Desiccation Tolerance In Resurrection Plants Research Articles

Lindernia brevidens: a novel desiccation‐tolerant vascular plant, endemic to ancient tropical rainforests

A particular adaptation to survival under limited water availability has been realized in the desiccation-tolerant resurrection plants, which tend to grow in a habitat with seasonal rainfall and long dry periods. One of the best-studied examples is Craterostigma plantagineum. Here we report an unexpected finding: Lindernia brevidens, a close relative of C. plantagineum, exhibits desiccation tolerance, even though it is endemic to the montane rainforests of Tanzania and Kenya, where it never experiences seasonal dry periods. L. brevidens has been found exclusively in two fragments of the ancient Eastern Arc Mountains, which were protected from the devastating Pleistocene droughts by the stable Indian Ocean temperature. Analysis of the microhabitat reveals that L. brevidens is found in the same habitat as hygrophilous plant species, which further indicates that the plant never dries out completely. The objective of this investigation was to address whether C. plantagineum and L. brevidens have desiccation-related pathways in common, or whether L. brevidens has acquired novel pathways. A third, closely related, desiccation-sensitive species, Lindernia subracemosa, has been included for comparison. Mechanisms that confer cellular protection during extreme water loss are well conserved between C. plantagineum and L. brevidens, including the interconversion of 2-octulose to sucrose within the two desiccation-tolerant species. Furthermore, transcriptional control regions of desiccation-related genes belonging to the late embryogenesis abundant (LEA) protein family are also highly conserved. We propose that L. brevidens is a neoendemic species that has retained desiccation tolerance through genome stability, despite tolerance being superfluous to environmental conditions.

The ectopic overexpression of a seed‐specific transcription factor, HaHSFA9, confers tolerance to severe dehydration in vegetative organs

Most plant seeds tolerate desiccation, but vegetative tissues are intolerant to drastic dehydration, except in the case of resurrection plants. Therefore, changes in the regulation of genes normally expressed in seeds are thought to be responsible for the evolutionary origin of desiccation tolerance in resurrection plants. Here, we show that constitutive overexpression of the seed-specific HSFA9 transcription factor from sunflower is sufficient to confer tolerance to severe dehydration, outside of the developing seed context, to vegetative tissues of transgenic tobacco. Whole 3-week-old seedlings could survive severe dehydration. This was quantified as a water loss to 1.96 +/- 0.05% of the initial water content, which corresponds to a water potential of approximately -40 MPa. Survival depended on the water potential, from 40% survival at approximately -20 MPa to 6.5% survival at approximately -40 MPa. Whole-seedling survival was limited by the dehydration sensitivity of the roots. Survival correlated with the ectopic expression of a genetic program involving seed-specific, small heat-shock proteins, but not late embryogenesis abundant proteins. The accumulation of sucrose or raffinose family oligosaccharides was not altered by HSFA9. The observed tolerance was achieved without a reduction of growth and development. Our results strongly support the previously suggested contribution of small heat-shock proteins to the desiccation tolerance of seeds. We provide a successful system for analyzing tolerance to severe dehydration in all vegetative organs of seedlings. We propose that HSFA9 is a potential genetic switch involved in the evolution of tolerance to vegetative desiccation.

Desiccation of the resurrection plant Craterostigma plantagineum induces dynamic changes in protein phosphorylation

Reversible phosphorylation of proteins is an important mechanism by which organisms regulate their reactions to external stimuli. To investigate the involvement of phosphorylation during acquisition of desiccation tolerance, we have analysed dehydration-induced protein phosphorylation in the desiccation tolerant resurrection plant Craterostigma plantagineum. Several dehydration-induced proteins were shown to be transiently phosphorylated during a dehydration and rehydration (RH) cycle. Two abundantly expressed phosphoproteins are the dehydration- and abscisic acid (ABA)-responsive protein CDeT11-24 and the group 2 late embryogenesis abundant (LEA) protein CDeT6-19. Although both proteins accumulate in leaves and roots with similar kinetics in response to dehydration, their phosphorylation patterns differ. Several phosphorylation sites were identified on the CDeT11-24 protein using liquid chromatography-tandem mass spectrometry (LCMS/MS). The coincidence of phosphorylation sites with predicted coiled-coil regions leads to the hypothesis that CDeT11-24 phosphorylations influence the stability of coiled-coil interactions with itself and possibly other proteins.

Transition from Natively Unfolded to Folded State Induced by Desiccation in an Anhydrobiotic Nematode Protein

Late embryogenesis abundant (LEA) proteins are associated with desiccation tolerance in resurrection plants and in plant seeds, and the recent discovery of a dehydration-induced Group 3 LEA-like gene in the nematode Aphelenchus avenae suggests a similar association in anhydrobiotic animals. Despite their importance, little is known about the structure of Group 3 LEA proteins, although computer modeling and secondary structure algorithms predict a largely alpha-helical monomer that forms coiled coil oligomers. We have therefore investigated the structure of the nematode protein, AavLEA1, in the first such analysis of a well characterized Group 3 LEA-like protein. Immunoblotting and subunit cross-linking experiments demonstrate limited oligomerization of AavLEA1, but analytical ultracentrifugation and gel filtration show that the vast majority of the protein is monomeric. Moreover, CD, fluorescence emission, and Fourier transform-infrared spectroscopy indicate an unstructured conformation for the nematode protein. Therefore, in solution, no evidence was found to support structure predictions; instead, AavLEA1 seems to be natively unfolded with a high degree of hydration and low compactness. Such proteins can, however, be induced to fold into more rigid structures by partner molecules or by altered physiological conditions. Because AavLEA1 is associated with desiccation stress, its Fourier transform-infrared spectrum in the dehydrated state was examined. A dramatic but reversible increase in alpha-helix and, possibly, coiled coil formation was observed on drying, indicating that computer predictions of secondary structure may be correct for the solid state. This unusual finding offers the possibility that structural shifts in Group 3 LEA proteins occur on dehydration, perhaps consistent with their role in anhydrobiosis.

A novel abscisic acid- and dehydration-responsive gene family from the resurrection plant Craterostigma plantagineum encodes a plastid-targeted protein with DNA-binding activity.

In the desiccation-tolerant resurrection plant Craterostigma plantagineum Hochst. the chloroplasts undergo major ultrastructural changes during dehydration, which are reversible upon rehydration. Such alterations argue the need for efficient protective/stabilising mechanisms to exist. Here we describe a novel gene family that is rapidly and transiently expressed in response to both dehydration and exogenously applied abscisic acid, mostly in the chloroplast-rich palisade layer on the adaxial side of the leaf. Analysis of the putative coding region suggests that the resulting protein is plastid-targeted. This was confirmed using a chimeric green fluorescent protein (GFP) reporter construct in transgenic tobacco plants - hence the gene family is termed Plastid Targeted Protein ( CpPTP). Fluorescence microscopy also revealed that CpPTP was localised in structures similar to proplastid nucleoids in transgenic tobacco ( Nicotiana tabacum L.) BY-2 cells. The ability of CpPTP to interact with DNA was demonstrated through a DNaseI protection assay. A structure-prediction programme suggests that the mature CpPTP is composed almost entirely of a pattern of hydrophobic and hydrophilic residues that form heptad repeats, which are the hallmarks of a coiled-coil domain. Given the localisation and DNA-binding property of the protein, we propose that CpPTP plays a role during the early stages of dehydration-induced chloroplast remodelling.

The isolation of genes from the resurrection grass Sporobolus stapfianus which are induced during severe drought stress

ABSTRACTA modification of the ‘cold plaque’ screening technique (Hodge et al., Plant Journal1992, 2, 257–260) was used to screen a cDNA library constructed from drought‐stressed leaf tissue of the desiccation tolerant (‘resurrection’) grass Sporobolus stapfianus. This technique allowed a large number of clones representing genes expressed at low abundance to be isolated. An examination of expression profiles revealed that several of these genes are induced in desiccation‐tolerant tissue experiencing severe drought stress. Further characterization indicated that the gene products encoded include an eIF1 protein translation initiation factor and a glycine‐ and proline‐rich protein which have not previously been associated with drought stress. In addition, genes encoding a serine/threonine phosphatase type 2C, a tonoplast‐intrinsic protein (TIP) and an early light‐inducible protein (ELIP) were isolated. A number of these genes are expressed differentially in desiccation‐tolerant and desiccation‐sensitive tissues, suggesting that they may be associated with the desiccation tolerance response of S. stapfianus. The results indicate that there may be unique gene regulation processes occurring during induction of desiccation tolerance in resurrection plants which allow different drought‐responsive genes to be selectively expressed at successive levels of water loss.

New desiccation-tolerant plants, including a grass, in the central highlands of Mexico, accumulate trehalose

A grass endemic to Mexico, Sporobolus atrovirens, was identified for the first time as a desiccation-tolerant resurrection plant. Nine species of desiccation-tolerant vascular plants were found in the highland area of Mexico, including four species of ferns (Cheilanthes and Pellaea species) and three species of Selaginella. Two other grasses collected were known desiccation-tolerant species distributed from South America (Cordoba) to North America (Georgia). The ferns Ch. bonariensis, Ch. integerrima, Ch. myriophylla and P. sagittata are newly reported as desiccation-tolerant plants. The osmoprotectant trehalose which has been recorded as rare in plants was found in air-dry foliage of representative species of widely different taxa (9–291 µM g–1 dry weight). The flora of desiccation-tolerant species in Mexico is discussed in connection with its ability to accumulate trehalose.

Desiccation- and abscisic acid-responsive genes encoding major intrinsic proteins (MIPs) from the resurrection plant Craterostigma plantagineum.

Major intrinsic proteins (MIPs) are a family of channel proteins that are mainly represented by aquaporins in plants. These are divided into TIPs (tonoplast intrinsic proteins) and PIPs (plasma membrane intrinsic proteins) according to their subcellular localization. Homologues to PIPs and TIPs were isolated from the desiccation-tolerant resurrection plant Craterostigma plantagineum by two approaches: firstly, a cDNA library constructed from RNA of dehydrated C. plantagineum leaves was screened with an Arabidopsis thaliana Ath-PIP1b cDNA probe and, secondly, a cDNA library was screened differentially to isolate early drought-induced transcripts. According to sequence homologies the isolated cDNA clones were grouped as follows: Cp-PIPa, Cp-PIPb, Cp-PIPc and Cp-TIP. Cp-PIPa, Cp-PIPc and Cp-TIP transcript accumulation was regulated by dehydration and abscisic acid (ABA). Within the Cp-PIPa group transcripts were regulated either by drought only or by drought and ABA, indicating that ABA-dependent and -independent signal transduction pathways lead to Cp-PIPa expression. Comparison of Cp-PIPa expression in detached leaves and in whole plants suggested the involvement of a signal transmitted in the whole plant in response to drought. Cp-PIPb transcript levels were constitutive in all organs tested. Antibodies raised against a Cp-PIPA protein recognized a polypeptide with an apparent molecular mass of 28 kDa. Using these antibodies it was shown that both Cp-PIPA and Cp-PIPB proteins were localized to the plasma membrane. The role of different members of the MIP group in the dehydration response is discussed.