Plant genes and their expression

The transcription of protein encoding genes and their regulation in plant cells shows the signature of a typical eukaryotic system. The major outcome of research in this area is recognition of variability in components assembled to design constitutive or inducible promoters, including those constituting enhancers and boundary elements. Such cis-acting elements respond to diverse intrinsic and extrinsic signals to take care of the needs of sessile plants. While much remains to be defined about the basic transcription machinery in plants, it is expected to follow the pattern as deciphered in animals. At the same time, a large number of genes for regulatory proteins have been cloned leading to information about DNA binding domains, activation domains, nuclear localization signals and oligomerization sites. However, most of this information needs to be tested by experimentation. Availability of transgenic systems in plants and biotechnological needs have given impetus to research for designing tailor-made inducible/ repressible promoters and to manipulate plant processes by deploying transcription factors. While considerable progress has been made, much remains to be learned about the biochemical and molecular basis of regulation of plant gene expression

OsIAA1, an Aux/IAA cDNA from rice, and changes in its expression as influenced by auxin and light

The Aux/IAA class of genes are rapidly induced by exogenous auxins and have been characterized extensively from many dicot species like Arabidopsis, Glycine max and Pisum sativum. We report here the isolation and characterization of rice (Oryza sativa L. subsp. Indica) OsIAA1 cDNA as a monocot member of the Aux/IAA gene family. The predicted amino acid sequence of OsIAA1 corresponds to a protein of ca. 26 kDa, which harbors all four characteristic domains known to be conserved in Aux/IAA proteins. The conservation of these Aux/IAA genes indicates that auxins have essentially a similar mode of action in monocots and dicots. Northern blot analysis revealed that the OsIAA1 transcript levels decrease in the excised coleoptile segments on auxin starvation, and the level is restored when auxin is supplemented; the increase in OsIAA1 transcript level was apparent within 15 to 30 min of auxin application. Auxin-induced OsIAA1 expression appears to be correlated with the elongation of excised coleoptile segments. In light-grown rice seedlings, OsIAA1 is preferentially expressed in roots and basal segment of the seedling, whereas in the etiolated rice seedlings, the OsIAA1 transcripts are most abundant in the coleoptile. A comparative analysis in light- and dark-grown seedling tissues indicates that the OsIAA1 transcript levels decrease on illumination

Sodium chloride resistant cell line from haploid Datura innoxia mill. A resistance trait carried from cell to plantlet and vice versa in Vitro

A cell line resistant to sodium chloride was selected from callus cultures of haploid Datura innoxia by cloning under selective pressure. Cells of the resistant cell line retained their resistance even after subculture in absence of NaCl. Plantlets could be regenerated from resistant cells in the presence as well as absence of NaCl. In contrast, regeneration of plantlets was not possible from normal cells in the presence of NaCl, although regeneration readily occurred in the absence of NaCl. To examine the stability of the resistance in the long-term, callus cultures were initiated in presence of NaCl from stem expiants of the differentiated plantlets. All expiants of plantlets derived from resistant cells showed callus formation. This callus, derived from resistant explants, retained the trait of resistance upon subculture

Molecular characterization and differential expression of cytokinin-responsive type-A response regulators in rice (Oryza sativa)

Background The response regulators represent the elements of bacterial two-component system and have been characterized from dicot plants like Arabidopsis but little information is available on the monocots, including the cereal crops. The aim of this study was to characterize type-A response regulator genes from rice, and to investigate their expression in various organs as well as in response to different hormones, including cytokinin, and environmental stimuli. Results By analysis of the whole genome sequence of rice, we have identified ten genes encoding type-A response regulators based upon their high sequence identity within the receiver domain. The exon-intron organization, intron-phasing as well as chromosomal location of all the RT-PCR amplified rice (Oryza sativa) response regulator (OsRR) genes have been analyzed. The transcripts of OsRR genes could be detected by real-time PCR in all organs of the light- and dark-grown rice seedlings/plants, although there were quantitative differences. The steady-state transcript levels of most of the OsRR genes increased rapidly (within 15 min) on exogenous cytokinin application even in the presence of cycloheximide. Moreover, the expression of the OsRR6 gene was enhanced in rice seedlings exposed to salinity, dehydration and low temperature stress. Conclusion Ten type-A response regulator genes identified in rice, the model monocot plant, show overlapping/differential expression patterns in various organs and in response to light. The induction of OsRR genes by cytokinin even in the absence of de novo protein synthesis qualifies them to be primary cytokinin response genes. The induction of OsRR6 in response to different environmental stimuli indicates its role in cross-talk between abiotic stress and cytokinin signaling. These results provide a foundation for further investigations on specific as well as overlapping cellular functions of type-A response regulators in rice

High frequency production of embryos in Datura innoxia from isolated pollen grains by combined cold treatment and serial culture of anthers in liquid medium

This study concerns the development of pollen embryos as affected by various physical conditions of culture in media devoid of hormones. Freshly isolated pollen, from anthers of Datura, failed to form embryos regardless of whether they were cultured on liquid or solid medium. In contrast, pollen isolated from anthers precultured on solid medium did form embryos and the response could be increased by prior cold treatment of anthers at 4 °C for 4 days. However, the best results were obtained when anthers were cultured from the very beginning in liquid medium and transferred serially to fresh medium. Under such conditions, the anthers dehisced, allowing spontaneous shedding of pollen grains. It was thus possible to have several fractions of shed pollen continuing their development into embryos. When serial culture was started with anthers from cold-treated buds not only were embryos formed in all the fractions of shed pollen but the frequency was also considerably higher than in any mode of culturing

Involvement of G-proteins, calmodulin and tagetitoxin-sensitive RNA polymerase in light-regulated expression of plastid genes (psbA, psaA and rbcL) in rice (Oryza sativa L.)

The regulation of chloroplast gene expression by light involves multiple signaling components. In an earlier study, we demonstrated the role of calcium and phosphorylation in regulating the expression of photosynthesis-related plastid genes, psbA, psaA and rbcL, using rice as a model monocot system. This work has been extended further to examine the possible involvement of heterotrimeric GTP-binding proteins and calmodulin. Vacuum infiltration of 5-day-old etiolated rice seedlings with G-protein agonists, cholera toxin and GTPγS, increased the steady-state transcript levels of the plastid genes. The antagonists/inhibitors of calmodulin action, trifluoperazine and W7, inhibited the light-induced increase in steady-state transcript levels of these genes. The light-regulated expression of photosynthetic genes was also adversely affected by tagetitoxin, a specific inhibitor of plastid-encoded RNA polymerase. These results indicate the involvement of various signaling components in transduction of light signal that probably also recruits PEP to regulate plastid gene expression

Genome-wide identification, classification, evolutionary expansion and expression analyses of homeobox genes in rice

Homeobox genes play a critical role in regulating various aspects of plant growth and development. In the present study, we identified a total of 107 homeobox genes in the rice genome and grouped them into ten distinct subfamilies based upon their domain composition and phylogenetic analysis. A significantly large number of homeobox genes are located in the duplicated segments of the rice genome, which suggests that the expansion of homeobox gene family, in large part, might have occurred due to segmental duplications in rice. Furthermore, microarray analysis was performed to elucidate the expression profiles of these genes in different tissues and during various stages of vegetative and reproductive development. Several genes with predominant expression during various stages of panicle and seed development were identified. At least 37 homeobox genes were found to be differentially expressed significantly (more than two-fold; P <0.05) under various abiotic stress conditions. The results of the study suggest a critical role of homeobox genes in reproductive development and abiotic stress signaling in rice, and will facilitate the selection of candidate genes of agronomic importance for functional validation

Developmental and light-dependent cues interact to establish steady-state levels of transcripts for photosynthesis-related genes (psbA, pbsD, psaA and rbcL) in rice (Oryza sativa L.)

The steady-state transcript levels for psbA, psbD, psaA and rbcL are low in dark-grown rice seedlings as compared to those grown in light. Following seed germination, they accumulate in an age-dependent manner, in dark as well as light, reaching a maximal level on the 7th or 8th day, before a slow decline sets in. But transcripts for psbA and psbD continue to maintain relatively-high levels even after 10 days of growth in light. Exposure of 5-day-old dark-grown seedlings to light results in an approximately 25-60-fold increase in transcripts during a period of 72 h, followed by a decrease. An analysis of data from both lines of investigation reveals that the developmental programme increases the transcript levels for psbA, psbD, psaA and rbcL by about 10-, 2.3-, 7.0- and 8.0-fold, respectively, between 5-8 days after germination and it is independent of light. At the same time, exposure of the seedlings to light during this period further enhances transcript levels by 5-, 11.4-, 6.6- and 7.8-fold, respectively. Thus, both developmental and light-dependent cues contribute to establish steady-state levels of transcripts for the chloroplast genes investigated

Overexpression of putative topoisomerase 6 genes from rice confers stress tolerance in transgenic Arabidopsis plants

DNA topoisomerase 6 (TOP6) belongs to a novel family of type II DNA topoisomerases present, other than in archaebacteria, only in plants. Here we report the isolation of full-length cDNAs encoding putative TOP6 subunits A and B from rice (Oryza sativa ssp. indica), preserving all the structural domains conserved among archaebacterial TOP6 homologs and eukaryotic meiotic recombination factor SPO11. OsTOP6A1 was predominantly expressed in prepollinated flowers. The transcript abundance of OsTOP6A2, OsTOP6A3 and OsTOP6B was also higher in prepollinated flowers and callus. The expression of OsTOP6A2, OsTOP6A3 and OsTOP6B was differentially regulated by the plant hormones, auxin, cytokinin, and abscisic acid. Yeast two-hybrid analysis revealed that the full-length OsTOP6B protein interacts with both OsTOP6A2 and OsTOP6A3, but not with OsTOP6A1. The nuclear localization of OsTOP6A3 and OsTOP6B was established by the transient expression of their β-glucuronidase fusion proteins in onion epidermal cells. Overexpression of OsTOP6A3 and OsTOP6B in transgenic Arabidopsis plants conferred reduced sensitivity to the stress hormone, abscisic acid, and tolerance to high salinity and dehydration. Moreover, the stress tolerance coincided with enhanced induction of many stress-responsive genes in transgenic Arabidopsis plants. In addition, microarray analysis revealed that a large number of genes are expressed differentially in transgenic plants. Taken together, our results demonstrate that TOP6 genes play a crucial role in stress adaptation of plants by altering gene expression

The psbO gene for 33-kDa precursor polypeptide of the oxygen-evolving complex in Arabidopsis thaliana - nucleotide sequence and control of its expression

The 33-kDa polypeptide of the oxygen-evolving complex of photosystem II is nuclear-encoded. The single psbO gene of Arabidopsis thaliana, as suggested by Southern hybridization, has been isolated from the genomic library and sequenced. The sequence analysis has revealed that the psbO gene harbors two introns and encodes a precursor polypeptide of 332 amino acid residues; the first 85 amino acid residues represent the transit peptide and the following 247 amino acids constitute the mature polypeptide. The hydrophilic nature of the 33-kDa protein is confirmed by the presence of 27% charged residues. Northern analysis of the total RNA from Arabidopsis indicates that a 1.2-kb transcript represents the psbO gene. It is expressed in a tissue-specific manner - the steady-state transcript levels being highest in the leaves and virtually undetectable in the roots. Also, expression of the psbO gene is development-dependent and regulated by light in young Arabidopsis seedlings. In a constitutively photomorphogenic mutant of Arabidopsis, pho2 (plumular hook open 2), the psbO gene is de-repressed in young, dark-grown seedlings, resulting in increased transcript abundance compared to the wild-type. These studies, thus, define the influence of at least one regulatory component for psbO expression