showed slightly desorbing properties with increasing concentrations. Thus, it may be concluded that Na+ ions play the role as "first messenger' in flow-dependent vasodilation.

Anionic biopolymers as blood flow sensors.

The finding of flow-dependent vasodilation rests on the basic observation that with an increase in blood flow the vessels become wider, with a decrease the vascular smooth muscle cells contract. Proteoheparan sulphate could be the sensor macromolecule at the endothelial cell membrane-blood interface, that reacts on the shear stress generated by the flowing blood, and that informs and regulates the vascular smooth muscle cells via a signal transduction chain. This anionic biopolyelectrolyte possesses viscoelastic and specific ion binding properties which allow a change of its configuration in dependence on shear stress and electrostatic charge density. The blood flow sensor undergoes a conformational transition from a random coil to an extended filamentous state with increasing flow, whereby Na+ ions from the blood are bound. Owing to the intramolecular elastic recoil forces of proteoheparan sulphate the slowing of a flow rate causes an entropic coiling, the expulsion of Na+ ions and thus an interruption of the signal chain. Under physiological conditions, the conformation and Na+ binding proved to be extremely Ca(2+)-sensitive while K+ and Mg2+ ions play a minor role for the susceptibility of the sensor. Via counterion migration of the bound Na+ ions along the sensor glycosaminoglycan side chains and following Na+ passage through an unspecific ion channel in the endothelial cell membrane, the signal transduction chain leads to a membrane depolarization with Ca2+ influx into the cells. This stimulates the EDRF/NO production and release from the endothelial cells. The consequence is vasodilation.

Ac/Ds transposon mutagenesis in Arabidopsis thaliana: mutant spectrum and frequency of Ds insertion mutants.

Using a two-component Ac/Ds system consisting of a stabilized Ac element (Acc1) and a non-autonomous element (DsA), 650 families of plants carrying independent germinal DsA excisions/transpositions were isolated. Progenies of 559 of these Acc1/DsA families, together with 43 families of plants selected for excision/transposition of wild-type (wt) Ac, were subjected to a broad screening program for mutants exhibiting visible alterations. This resulted in the identification of 48 mutants showing a wide variety of mutant phenotypes, including embryo lethality (24 mutants), chlorophyll defects (5 mutants), defective seedlings (2 mutants), reduced fertility (5 mutants), reduced size (3 mutants), altered leaf morphology (2 mutants), dark green, unexpanded rosette leaves (3 mutants), and aberrant flower or shoot morphology (4 mutants). To whether these mutants were due to transposon insertions, a series of Southern blot experiments was performed on 28 families, comparing in each case several mutant plants with others showing the wild-type phenotype. A preliminary analysis revealed in 4 of the 28 families analyzed a common, novel DsA fragment in all mutant plants, which was present only in heterozygous plants with wt phenotype, as expected for DsA insertion mutations. These four mutants included two showing embryo lethality, one with dark green, unexpanded rosette leaves and stunted inflorescences, and one with curly growth of stems, leaves and siliques. Further evidence for DsA insertion mutations was obtained for one embryo lethal mutant and for the stunted mutant, while in case of the second embryo lethal mutant, the DsA insertion could be separated from the mutant locus by genetic recombination.

Evolution of the NH2- and COOH-terminal extensions of chloroplast ribosomal protein S18. Nucleotide sequence of pea and rye chloroplast rps 18 genes.

An unusual, variably repeated heptapeptide motif is present in most chloroplast ribosomal protein S18 sequences (Weglöhner and Subramanian, FEBS Lett. 269, 193-197, 1991), whereas it is absent in bacterial, cyanelle, and in the chloroplast S18 of the lower plant liverwort. In order to understand the evolution of this higher plant-specific motif, we have cloned and sequenced chloroplast rps18 genes from pea, a dicot plant of the large legume family and rye, a monocot plant with temperature-sensitive chloroplast ribosome formation. The derived amino acid sequence of pea S18 protein shows two and that of rye seven repeats of this motif. We also show that a different heptapeptide motif is discernible in the recently published chloroplast S18 sequence of Pinus thunbergii (a gymnosperm), which can however be derived convergently from a putative progenitor of angiosperm-gymnosperm chloroplast S18. The presence of a 3-fold repeat of an asparagine-rich heptapeptide in the C-terminal extensions of all cereal S18 is also shown here. The results are further discussed in terms of possible origin of these repeats and the ribosomal protein evolution in general.