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Adenosine diphosphate-ribosylation factors (ARFs) are small guanine nucleotide-binding proteins that play an important role in intracellular protein trafficking necessary for undertaking multiple physiological functions in plant growth and developmental processes. However, little is known about the mechanism of ARF functioning at the molecular level, as well as its involvement in abiotic stress tolerance. In this study, we demonstrated the direct involvement of an ARF gene SaARF from a grass halophyte Spartina alterniflora in abiotic stress adaptation for the first time. SaARF, which encodes a protein with predicted molecular mass of 21 kDa, revealed highest identity with ARF of Oryza sativa. The SaARF gene is transcriptionally regulated by salt, drought, cold, and ABA in the leaves and roots of S. alterniflora. Arabidopsis plants overexpressing SaARF showed improved seed germination and survival of seedlings under salinity stress. Similarly, SaARF transgenic Arabidopsis plants were more tolerant to drought stress, compared to wild-type plants, by maintaining chlorophyll synthesis, increasing osmolyte synthesis, and stabilizing membrane integrity. Oxidative damage due to moisture stress in transgenic Arabidopsis was also reduced possibly by activating antioxidant genes, AtSOD1 and AtCAT. Our results suggest that enhanced drought and salinity tolerance conferred by the SaARF gene may be due to its role in mediating multiple abiotic stress tolerance mechanisms.

Background: SUMO (Small Ubiquitin related Modifier) conjugation is a post translational regulatory process found in all eukaryotes, mediated by SUMO activating enzyme, SUMO conjugating enzyme, and SUMO ligase for the attachment of SUMO to its target protein. Although the mechanism for regulation of SUMO conjugation pathway genes under abiotic stress has been studied to certain extent, the role of SUMO conjugating enzyme in improving abiotic stress tolerance to plant is largely unexplored. Here, we have characterized a SUMO conjugating enzyme gene 'SaSce9' from a halophytic grass Spartina alterniflora and investigated its role in imparting abiotic stress tolerance., Results: SaSce9 gene encodes for a polypeptide of 162 amino acids with a molecular weight of ~18 kD and isoelectric point 8.43. Amino acid sequence comparisons of SaSce9 with its orthologs from other plant species showed high degree (~85-93%) of structural conservation among each other. Complementation analysis using yeast SCE mutant, Ubc9, revealed functional conservation of SaSce9 between yeast and S. alterniflora. SaSce9 transcript was inducible by salinity, drought, cold, and exogenously supplied ABA both in leaves and roots of S. alterniflora. Constitutive overexpression of SaSce9 in Arabidopsis through Agrobacterium mediated transformation improved salinity and drought tolerance of Arabidopsis. SaSce9 overexpressing Arabidopsis plants retained more chlorophyll and proline both under salinity and drought stress. SaSce9 transgenic plants accumulated lower levels of reactive oxygen under salinity stress. Expression analysis of stress responsive genes in SaSce9 Arabidopsis plants revealed the increased expression of antioxidant genes, AtSOD and AtCAT, ion antiporter genes, AtNHX1 and AtSOS1, a gene involved in proline biosynthesis, AtP5CS, and a gene involved in ABA dependent signaling pathway, AtRD22., Conclusions: These results highlight the prospect of improving abiotic stress tolerance in plants through genetic engineering of the sumoylation pathway. The study provides evidence that the overexpression of SaSce9 in plant can improve salinity and drought stress tolerance by protecting the plant through scavenging of ROS, accumulation of an osmolyte, proline, and expression of stress responsive genes. In addition, this study demonstrates the potential of the halophyte grass S. alterniflora as a reservoir of abiotic stress related genes for crop improvement.

To understand the tolerance mechanism associated with high temperature stress in a halophyte smooth cordgrass (Spartina alterniflora), a few selected salt-induced genes including transcription factors were studied for their transcript abundance. The mRNA analysis of eight genes and nine transcription factors known to be involved in various salt tolerance mechanisms revealed temporal and tissue-dependent variation in their expression under high temperature stress. Differential response of genes under heat and salt stress (reported earlier) indicated different mode of action in the metabolic pathway in response to different environmental cues, and a few common genes responsive to multiple stresses showed temporal and tissue-dependent variation in their expression. This study demonstrates that S. alterniflora could be a potential source of candidate genes conferring tolerance against high temperature in addition to salt tolerance for crop improvement.

The response of a grass halophyte Spartina alterniflora at early stages of salt stress was investigated through generation and systematic analysis of expressed sequence tags (ESTs) from both leaf and root tissues. Random EST sequencing produced 1,227 quality ESTs, which were clustered into 127 contigs, and 368 were singletons. Of the 495 unigenes, 27% represented genes for stress response. Comparison of the 368 singletons against the Oryza sativa gene index showed that >85% of these genes had similarity with the rice unigenes. Moreover, the phylogenetic analysis of an EST similar to myo-inositol 1-phosphate synthase of Spartina and some selected grasses and halophytes showed closeness of Spartina with maize and rice. Transcript abundance analysis involving eight known genes of various metabolic pathways and nine transcription factor genes showed temporal and tissue-dependent variation in expression under salinity. Reverse northern analysis of a few selected unknown and ribosomal genes exhibited much higher abundance of transcripts in response to salt stress. The results provide evidence that, in addition to several unknown genes discovered in this study, genes involved in ion transport, osmolyte production, and house-keeping functions may play an important role in the primary responses to salt stress in this grass halophyte.

Uniola paniculata, commonly known as sea oats, is a C4 perennial grass capable of stabilizing sand dunes. It is most abundant along the Gulf of Mexico and southeastern Atlantic coastal regions of the United States. The species exhibits low seed set and low rates of germination and seedling emergence, and so extensive clonal reproduction is achieved through production of rhizomes, which may contribute to a decline in genetic diversity. To date, there has been no systematic assessment of genetic variability and population structure in naturally occurring stands in the USA. This study was conducted to assess the genetic relationship and diversity among nineteen U. paniculata accessions representing eight states: Texas, Louisiana, Mississippi, Alabama, Florida, South Carolina, North Carolina, and Virginia, using amplified fragment length polymorphism (AFLP). Twelve AFLP EcoRI + MseI primer combinations generated a wide range of polymorphisms (42-81%) with a mean of 59%. Overall, the sea oats plants exhibited a low range of genetic similarity. Florida accessions, FL-33 and FL-39, were most genetically diverse and the accessions from both Carolinas and Virginia (NC-1, NC-11, SC-15, and VA-53) harbored less genetic variability. Cluster analysis using the UPGMA approach separated U. paniculata plants into four major clusters which were also confirmed by principal coordinate analysis (PCO). Further examination of the different components of genetic variation by analysis of molecular variance (AMOVA) indicated the largest proportion of variability at the state level (47.8%) followed by the variation due to the differences among the genotypes within an accession (34.4%), and the differences among the accessions within a state (17.8%). The relationship between genetic diversity and geographic source of sea oats populations of the United States as revealed through this comprehensive study will be helpful to resource managers and commercial nurseries in identifying suitable plant materials for restoration of new areas without compromising the adaptation and genetic diversity.

Drought is a major abiotic stress factor limiting crop production. Identification of genetic factors involved in plant responses to drought stress will provide a solid foundation to improve drought resistance. Sorghum is well adapted to hot dry environments and regarded as a model for studying drought resistance among the grasses. Significant progress in genome mapping of this crop has also been made. In sorghum, rapid premature leaf death generally occurs when water is limited during the grain filling period. Premature leaf senescence, in turn, leads to charcoal rot, stalk lodging, and significant yield loss. More than 80% of commercial sorghum hybrids in the United States are grown under non-irrigated conditions and although most of them have pre-flowering drought resistance, many do not have any significant post-flowering drought resistance. Stay-green is one form of drought resistance mechanism, which gives sorghum resistance to premature senescence under soil moisture stress during the post-flowering period. Quantitative trait locus (QTL) studies with recombinant inbred lines (RILs) and near-isogenic lines (NILs) identified several genomic regions associated with resistance to pre-flowering and post-flowering drought stress. We have identified four genomic regions associated with the stay-green trait using a RIL population developed from B35 x Tx7000. These four major stay-green QTLs were consistently identified in all field trials and accounted for 53.5% of the phenotypic variance. We review the progress in mapping stay-green QTLs as a component of drought resistance in sorghum. The molecular genetic dissection of the QTLs affecting stay-green will provide further opportunities to elucidate the underlying physiological mechanisms involved in drought resistance in sorghum and other grasses.

Several molecular maps have been constructed in sorghum (Sorghum bicolor L. Moench) using a variety of probes from different grass species such as sorghum, maize, sugarcane, rice, oat, and barley. In order to enhance the utility of the existing mapping information by the sorghum research community, alignment and integration of all major molecular maps is necessary. To achieve this objective, a genetic map of 214 loci with a total map distance of 1200 cM was constructed using 98 F7 sorghum recombinant inbred lines (RILs) from a cross between two inbred lines, B35 and Tx7000. Few cDNA clones of sorghum and maize related to photosynthesis and drought stress were mapped on this map for the first time. Five major restriction fragment length polymorphism (RFLP) maps independently developed in this species were used for alignment purpose. The distributions of previously mapped markers were compared with their respective sorghum maps to align each of the linkage groups. In general, consistent linear order among markers was maintained in all the linkage maps. The successful alignment of these RFLP maps will now allow selection of a large number of markers for any region of the sorghum genome with many potential applications ranging from fine mapping and marker-assisted selection to map-based cloning for the improvement of sorghum and related species.