Ovule-specific MADS-box proteins have conserved protein-protein interactions in monocot and dicot plants

OsMADS13 is a rice MADS-box gene that is specifically expressed in developing ovules. The amino acid sequence of OsMADS13 shows 74% similarity to those of FLORAL BINDING PROTEIN 7 (FBP7) and FBP11, the products of two MADS-box genes that are necessary and sufficient to determine ovule identity in Petunia. To assess whether OsMADS13, the putative rice ortholog of FBP7 and FBP11, has an equivalent function, several analyses were performed. Ectopic expression of FBP7 and FBP11 in Petunia results in ectopic ovule formation on sepals and petals. Here we show that ectopic expression of OsMADS13 in rice and Arabidopsis does not result in the formation of such structures. Furthermore, ectopic expression of FBP7 and FBP11 in Arabidopsis also fails to induce ectopic ovule formation. To determine whether protein-protein interactions involving putative class D MADS-box proteins have been conserved, yeast two-hybrid assays were performed. These experiments resulted in the identification of three putative partners of OsMADS13, all of them encoded by AGL2-like genes. Interestingly the Petunia FBP7 protein also interacts with AGL2-like proteins. The evolutionary conservation of the MADS-box protein partners of these ovule-specific factors was confirmed by exchange experiments which showed that the protein partners of OsMADS13 interact with FBP7 and vice versa.     

A novel and conserved salt-induced protein is an important determinant of salt tolerance in yeast.

We have isolated a novel yeast gene, HAL1, which upon overexpression improves growth under salt stress. In addition, disruption of this gene decreases salt tolerance. Therefore HAL1 constitutes a rate-limiting determinant for halotolerance. It encodes a polar protein of 32 kDa located in the yeast cytoplasm and unrelated to sequences in data banks. The expression of this gene is increased by high concentrations of either NaCl, KCl or sorbitol. On the other hand, the growth advantage obtained by overexpression of HAL1 is specific for NaCl stress. In cells overexpressing HAL1, sodium toxicity seems to be counteracted by an increased accumulation of potassium. The HAL1 protein could interact with the transport systems which determine intracellular K+ homeostasis. The HAL1 gene and encoded protein are conserved in plants, being induced in these organisms by salt stress and abscisic acid. These results suggest that yeast serves as a convenient model system for the molecular biology of plant salt tolerance.     

A promoter from sugarcane bacilliform badnavirus drives transgene expression in banana and other monocot and dicot plants

A 1369 bp DNA fragment (Sc) was isolated from a full-length clone of sugarcane bacilliform badnavirus (ScBV) and was shown to have promoter activity in transient expression assays using monocot (banana, maize, millet and sorghum) and dicot plant species (tobacco, sunflower, canola and Nicotiana benthamiana). This promoter was also tested for stable expression in transgenic banana and tobacco plants. These experiments showed that this promoter could drive high-level expression of the -glucuronidase (GUS) reporter gene in most plant cells. The expression level was comparable to the maize ubiquitin promoter in standardised transient assays in maize. In transgenic banana plants the expression levels were variable for different transgenic lines but was generally comparable with the activities of both the maize ubiquitin promoter and the enhanced cauliflower mosaic virus (CaMV) 35S promoter. The Sc promoter appears to express in a near-constitutive manner in transgenic banana and tobacco plants. The promoter from sugarcane bacilliform virus represents a useful tool for the high-level expression of foreign genes in both monocot and dicot transgenic plants that could be used similarly to the CaMV 35S or maize polyubiquitin promoter.     

Dephosphorylation of photosystem II core proteins is light-regulated in vivo.

A number of photosystem II (PSII)-associated proteins, including D1, D2, CP43 and LHCII, are phosphorylated post-translationally by a membrane-bound, redox-regulated kinase activity. In vitro studies have demonstrated that these proteins can be dephosphorylated by membrane-bound phosphatase activity, reportedly insensitive to light or redox control. We demonstrate here that the PSII core proteins, D1, D2 and CP43, undergo light-stimulated, linear electron-transport-independent dephosphorylation in vivo. The in vivo dephosphorylation of D1 was characterized further and shown to depend upon light intensity, and to occur throughout the visible light spectrum with characteristics most consistent with light absorption by chlorophyll. PSII core protein dephosphorylation in vivo was stimulated by photosystem I (PSI)-specific far-red light, and inhibited by 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone, an inhibitor of plastoquinol oxidation by the cytochrome b6f complex. Based on these findings, we propose that PSI excitation is involved in regulating dephosphorylation of PSII core proteins in vivo.     

Maize rbcS promoter activity depends on sequence elements not found in dicot rbcS promoters.

Although the molecular mechanisms of dicot photosynthetic gene regulation have been pursued actively, comparable studies of monocot regulation have been slow to come forth. We show here that monocot (maize and wheat) but not dicot (pea, tobacco, and Arabidopsis) ribulose-1,5-bisphosphate carboxylase small subunit (rbcS) gene promoters are active in maize mesophyll protoplasts. The evolutionarily conserved GT and G boxes of dicot rbcS promoters are not essential for light-responsive expression in monocot leaf cells. Instead, at least six constitutive and light-sensitive regulatory elements are likely important for maize rbcS expression. Synergism between upstream and downstream promoter elements is required. Whereas in dicots, light triggers coupled leaf development and photosynthetic gene expression, in monocots, light regulation of rbcS is uncoupled from leaf development. Light regulation of maize rbcS may be divided into direct and indirect contributions mediated by different regulatory elements. Because wheat and maize rbcS promoters show sequence homologies and similar expression patterns in monocot and dicot leaf cells, it appears likely that monocots share conserved regulatory elements irrespective of whether they utilize the C3 or C4 pathway for carbon fixation.     

A Y-chromosomal DNA fragment is conserved in human and chimpanzee.

A human male-specific Y-chromosomal DNA fragment (lambda YH2D6) has been isolated. By deletion-mapping analysis, 2D6 has been localized to the euchromatic portion of the long arm (Yq11) of the human Y chromosome. Among great apes, this fragment was found to be conserved in male chimpanzee but was lacking in male gorilla and male orangutan. No homologous fragments were detected in females of orangutan, gorilla, chimpanzee, or human. Nucleotide sequence analysis indicated the presence of partial-Alu-elements and of sequences similar to the GATA repeats of the snake Bkm sequence.     

A novel Arabidopsis gene required for ethanol tolerance is conserved among plants and archaea

A novel ethanol-hypersensitive mutant, gek1, of Arabidopsis shows 10-100 times greater sensitivity to ethanol compared to the wild type, while it grows normally in the absence of ethanol, and responds normally to other alcohols and to environmental stresses such as heat shock and high salinity. Mapping of the gek1 locus indicated it is a previously unreported locus. In order to address the GEK1 function, we identified the GEK1 gene by means of map-based cloning. The GEK1 gene encodes a novel protein without any known functional motifs. Transgenic Arabidopsis plants overexpressing GEK1 displayed an enhanced tolerance to ethanol and acetaldehyde, suggesting that GEK1 is directly involved in the tolerance to those chemicals. By contrast, expression of GEK1 in E. coli and yeasts did not increase their tolerance to ethanol or acetaldehyde. Interestingly, a similarity search revealed that GEK1-related genes are conserved only in plants and archaea. These results might suggest that plants, and presumably archaea, have a novel mechanism for protection from acetaldehyde toxicity.     

A novel calcium/calmodulin-regulated kinesin-like protein is highly conserved between monocots and dicots

Recently, a novel kinesin-like protein (KCBP) that is regulated by Ca2+/calmodulin was isolated from dicot plants. A homolog of KCBP has not been reported in monocots. To determine if this motor protein is present in phylogenetically divergent flowering plants, Arabidopsis KCBP cDNA was used as a probe to screen a genomic library of maize, an evolutionarily divergent species. This screening resulted in isolation of a KCBP homolog. Comparison of the predicted amino acid sequence of the KCBP from maize (ZmKCBP), a monocot, with the previously reported KCBP sequences from dicot species showed that the amino acid sequence, domain organization, and gene structure are highly conserved between monocots and dicots. The C-terminal region of ZmKCBP, containing the motor domain and the calmodulin-binding domain, and the N-terminal tail, with a myosin tail homology region (MyTH4) and talin-like region, showed strong sequence similarity to the KCBP homolog from dicots. However, the coiled-coil region is less conserved between monocots and dicots. The ZmKCBP gene contained 22 exons and 21 introns. The location of 19 of the 21 introns of ZmKCBP is also conserved. The ZmKCBP protein is encoded by a single gene and expressed in all tissues. Affinity-purified antibody to the calmodulin-binding domain of Arabidopsis KCBP detected a protein in both the soluble and the microsomal fractions. The C-terminal region of ZmKCBP, containing the motor and calmodulin-binding domains, bound calmodulin in the presence of calcium and failed to bind in the presence of EGTA. The ZmKCBP, along with other KCBPs from dicots, was grouped into a distinct group in the C-terminal subfamily of kinesin-like proteins. These data suggest that the KCBP is ubiquitous and highly conserved in all flowering plants and the origin of KCBP predated the divergence of monocots and dicots.     

A stationary-phase gene in Saccharomyces cerevisiae is a member of a novel, highly conserved gene family.

The regulation of cellular growth and proliferation in response to environmental cues is critical for development and the maintenance of viability in all organisms. In unicellular organisms, such as the budding yeast Saccharomyces cerevisiae, growth and proliferation are regulated by nutrient availability. We have described changes in the pattern of protein synthesis during the growth of S. cerevisiae cells to stationary phase (E. K. Fuge, E. L. Braun, and M. Werner-Washburne, J. Bacteriol. 176:5802-5813, 1994) and noted a protein, which we designated Snz1p (p35), that shows increased synthesis after entry into stationary phase. We report here the identification of the SNZ1 gene, which encodes this protein. We detected increased SNZ1 mRNA accumulation almost 2 days after glucose exhaustion, significantly later than that of mRNAs encoded by other postexponential genes. SNZ1-related sequences were detected in phylogenetically diverse organisms by sequence comparisons and low-stringency hybridization. Multiple SNZ1-related sequences were detected in some organisms, including S. cerevisiae. Snz1p was found to be among the most evolutionarily conserved proteins currently identified, indicating that we have identified a novel, highly conserved protein involved in growth arrest in S. cerevisiae. The broad phylogenetic distribution, the regulation of the SNZ1 mRNA and protein in S. cerevisiae, and identification of a Snz protein modified during sporulation in the gram-positive bacterium Bacillus subtilis support the hypothesis that Snz proteins are part of an ancient response that occurs during nutrient limitation and growth arrest.